Name: a procedure to observe context-induced renewal of pavlovian-conditioned alcohol-seeking behavior in rats
Uploaded: 2014-09-19T14:00:00.000+00:00
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<p class='jove_content'>This work was supported by the National Institute on Alcohol Abuse and Alcoholism (RO1 AA014925 awarded to Dr. Patricia H. Janak at University of California, San Francisco, CA, USA), Fonds de recherche du Quebec &#8211; Sant&#233; (NC) and the Natural Sciences and Engineering Research Council (NC, 387224-2010). The open-access publication of this article was made possible by sponsorship from Med Associates Inc. </p>

<p class="jove_content"> Tutte le procedure sono approvate dal Comitato di Ricerca Animal Ethics alla Concordia University e concordano con le raccomandazioni del Consiglio canadese Animal Care. </p><p class="jove_title"> 1. Gli animali </p><ol><li> Ottenere ratti maschio lungo-Evans (220 - 240 g all&#39;arrivo). </li><li> Mantenere i ratti in una temperatura (21 ° C) e umidità (44%), controllata di cura degli animali, su un ciclo luce-buio 12 ore con le luci accese alle ore 7:00. </li><li> Pair-house ratti all&#39;arrivo. Dopo 3 giorni, i ratti casa singolarmente in gabbie scatola da scarpe di plastica con biancheria da letto di beta-chip, un osso di nylon, e l&#39;accesso gratuito al ratto chow standard ed acqua. Utilizzare coperchi gabbia che hanno doppi occhielli per consentire la successiva disposizione di una seconda bottiglia, oltre alla borraccia. NOTA: I ratti possono essere alloggiati individualmente, non appena arrivano nella struttura di cura degli animali. </li><li> Inizia a gestire ratti ogni giorno. NOTA: Come guida, una settimana di trattamento è di solito sufficiente per loro ambientarsi ad essere picked e gestito dallo sperimentatore. </li></ol><p class="jove_title"> 2. intermittente etanolo Access nella casa Cage </p><p class="jove_content"> NOTA: Comportamento intermittente accesso etanolo nella gabbia casa per garantire che i ratti saranno bere quantità fisiologicamente rilevanti di etanolo prima della fase di formazione comportamentale dell&#39;esperimento inizia <sup>17-19.</sup> Avviare la procedura durante la fase di luce, anche se potrebbe essere iniziato in qualsiasi momento durante il ciclo di chiaro o scuro. </p><ol><li> Preparare due bottiglie per ogni ratto. <ol><li> Utilizzare una plastica da 100 ml cilindro graduato come la bottiglia di etanolo e una bottiglia d&#39;acqua di plastica standard di 473 ml. Utilizzare una relativamente piccola bottiglia di capacità etanolo, riempita quasi a capacità massima di volume, per ridurre al minimo la perdita di etanolo a causa di perdite ed evaporazione. </li><li> Riempire una bottiglia con il 15% di etanolo, fatta diluendo etanolo al 95% in acqua di rubinetto. NOTA: altre concentrazioni di etanolo, come ad esempio il 10% o il 20%, può essere utilizzato al posto del 15% di etanolo. </li><li> Fill la seconda bottiglia con acqua del rubinetto. </li><li> Inserire un tappo di gomma, con un tubo di aspirazione che contiene un cuscinetto a sfere per ridurre al minimo le perdite, in ogni bottiglia. </li></ol></li><li> Impostare due gabbie di controllo, trattati in modo identico alle gabbie a casa con l&#39;eccezione che le gabbie di controllo non contengono ratti o biancheria da letto. <ol><li> Preparare bottiglie di etanolo e acqua per le gabbie di controllo come specificato nei punti 2.1.1 - 2.1.4. </li><li> Utilizzare le misure ottenute dalle gabbie di controllo per controllare per spandimenti e / o evaporazione che possono verificarsi nel corso della sessione di accesso etanolo (come descritto ai punti 2.8.1 - 2.8.2). </li></ol></li><li> Pesare tutte le bottiglie di etanolo e di acqua. Registrare tutti i pesi. </li><li> Pesare tutti i topi. Registrare tutti i pesi. </li><li> Inserire la bottiglia etanolo e la bottiglia d&#39;acqua su ogni gabbia casa e controllare gabbia allo stesso tempo. Lasciare entrambe le bottiglie sulle gabbie per 24 ore. </li><li> Alla fine del periodo di 24 ore, rimuovere entrambe le bottiglie dallagabbie allo stesso tempo. </li><li> Pesare tutte le bottiglie di etanolo e di acqua. Registrare tutti i pesi. </li><li> Per determinare la quantità di etanolo e acqua è stata consumata durante il periodo di 24 ore, sottrarre il peso di ogni bottiglia quando è stato tolto dalla gabbia casa dal peso della bottiglia quando è stato posto sulla gabbia a casa 24 ore prima per ogni ratto. <ol><li> Fare lo stesso per le due gabbie di controllo, e quindi calcolare l&#39;importo medio di versamento per ciascun fluido. </li><li> Sottrarre la quantità media di etanolo e di fuoriuscita di acqua, rispettivamente dalle corrispondenti misure di etanolo e di consumo di acqua per ogni ratto. </li><li> Utilizzare queste misure di differenza fuoriuscita controllata per calcolare il consumo di etanolo in g / kg, consumo di acqua, etanolo e la preferenza. </li></ol></li><li> Per calcolare il consumo di etanolo (g / kg), prima convertire il peso (g) di soluzione di etanolo consumato in volume (ml) in base alla densità della soluzione totale (componenti etanolo + acqua), secondoal volume formula (ml) = Peso (g) / Densità (g / ml). NOTA: La densità della soluzione varierà in funzione della concentrazione di etanolo. <ol><li> Successivamente, moltiplicare il volume (ml) di soluzione di etanolo consumata dalla densità (g / ml) di etanolo in scala per la soluzione percentuale specifica utilizzata, a dare il peso (g) di etanolo consumato. </li><li> Dividere il peso (g) di etanolo consumata dal peso di ratto (kg) per ottenere la g / kg di misura del consumo di etanolo. </li></ol></li><li> Per calcolare etanolo preferenza, esprimere assunzione di etanolo (ml) in percentuale del totale di liquidi (acqua + etanolo, ml) di aspirazione. </li><li> Riportare il flacone solo acqua torna alla home gabbia del ratto. Per il prossimo 24 ore, dare la ratti accesso all&#39;acqua, ma non etanolo. Invece di una bottiglia di etanolo, posizionare una seconda bottiglia di pre-pesato acqua sulla gabbia di casa. </li><li> Alla fine del periodo di 24 ore, rimuovere entrambi bottiglie di acqua dalla gabbia allo stesso tempo. </li><li> Pesare entrambe le serie di bottiglie d&#39;acqua. Registrare tutti weicombatte. </li><li> Sottrarre il peso di ogni bottiglia quando è stato tolto dalla gabbia casa dal peso della bottiglia quando è stato posto sulla gabbia a casa 24 ore prima, sottraendo la misura media fuoriuscita ottenuto da gabbie di controllo come descritto ai punti 2.8.1 - 2.8.2. </li><li> Utilizzare queste misure differenza fuoriuscita controllata per calcolare il consumo di acqua. </li><li> Inizia il ciclo di nuovo, fornendo sia l&#39;etanolo e bottiglie d&#39;acqua ai ratti, come descritto nei punti da 2.1 a 2.14. <ol><li> Ripetere questo ciclo per più (circa 12) sedute di indurre livelli elevati e stabili di consumo di etanolo. Vedere <strong>la Figura 1</strong> per risultati rappresentativi di consumo di etanolo (g / kg) nella gabbia casa. </li><li> Alternate la posizione spaziale della bottiglia etanolo e acqua sul coperchio della gabbia tra le sessioni per prevenire lo sviluppo di una preferenza laterale. </li><li> Fare una soluzione di etanolo fresco una volta alla settimana per garantire la concentrazione accurata e stabile etanolo attraversosessioni. Ridurre al minimo l&#39;impatto di evaporazione di etanolo per periodi prolungati di tempo che utilizzano questo approccio. </li></ol></li></ol><p class="jove_title"> 3 Apparecchiatura </p><ol><li> Casa ciascuna camera di condizionamento all&#39;interno di un ventilato,-attenuazione del suono box. </li><li> Costruire ciascuna camera di condizionamento di un metallo in acciaio inox asta pavimento, un soffitto in policarbonato trasparente, in policarbonato trasparente parete posteriore e porta d&#39;ingresso, e fianchi in alluminio del pannello di metallo. <ol><li> Installare una porta fluido centrale, vicino alla base della parete laterale destra metallo. Dotare l&#39;ingresso al porto fluido con una fotocellula a raggi infrarossi per rilevare le entrate nella porta. </li><li> Collegare tubo di polietilene dal retro della porta fluido a una siringa da 20 ml contenente etanolo utilizzando un ago a punta smussata. Posizionare la siringa in una pompa a siringa situata al di fuori del box-attenuazione del suono. Per le sessioni di allenamento discriminazione di Pavlov, caricare etanolo nella siringa prima di metterlo nella pompa. Prime il tubo by avanzare manualmente la pompa, e poi asciugare l&#39;eccesso di etanolo dal porto fluido. </li><li> Sul fianco sinistro, installare un houselight bianca centrale, vicino al soffitto della camera. </li><li> Installare un clicker (2 ciclo Hz) e un generatore di rumore bianco con un altoparlante nell&#39;angolo superiore sinistro del fianco sinistro. </li></ol></li><li> Dotare ogni camera di condizionamento, con una specifica configurazione di visive, olfattive, tattili e stimoli per creare un contesto caratteristico. <ol><li> Utilizzare due contesti diversi, designato Contesto 1 e Context 2 Configurare la metà delle camere condizionata Contesto 1 e l&#39;altra metà come Context 2. </li><li> Per creare Contesto 1, posizionare pannelli neri di cartone sopra il soffitto e pareti anteriore e posteriore della camera, odore spruzzo di limone (10% v / v, fatta diluendo olio di limone in acqua di rubinetto) in una capsula di Petri e posizionarlo al centro di un vassoio di rifiuti metallici sotto il pavimento della camera e inserire un chiaro pannello di pavimento in policarbonato sopra il metallo rod pavimento. Foderare il contenitore scorie con tovagliolo di carta assorbente o da banco liner. </li><li> Per creare Contesto 2, lasciare il soffitto e pareti anteriore e posteriore della camera di scoperta, in modo che siano chiare, spruzzo odore di mandorle (10% v / v, fatta diluendo benzaldeide in acqua di rubinetto) in una capsula di Petri e posizionarlo centralmente in un contenitore scorie di metallo sotto il pavimento della camera e inserire un pannello perforato pavimento di metallo sul pavimento asta di metallo. Foderare il contenitore scorie con tovagliolo di carta assorbente o da banco liner. </li></ol></li></ol><p class="jove_title"> 4. assuefazione </p><p class="jove_content"> Abituare i ratti alla sala formazione comportamentale e ciascuno dei due contesti all&#39;interno condizionamento camere. Per questa fase dell&#39;esperimento, non caricare l&#39;etanolo nelle siringhe da 20 ml. </p><ol><li> Abituare i ratti al processo di essere trasportati dalla struttura di cura degli animali alla sala formazione comportamentale in laboratorio in due sessioni assuefazione brevi. <ol><li> Nella prima sessione, caricare i ratti in loro gabbie di casa su un carrello a ruote nello stabilimento di cura degli animali, e li trasportano al pronto formazione comportamentale in laboratorio. Illuminare le luci della stanza, accendere i ventilatori in ogni suono attenuazione cubicolo, uscite dalla stanza, chiudere la porta e lasciare che i ratti adattarsi a questo nuovo ambiente per 20 minuti, pur rimanendo nelle loro gabbie a casa sul carrello. Al termine di questo periodo di 20 minuti, restituire i ratti alla funzione di cura degli animali. </li><li> Nella seconda sessione, portare i topi alla sala formazione comportamentale in laboratorio. Come prima, lasciare le luci e ventilatori cubicolo nella stanza. Rimanere nella stanza, con la porta chiusa per ridurre al minimo le distrazioni, e la maniglia e pesare ogni ratto nella sala formazione comportamentale. Registrare i pesi, e quindi restituire i ratti alla struttura di cura degli animali. </li></ol></li><li> Abituare i ratti alle camere di condizionamento in due sessioni assuefazione brevi. <ol><li> Trasportare i ratti alla sala formazione comportamentale, registrare il peso di ciascun ratto,e caricare ogni ratto nella sua camera di condizionamento designato. </li><li> Nella prima sessione di assuefazione impostare la metà delle camere come Context 1 e il resto come Context 2. </li><li> Nella seconda sessione di assuefazione configurare ogni camera con il contesto che non è stato utilizzato nella sessione precedente. </li><li> Condurre sessioni di assuefazione in modo controbilanciato, in modo tale che la metà dei ratti vengono prima esposto a Context 1, seguito da Context 2, e l&#39;altra metà vengono prima esposti a Context 2, seguito da Context 1 Idealmente, condurre entrambe le sessioni di assuefazione nel stesso giorno, anche se potevano essere distribuite in due giorni, se necessario. </li><li> Per le sessioni di assuefazione, utilizzare un programma per computer per illuminare il houselight 1 min dopo che il programma è stato rilasciato. Annotare il numero totale di voci di porta fluido effettuate durante una sessione di 20 min. Al termine della sessione, programmare la houselight per spegnere. Non presenta alcun etanolo o segnali durante le sessioni di assuefazione. </li></ol></li&gt; <li> Rimuovere i ratti dalle camere, li tornare alle loro gabbie a casa, e trasportarli di nuovo alla funzione di cura degli animali. </li><li> Pulire le attrezzature <em>(ad esempio,</em> pannelli a pavimento, vassoi rifiuti, piastre di Petri). Vedere il punto 5.7 per ulteriori dettagli. </li></ol><p class="jove_title"> 5. pavloviano Discriminazione Training (PDT) </p><p class="jove_content"> Treno ratti per comportamentale discriminano tra un cue uditivo che è accoppiato con etanolo e un secondo spunto uditivo che non è. Per questa fase dell&#39;esperimento, riempire le siringhe da 20 ml con 15% di etanolo. </p><ol><li> Dotare ogni camera di condizionamento, con una specifica configurazione di visiva, olfattiva, e stimoli tattili per creare un contesto caratteristico, come descritto ai punti 3.3.2 e 3.3.3 <ol><li> Assegnare la metà dei ratti a Contesto 1 e l&#39;altra metà a Contesto 2 in modo controbilanciato, in base al consumo di etanolo e di preferenza in media attraverso le ultime 2 sessioni della procedura di accesso etanolo gabbia a casa. Fare riferimento alla context usato durante la PDT come Contesto A. NOTA: Ogni ratto deve solo provare un contesto in questa fase di formazione. </li><li> Assicurarsi che lo stimolo olfattivo è l&#39;ultimo stimolo contestuale da aggiungere ad ogni camera di condizionamento. Chiudere le porte della camera e la cabina-attenuazione del suono per contenere lo stimolo olfattivo e ridurre al minimo la comingling di odori diversi. </li></ol></li><li> Portare i topi nella stanza formazione comportamentale, registrare il peso di ciascun ratto, e caricare ogni ratto nella sua camera di condizionamento designato. </li><li> Avviare il programma per computer che controlla la seguente sequenza di eventi durante ogni sessione di PDT. <ol><li> A 5 minuti dopo che il programma è stato rilasciato, illuminare il houselight. </li><li> Presentare lo stimolo rumore bianco e il clicker stimolo 16 volte ciascuno. Designa uno di questi stimoli uditivi essere il CS + e l&#39;altro per essere il CS, in modo controbilanciato attraverso entrambi i contesti. </li><li> Rendere ogni CS + e CS-prova last 10 sec. </li><li> Consegnare 0,2 ml di etanolo nella porta del fluido durante l&#39;ultimo 6 sec del CS +. Non fornire alcun etanolo durante il CS. </li><li> Rendere ogni sessione PDT ultimi 58 minuti in media. Presentare il CS + e CS-ciascuno su una variabile tempo 67 sec programma. </li><li> Record voci fluido porto durante la 10 sec intervallo CS (10 sec durante la presentazione CS), il 10 sec intervallo pre-CS (10 sec immediatamente prima dell&#39;inizio CS), e il 10 sec intervallo post-CS (10 sec immediatamente seguente Offset CS) sia per CS + e CS-presentazioni, così come totale di voci di porta fluido per l&#39;intera sessione. Registrare anche la durata di ogni voce porta fatta durante questi intervalli. </li><li> Al termine della sessione, ruotare la houselight off. </li></ol></li><li> Rimuovere i ratti dalle camere, li tornare alle loro gabbie a casa, e trasportarli di nuovo alla funzione di cura degli animali. </li><li> Controllare le porte del fluido per garantire che i ratti hanno consumato i 3,2 ml di etanolo consegnati in ogni PDT sESSIONE. </li><li> Lavare l&#39;etanolo fuori dal tubo di polietilene con una siringa da 60 ml contenente una piccola quantità di acqua. Quindi, passare 60 ml di aria attraverso il tubo utilizzando una seconda, siringa vuota, per assicurarsi che sia asciutto. </li><li> Pulire l&#39;apparecchiatura. Per la pulizia quotidiana, lavare i pannelli di pavimento, vassoi rifiuti, e piastre di Petri. Una volta alla settimana, pulire l&#39;intera camera di condizionamento. Non utilizzare prodotti di pulizia profumati per evitare l&#39;introduzione di odori indesiderati nella apparato sperimentale. </li><li> Continuare sedute di PDT tutti i giorni fino ratti mostrano discriminazione stabili nelle loro risposte ingresso porta al CS + vs CS. NOTA: le voci della porta fatte in risposta al CS + dovrebbero aumentare nel corso sessioni, mentre voci di porta fatte in risposta al CS dovrebbero rimanere bassi. Il numero esatto di sedute di PDT necessario per conseguire discriminati condizionata risposta può variare, ma, come una guida, intenzione di circa 20 sessioni. </li></ol><p class="jove_title"> 6. Estinzione </p><ol><li> Outfit ogni condizionamento chamber con visiva, olfattiva, e stimoli tattili per creare un contesto che è distinto da quello dei ratti hanno sperimentato durante le sedute di PDT. Ad esempio, se Contesto 1 è stato utilizzato in PDT, configurare la camera come Contesto 2 per le sessioni di estinzione. Fare riferimento al contesto utilizzato durante l&#39;estinzione come Context B. </li><li> Condurre sessioni di estinzione identico a sedute di PDT, ma non fornire alcuna etanolo durante il CS +. <ol><li> Utilizzare lo stesso programma informatico che è stato utilizzato durante PDT per presentare il CS + e CS. </li><li> Per mantenere la coerenza tra le fasi dell&#39;esperimento, caricare una siringa vuota nella pompa a siringa, e lasciare la pompa accesa. Assicurarsi che il tubo di polietilene non contiene etanolo prima sessione di estinzione. </li></ol></li><li> Continuare sessioni di estinzione tutti i giorni fino ratti non è più spettacolo discriminazione risponde tra il CS + e CS. NOTA: le voci della porta fatte in risposta al CS + dovrebbe diminuire gradualmente tra le sessioni di estinzione, fino a quando non vi è alcuna difference tra voci di porta fatte in risposta al CS + e le voci di porta fatte in risposta al CS. Il numero esatto di sessioni di estinzione necessarie per estinguere condizionata rispondere può variare, ma come una guida, ha intenzione di circa 8 sessioni. </li><li> Pulire l&#39;apparecchiatura al termine di ogni sessione, come descritto nel passaggio 5.7. </li></ol><p class="jove_title"> 7 Context-indotta Rinnovo di prova </p><ol><li> Dotare ogni camera di condizionamento con i, olfattiva, e stimoli tattili visive per creare il contesto che è stato sperimentato durante le sedute di PDT. Ad esempio, se un determinato topo esperto Contesto 1 a sedute di PDT, configurare sua camera come Contesto 1 nella sessione di test. Fare riferimento al contesto utilizzato durante la prova di rinnovamento come Context A. </li><li> Effettuare il test di rinnovamento indotta contesto identico a sedute di PDT, ma non fornire alcuna etanolo durante la prova di rinnovo. Presentare il <br /> CS + e CS-come prima durante le sedute di PDT, ma trattenere l&#39;etanolo. <ol><li> Utilizzare lo stesso pr del computerOgram che è stato utilizzato durante PDT per presentare il CS + e CS. </li><li> Per mantenere la coerenza tra le fasi, caricare una siringa vuota nella pompa a siringa, e lasciare la pompa accesa. Assicurarsi che il tubo di polietilene non contiene etanolo prima della sessione di test. </li></ol></li><li> Osservare il rinnovo indotta contesto pavloviano condizionata comportamenti alcol-ricerca. NOTA: Questo effetto rinnovamento sarà evidente da un aumento selettivo del numero di voci portuali effettuate in risposta al CS +, con nessun cambiamento nel comportamento ingresso porto made in risposta al CS, rispetto al CS + e CS-rispondendo osservato il fine della formazione estinzione. </li><li> Pulire l&#39;attrezzatura al termine della sessione, come descritto al punto 5.7. </li></ol>

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H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Context-induced+relapse+of+conditioned+behavioral+responding+to+ethanol+cues+in+rats.">Context-induced relapse of conditioned behavioral responding to ethanol cues in rats.</a> <em style='font-style: italic'>Biol Psychiatry</em>. <b>64</b>, 203-210 (2008).</li><li>Crombag, H. S., Shaham, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Renewal+of+drug+seeking+by+contextual+cues+after+prolonged+extinction+in+rats.">Renewal of drug seeking by contextual cues after prolonged extinction in rats.</a> <em style='font-style: italic'>Behav Neurosci</em>. <b>116</b>, 169-173 (2002).</li><li>Fuchs, R. A., Eaddy, J. L., Su, Z. I., Bell, G. 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<p class="jove_content"> Tutte le procedure sperimentali sono state approvate dal comitato di ricerca Animal Ethics Concordia University e sono conformi alle linee guida del Canadian Council on Animal Care. <span style="font-size: 14px; line-height: 28px;">La pubblicazione open-access di questo articolo è stata resa possibile dalla sponsorizzazione da Med Associates Inc.</span> </p>

<p class="jove_content"> I risultati di questa procedura rivelano che stimoli ambientali discreti che normalmente accompagnano la consegna alcol può acquisire la capacità di guidare il comportamento-alcol in cerca. Essi dimostrano inoltre che gli stimoli contestuali associati alla disponibilità o l&#39;assenza di alcol prima può guidare condizionata risposte comportamentali a stimoli alcol-predittiva discreti. </p><p class="jove_step"> Passaggi critici all&#39;interno del protocollo </p><p class="jove_content"> I contesti nella presente compito incorporano stimoli esterni al ratto. Tuttavia, lo stato interoceptive del ratto a test può anche costituire un &#39;contesto&#39; che potrebbero influenzare il rinnovamento <sup>20.</sup> E &#39;quindi importante abituare i ratti a tutte le procedure che possono verificarsi prima della prova di rinnovamento e modificare il loro stato interoceptive, come microinfusions intracraniche o iniezioni sistemiche. Inoltre, assuefazione a microinfusions o iniezioni sistemiche dovrebbe essere condotta prima sia estinzione e PDT sessions, per evitare che tali procedure di diventare associata ad una specifica fase dell&#39;esperimento. </p><p class="jove_content"> Parametri temporali, come la durata CS, intervallo CS-USA, e l&#39;intervallo tra le prove (ITI), sono da tempo noti per influenzare condizionata rispondere in una varietà di paradigmi di condizionamento <sup>21-26.</sup> Analogamente, i parametri temporali possono influenzare l&#39;osservazione dell&#39;effetto rinnovamento nella nostra procedura. Ogni set di parametri diversi da quelli specificati nel protocollo deve essere convalidato sperimentalmente in studi pilota prima di procedere ad impiegarli nelle indagini più grandi. </p><p class="jove_step"> Modifiche e risoluzione dei problemi </p><p class="jove_content"> Precedenti ricerche hanno trovato che la quantità di etanolo che i ratti consumano nella gabbia casa può variare in funzione del fornitore da cui i ratti vengono acquistati <sup>17,27.</sup> Fornitore è quindi una considerazione importante per questa fase dello studio. Analogamente, la scelta del ceppo ratto dovrebbe essere considered con attenzione, in quanto vi sono numerose segnalazioni di differenze di ceppi di consumo di etanolo <sup>28-31.</sup> </p><p class="jove_content"> Prima opera in un paradigma di condizionamento avversivo ha dimostrato che l&#39;età evolutiva dei ratti influenza l&#39;osservazione di rinnovamento di un pavloviano condizionata risposta di paura <sup>32.</sup> La mancanza di un effetto di rinnovamento in ratti molto giovani (di età inferiore ai 20 giorni dopo la nascita) è stato suggerito di essere dovuto alla codifica compromissione di informazioni contestuali <sup>33.</sup> L&#39;età evolutiva di ratti da utilizzare nel presente procedimento deve quindi essere preso in considerazione. </p><p class="jove_content"> Rinnovo pavloviano condizionata comportamento alcol-ricerca è stato osservato quando il 10% <sup>9,16,</sup> 15% <sup>14,</sup> o 20% <sup>15</sup> soluzioni di etanolo sono stati utilizzati durante la gabbia di casa accesso etanolo e PDT fasi dell&#39;esperimento. I ricercatori potrebbero usare una qualsiasi di queste concentrazioni di etanolo, ai fini della costituzione di tale procedura nel loro laboratories. </p><p class="jove_content"> Durante l&#39;acquisizione di una formazione discriminazione pavloviano è importante controllare le porte del fluido dopo ogni sessione per garantire che tutti i etanolo che è stato consegnato è stato consumato. Lasciando topi nelle camere di condizionamento per un ulteriore 10 - 15 minuti dopo ogni sessione può essere fatto all&#39;inizio del PDT per incoraggiare il consumo di qualsiasi etanolo che non è stato consumato durante la sessione. Il numero di ratti che lasciano etanolo non consumato nei porti dovrebbe diminuire, come progredisce PDT. Se entro la fine di ratti PDT sono (a) non consumare tutto il etanolo e / o (b) non risponde al CS + allora queste discipline deve essere eliminato dallo studio. La maggior parte dei ratti affidabile rispondere più al + CS rispetto al CS. Tuttavia, alcuni ratti possono rispondere ad alti livelli per entrambe le indicazioni, quindi non mostra evidenza di discriminazione. I ratti che mostrano elevata rispondono sia al CS + e CS-durante PDT non vengono eliminati dallo studio. Nella nostra esperienza, questi soggetti mostrano discriminazione notevoledurante la prima sessione di estinzione in assenza di etanolo, suggerendo che i loro livelli elevati di rispondere alla CS durante PDT sono guidati dalla presenza di etanolo nella valvola liquido. Questi soggetti tendono a mostrare gli effetti di rinnovo robusti. </p><p class="jove_step"> Limiti della procedura </p><p class="jove_content"> Poiché il test rinnovo viene condotta in assenza di etanolo, voci di porta condizionati indotte dal CS + diminuiscono gradualmente tutta la sessione di test. Pertanto, l&#39;importo complessivo del comportamento generato prova, contro la quale i cambiamenti sperimentalmente indotte possono essere valutate, è piuttosto bassa. Perché in questo compito risposta tende ad essere superiore al via contro la fine della sessione di test, è importante esaminare gli effetti delle manipolazioni sperimentali sulla condizionato rispondono a titolo di prova-by-processo. </p><p class="jove_content"> Le voci nella presa del fluido forniscono la misura di dipendente principale in questo compito. Tuttavia, una voce porta errata potrebbe essere registrata se un animale è abbastanza vicino alla presa del fluido di rompere il raggio infrarosso attraverso l&#39;apertura della porta fluido con i baffi. Anche se poco frequenti, le risposte errate sono evidenti come un drammatico aumento in porto totale voci relative al comportamento del soggetto in sessioni precedenti. Videoregistrando gli animali faciliterebbe l&#39;individuazione di possibili risposte errate, e consentire un&#39;analisi più raffinata dei comportamenti esibiti dai ratti per tutta la formazione e la sperimentazione. </p><p class="jove_step"> Importanza della tecnica rispetto ai metodi esistenti / alternative </p><p class="jove_content"> Rinnovamento Contesto-indotta del comportamento farmaco-seeking è ampiamente studiata utilizzando un compito di condizionamento strumentale in cui la variabile dipendente primaria è una risposta operante, come ad esempio una pressa a leva <sup>8,34</sup> o il naso colpire <sup>12.</sup> Nel presente procedimento, la risposta in cerca di droga è &#39;approccio condizionato&#39; alla porta del fluido che si manifesta da presentazioni diil + CS. Questa risposta è probabilmente acquisita attraverso l&#39;apprendimento pavloviano, che è il processo di apprendimento che media la formazione di associazioni tra stimoli ambientali e gli effetti farmacologici delle sostanze d&#39;abuso negli esseri umani. </p><p class="jove_step"> Le applicazioni future </p><p class="jove_content"> Questo compito può essere utilizzato in combinazione con neurofarmacologia, neurochimica optogenetics e per studiare i meccanismi neurali che sono coinvolti nel rinnovamento indotta contesto dei comportamenti alcol-cerca pavloviano condizionata <sup>16.</sup> Inoltre, i meccanismi comportamentali e neurali che regolano l&#39;acquisizione e l&#39;estinzione di pavloviano condizionata comportamenti alcol-ricerca possono essere studiate. Infine, questo compito può essere utilizzato per esplorare manipolazioni di estinzione che potrebbero impedire il rinnovamento, una direzione importante per la ricerca traslazionale volta a ridurre l&#39;impatto del farmaco associato contesti ambientali sulla reattività di droga segnali predittivi dopo la terapia di esposizione spunto in addi umanocts. </p>

<p class="jove_content"> Rimanendo sobrio è una notevole sfida per gli individui che soffrono di disturbi da abuso di alcool. L&#39;astinenza è un momento di vulnerabilità per l&#39;impatto comportamentale, psicologico e fisiologico degli stimoli ambientali che abitualmente accompagnano l&#39;uso di alcol, che possono, attraverso il condizionamento pavloviano, diventano associati ebbrezza <sup>1,2.</sup> L&#39;esposizione a stimoli predittivi alcol può suscitare risposte condizionate come alcol desiderio, che possono promuovere comportamenti alcol-cerca che facilitano la ricaduta <sup>3,4.</sup> </p><p class="jove_content"> Le sequenze stereotipate di comportamento che portano al consumo di alcol può causare alcuni tipi di stimoli per essere regolarmente sperimentato immediatamente prima che gli effetti farmacologici di assunzione di alcol. Ad esempio, la vista, l&#39;olfatto e il gusto di alcol sono proprietà sensoriali di alcol che precedono affidabile intossicazione. Oltre a tali segnali, che sono indicati come &#39;discreti&#39; o &#39;Proximaspunti l &#39;, contesti ambientali in cui i farmaci vengono regolarmente utilizzati possono anche provocare desiderio <sup>5,6.</sup> L&#39;esposizione a luoghi fisici in cui i farmaci sono stati precedentemente utilizzati possono dunque essere un trigger critico per recidiva <sup>7.</sup> </p><p class="jove_content"> I modelli animali sono stati sviluppati per studiare i meccanismi neurali che mediano l&#39;impatto della droga contesti associati al comportamento di droga ricerca di <sup>8-13.</sup> La procedura qui descritta permette l&#39;indagine di come contesti associati al consumo di alcol possono modulare il comportamento-alcol alla ricerca che si manifesta da un discreto, alcool predittivo cue pavloviano. </p><p class="jove_content"> Formazione discriminazione pavloviano è condotto in un contesto ambientale specifico, in cui i ratti sono addestrati a behaviorally distinguere tra due stimoli uditivi condizionata, un + CS che è associato con l&#39;alcol, e un CS che non è. Sessioni di estinzione vengono poi condotti in un contesto diverso, in cui rispondendo alla CS+ Diminuisce a causa di alcol trattenuta. Successivamente, riesposizione al alcol associato, nell&#39;ambito della formazione pavloviano innesca un aumento selettivo dei comportamenti alcol-cerca suscitato dal CS +, con nessun cambiamento nel rispondere alla CS. Questo risultato, che abbiamo sempre replicato <sup>9,14-16,</sup> si estende risultati di procedure strumentali di condizionamento in cui sono stati trovati per stimolare il rinnovamento delle risposte operanti connessi con la somministrazione di farmaci <sup>10,13</sup> contesti di droga. </p>

<table><tbody><tr><td>Heavy-duty utility cart</td><td>Rubbermaid</td><td>#4520-88</td><td>Used to transport rats in their home cage from the colony room to the behavior room or other facilities</td></tr><tr><td>Rats, Long-Evans</td><td>Harlan</td><td></td><td>Rats weighed 220-240 g on arrival</td></tr><tr><td>Ohaus scale</td><td>Fisher</td><td>#S402421</td><td>For weighing rats; tare the bottom of a plastic container first, then put the rat in it to weigh it</td></tr><tr><td>Beta-chips, aspen</td><td>Harlan</td><td>#7090A</td><td>Bedding for home cage</td></tr><tr><td>Nylabone</td><td>Bio-Serv</td><td>#K3580</td><td>Enrichment provided in the home cage</td></tr><tr><td>Autoclavable rodent chow</td><td>Agribrands, Charles River</td><td>#5075</td><td>For feeding rats</td></tr><tr><td>Wire bar cage lid</td><td>Ancare</td><td>#R20SS2B</td><td>Custom made with two slots for&amp;nbsp; bottles</td></tr><tr><td>Cage bottom</td><td>Ancare</td><td>#R20PC</td><td>To house rats</td></tr><tr><td>Stoppers</td><td>Ancare</td><td>#8.5</td><td>Rubber stoppers to occlude opening in water bottles &amp;amp; graduated cylinders; goes with ball point sipper tubes&amp;nbsp;</td></tr><tr><td>Ball point tubes, 1.5&quot; length</td><td>Ancare</td><td>#TD-99</td><td>Inserted in stoppers, enables rats to drink from water bottles and graduated cylinders</td></tr><tr><td>Water bottles</td><td>Ancare</td><td>#FSPC8HT</td><td>473 ml (16 oz) capacity, needs to be occluded with stopper</td></tr><tr><td>Graduated cylinders, 100 ml</td><td>Fisher</td><td>#0300741</td><td>To contain ethanol during alcohol exposure in the home cage; spouted end needs to be cut off to enable occlusion with stoppers</td></tr><tr><td>Mettler scale</td><td>Mettler Toledo</td><td>#MS6001S</td><td>For weighing water bottles and ethanol cylinders</td></tr><tr><td>Polyethylene tubing, 1/32&quot; inner diameter,&amp;nbsp; 1/32&quot; wall thickness, 3/32&quot; outside diameter</td><td>Fisher</td><td>#14-169-1A</td><td>To connect 20 ml ethanol syringe to fluid port</td></tr><tr><td>Needle, 18 g, 1.5&quot; length</td><td>Fisher</td><td>#B305199</td><td>To connect polyethylene tubing to ethanol syringes; needs to be filed</td></tr><tr><td>Syringes, 20 ml, Luer-Lok non-sterile</td><td>Fisher</td><td>#1482316J</td><td>To contain ethanol on the fluid pumps; needs to be connected to the polyethylene tubing by a 18 G 1 1/2 needle</td></tr><tr><td>Syringes, 60 ml, Luer-Lok non-sterile</td><td>Fisher</td><td>#14-820-11</td><td>To flush polyethylene tubing after PDT sessions</td></tr><tr><td>Single speed syringe pump, 3.33 rpm</td><td>Med Associates Inc.</td><td>#ENV-018MD</td><td>To hold 20 ml ethanol syringes and deliver ethanol</td></tr><tr><td>Fluid port</td><td>Med Associates Inc.</td><td>#ENV-254-CB</td><td>To enable ethanol delivery during PDT sessions; connected to ethanol syringes with polyethylene tubing</td></tr><tr><td>Port entry infrared detector</td><td>Med Associates Inc.</td><td>#ENV-215M</td><td>Fixed on both sides of&amp;nbsp; fluid port; need 2 per fluid port</td></tr><tr><td>Houselight, 28 V, 100 mA</td><td>Med Associates Inc.</td><td>#ENV-135M</td><td>Provides general lighting in conditioning chamber</td></tr><tr><td>Clicker module</td><td>Med Associates Inc.</td><td>#ENV-225SM</td><td>Auditory stimulus for conditioning chamber</td></tr><tr><td>White noise generator with speaker</td><td>Med Associates Inc.</td><td>#PHM-100</td><td>Auditory stimulus for conditioning chamber</td></tr><tr><td>Bar floor</td><td>Med Associates Inc.</td><td>#ENV-009A-GF</td><td>To go with modular test chamber</td></tr><tr><td>Bench coat, absorbent</td><td>VWR</td><td>#89126-790</td><td>Absorbent paper to put in metal tray, needs to be cut to fit the metal tray</td></tr><tr><td>Polycarbonate sheet, 1/8&quot; thick, 12-5/8&quot; x 10&quot;</td><td>Johnston Plastics</td><td>#30102515</td><td>To make smooth transparent floor insert used for context 1. Needs to be cut in 11.5&quot; x 12.25&quot; pieces</td></tr><tr><td>Aluminium sheet perforated, 0.063&quot; thick, 48&quot; x 96&quot;, 0.250&quot; holes x 0.375&quot; centers</td><td>Anica Steel</td><td></td><td>To make perforated floor insert used for context 2. Needs to be cut in 11.5&quot; x 12.25&quot; pieces</td></tr><tr><td>Black cardboard, 0.053&quot; thick, 28&quot; x 44&quot;</td><td>Omer DeSerres</td><td>#P1909</td><td>To construct panels covering the ceiling, door and back wall of the conditioning chamber in context 1. Needs to be cut to size. Each chamber needs three panels measuring approximately 11.5&quot; x 13.25&quot;. The panels can be joined by the narrower side using duct tape.</td></tr><tr><td>All-purpose strength duct tape, 1.88&quot; x 45 yd.</td><td>Duck Tape</td><td></td><td>Adhesive material to connect the black cardboard panels used to cover the back wall, ceiling and door in context 1.</td></tr><tr><td>Modular test chamber</td><td>Med Associates Inc.</td><td>#ENV-009A</td><td>Conditioning chamber</td></tr><tr><td>Sound attenuating cubicles</td><td>Med Associates Inc.</td><td>#ENV-200R3AM</td><td>Comes with installed 28 V DC fan (#ENV-025F)</td></tr><tr><td>Petri dish</td><td>Fisher</td><td>#08748B</td><td>Bottom only, to contain the sprayed solution that provides the odor stimulus; needs to be centered in the metal tray in conditioning chambers</td></tr><tr><td>PCI interface package</td><td>Med Associates Inc.</td><td>#DIG-700P2-R2</td><td>Connects tabletop interface cabinet to computer; includes 1 interface card, 1 Decode card, 1 ribbon cable and 1 28 V DC power cable</td></tr><tr><td>Large tabletop cabinet and power supply</td><td>Med Associates Inc.</td><td>#SG-6510D</td><td>Used to contain the 12 cards included in the SmartCrtrl 8 Input, 16 Output package</td></tr><tr><td>SmartCtrl 8 Input, 16 Output package</td><td>Med Associates Inc.</td><td>#DIG-716P2</td><td>Used to connect and control the devices in the conditioning chamber. The package includes all the cables, the interface module and the connection panel.</td></tr><tr><td>Med PC IV software</td><td>Med Associates Inc.</td><td>#SOF-735</td><td>Software to run programs. Needs to be installed on the computer.</td></tr><tr><td>Monitor LG, 19&quot; LCD</td><td>Concordia computer store</td><td>#W1942TQ-BF</td><td>Computer monitor; to go with computer running Med PC IV</td></tr><tr><td>HP Z200 tower workstation</td><td>Concordia computer store</td><td>#BZ788US#ABA</td><td>Computer on which Med PC IV is run</td></tr><tr><td>Benzaldehyde, &amp;ge;99%</td><td>Les produits chimiques OMEGA Chemical Company Inc.</td><td>#B37-50</td><td>Dilute in tap water for almond context odor</td></tr><tr><td>Lemon oil, cold pressed, California</td><td>Sigma Aldrich</td><td>#W262528-1KG-K</td><td>Dilute in tap water for lemon context odor</td></tr><tr><td>Ethyl alcohol, 95%</td><td>Commercial Alcohols</td><td></td><td>Dilute in tap water to desired concentration</td></tr></tbody></table>

a procedure to observe context-induced renewal of pavlovian-conditioned alcohol-seeking behavior in rats

<p class="jove_content"> Contesti ambientali in cui l&#39;abuso di droghe vengono consumate può innescare desiderio, una soggettiva risposta pavloviano condizionata che può facilitare il comportamento di droga ricerca e tempestiva ricaduta in tossicodipendenti astinenti. Abbiamo sviluppato una procedura per studiare i processi comportamentali e neurali che mediano l&#39;impatto del contesto sul comportamento alcol-seeking nei ratti. Dopo acclimatazione al gusto e gli effetti farmacologici del 15% di etanolo nella gabbia casa, ratti maschi a lungo Evans ricevano una formazione discriminazione pavloviano (PDT) in camere di condizionamento. In ciascuna sessione giornaliera (lun-ven) PDT, 16 studi ciascuno dei due stimoli diversi 10 sec uditivo condizionata verificano. Durante uno stimolo, il CS +, 0,2 ml di 15% etanolo viene consegnato in una porta di fluido per il consumo orale. Il secondo stimolo, il CS, non è accoppiato con etanolo. Attraverso sessioni, le voci nella porta del fluido durante l&#39;aumento CS +, mentre le voci durante il CS stabilizzare a un livello inferiore, indicando che una predictive associazione tra il CS + e l&#39;etanolo è acquisito. Durante PDT ciascuna camera è dotata di una configurazione specifica di stimoli contestuali visivi, olfattivi e tattili. A seguito di PDT, formazione estinzione è condotto nella stessa camera che è ora dotato di una diversa configurazione di stimoli contestuali. Il CS + e CS sono presentati come prima, ma l&#39;etanolo sia ritirato, che causa un graduale declino in voci di porta durante il CS +. Alla prova, i ratti vengono inseriti di nuovo nel contesto PDT e presentati con il CS + e CS-come prima, ma senza etanolo. Questa manipolazione innesca un aumento robusto e selettivo il numero di voci di porta fatte durante l&#39;alcool predittivo CS +, con nessun cambiamento nel rispondere durante il CS. Questo effetto, denominato rinnovamento indotta contesto, illustra la potente capacità di contesti legati al consumo di alcol per stimolare comportamenti alcol-cerca in risposta a stimoli di alcol di Pavlov. </p>

<p class="jove_content"> Accesso etanolo intermittente nella gabbia di casa: Il consumo di etanolo (g / kg) e l&#39;etanolo aumento preferenza attraverso sessioni di accesso etanolo intermittente nella gabbia di casa, in genere di raggiungere livelli stabili entro 8-12 sessioni <strong>(Figura 1).</strong> Nel set di dati mostrata nella <strong>figura 1,</strong> il consumo stabile di etanolo media attraverso le ultime due sessioni di gabbia di casa accesso etanolo variava 1,33-6,44 g / kg per i singoli ratti. La gabbia casa fase il consumo di etanolo dell&#39;esperimento può essere interrotto una volta che il g / kg il consumo è stabilizzato, e ratti sono mantenuta a questo livello stabile di bere per un certo numero di sedute. </p><p class="jove_content"> Per analizzare i dati di questa fase, condurre analisi separata misure ripetute della varianza (ANOVA) attraverso il fattore entro i soggetti della sessione per il consumo di etanolo (g / kg) e l&#39;etanolo preferenza. </p><p class="jove_content"> Schematica della procedura di rinnovo indotta contesto: Ogni fase delprocedura sperimentale (PDT, l&#39;estinzione, prova rinnovo) è illustrato nella <strong>figura 2.</strong> Vedere la didascalia per ulteriori dettagli. </p><p class="jove_content"> Risposta misure e analisi dei dati: In tutte le fasi comportamentali descritte di seguito, le risposte di ingresso della porta fatte durante il CS + e CS sono normalizzati con riferimento alle pre-CS basali risposte di ingresso della porta. Per creare le misure normalizzate CS, sottrarre il numero di voci portuali effettuate durante l&#39;intervallo di riferimento pre-CS (10 secondi immediatamente prima dell&#39;inizio CS) dal numero di voci portuali effettuati durante le presentazioni del CS + e CS. Le misure CS normalizzati normalizzato CS + e servono come indicatori di comportamento suscitato particolare da presentazioni CS, elevati sopra dei livelli basali. </p><p class="jove_content"> Condurre misure ripetute ANOVA separati con i fattori oggetto entro ripetute di sessione e tipo CS (CS + vs CS) per le fasi di estinzione e PDT. Per i confronti tra fase (PDT, estinzione, rinnovareal test), i dati di utilizzo media attraverso le ultime due sedute di PDT e l&#39;estinzione e la una sessione di dati di test di rinnovo in ripetute misure ANOVA. Per dare seguito a significativi effetti principali e le interazioni, t-test uso <em>post hoc</em> per campioni indipendenti o appaiati che sono corretti per confronti multipli. Criterio di significatività è fissato a α = 0.05. </p><p class="jove_content"> PDT: risposte ingresso porta per l&#39;aumento CS + attraverso sedute di PDT, considerando che le risposte di ingresso porta al CS rimane bassa e stabile, nel complesso. Si noti che le risposte di ingresso porta al CS può aumentare leggermente nei primi sessioni di allenamento PDT; Tuttavia, voci di porta fatte in risposta alla CS stabilizzare a un livello molto più basso rispetto a quelli realizzati in risposta al CS +. Discriminazione di successo è evidenziato da un innalzamento statisticamente significativo in voci di porta durante il CS + rispetto al CS <strong>(Figura 3).</strong> Questo risultato indica che pavloviano condizionata comportamenti alcol-ricerca ha essereen istituito per il CS +, ma non il CS. </p><p class="jove_content"> Estinzione: risposte ingresso porta per il CS + graduale declino tra le sessioni di estinzione. Risposte ingresso di porta al CS rimangono bassi e stabili tra le sessioni di estinzione. Idealmente, discriminati risposta è abolito entro la fine della formazione di estinzione, come dimostra l&#39;assenza di una differenza statisticamente significativa tra i valori delle porte al CS + e CS. Tuttavia, non è raro che un certo grado di discriminazione rispondere a rimanere, anche alla fine della formazione estinzione. In questo caso, è importante verificare che le risposte di ingresso porta al CS + alla fine della formazione estinzione sono statisticamente inferiore risposte ingresso porta al CS + all&#39;inizio di estinzione, nonché al termine della PDT <strong>(figura 3)</strong> . Questo modello di risultati indica che pavloviano condizionata comportamento alcol-ricerca è stata notevolmente estinta. </p><p class="jove_content"> Context-indotta prova rinnovo: Portrisposte ingresso all&#39;aumento CS + nel test rinnovo indotta contesto, relativa alle voci portuali CS + osservati alla fine della formazione estinzione. Tale incremento è selettivo per il CS +, come voci di porta al CS rimangono inalterate. Inoltre, voci di porta al CS + sono statisticamente elevati rispetto a voci di porta al CS nel test rinnovo indotta contesto <strong>(Figura 4).</strong> Conferma statistica dei confronti tra voci di porta CS + tra l&#39;estinzione e la prova, così come i confronti di CS + e CS voci di porta all&#39;interno di test, indica un rinnovamento selettivo di comportamento-alcol in cerca pavloviano condizionata indotta dal contesto di alcol associato a test. </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 1" fo:content-width="4in" src="/files/ftp_upload/51898/51898fig1highres.jpg" width="400" /><br /> <strong>Figura 1 Escalation del consumo di etanolo e di preferenza indotta da etanolo accesso intermittente nella gabbia casa.</strong> Media &amp;# 177; SEM <strong>(A)</strong> consumo di etanolo (g / kg) e <strong>(B)</strong> la preferenza etanolo attraverso 12 sessioni di accesso etanolo intermittente nella gabbia casa. Entrambe le misure aumentano tipicamente tra le sessioni e poi si stabilizzano. Consumo di etanolo Stabile media attraverso le ultime due sessioni di gabbia di casa accesso etanolo variava 1,33-6,44 g / kg per i singoli ratti. Questi dati sono stati ottenuti utilizzando ratti maschi lungo Evans potabile 15% (v / v) di etanolo. </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 2" fo:content-width="4in" src="/files/ftp_upload/51898/51898fig2highres.jpg" width="400" /><br /> <strong>. Figura 2 Uno schema della procedura sperimentale</strong> La procedura si compone di tre fasi: formazione discriminazione pavloviano (PDT), l&#39;estinzione, e test di rinnovamento indotta contesto. Durante ogni sessione di PDT, ratti ricevono 16 studi di uno stimolo uditivo condizionato (CS +) che segnala la consegna di etanolo e 16 studi diun secondo stimolo uditivo (CS), che non è associato con la consegna di etanolo. Sedute di PDT avvengono in Context A, composto da una configurazione specifica di visivo, olfattivo e stimoli tattili. Durante le sessioni di estinzione, che si svolgono nel contesto B, composto da un diverso insieme di stimoli visivi, olfattivi, tattili e, il CS + e CS vengono presentati, ma l&#39;etanolo sia ritirato. Per la prova di rinnovamento indotta contesto, i ratti vengono restituiti Contesto A, e il CS + e CS sono presentati senza etanolo. <a href="https://www.jove.com/files/ftp_upload/51898/51898fig2highres.jpg" target="_blank">Cliccate qui per vedere una versione più grande di questa figura.</a> </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 3" fo:content-width="4in" src="/files/ftp_upload/51898/51898fig3highres.jpg" width="400" /><br /> <strong>Figura 3 Acquisizione ed estinzione di discriminate Pavlov condizionata, di rispondere per l&#39;etanolo.</strong> Media ± SEM voci di porta normalizzato made durante il CS + e CS-attraverso PDT (sessioni 1-18) in contesto A e di estinzione (sessioni 19-26) in contesto B. Questi dati sono stati ottenuti utilizzando ratti maschi a lungo Evans, addestrato con il 10% (v / v) etanolo come stimolo incondizionato (US) che è stato presentato in collaborazione con il + CS durante la PDT. * P &lt;0,05, CS + rispetto al CS. Figura originariamente pubblicato in Chaudhri, Sahuque, e Janak (2008). Usato con il permesso di Elsevier. </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 4" fo:content-width="5in" src="/files/ftp_upload/51898/51898fig4highres.jpg" width="500" /><br /> <strong>Figura 4 Context-indotta rinnovo del</strong> ± SEM voci <strong>di Pavlov condizionata, di rispondere per l&#39;etanolo.</strong> Media normalizzata portuali effettuate durante il CS + e CS-in sedute di PDT in contesto A, sessioni di estinzione in contesto B, e test di rinnovamento nel contesto A. I dati sono media degli ultimi due sessioni per PDT e di estinzione; il test di rinnovamento è una singola sessione. Questei dati sono stati ottenuti utilizzando ratti maschi a lungo Evans, addestrati con il 10% (v / v) di etanolo come stimolo incondizionato (US) che è stato presentato in collaborazione con il CS + durante la PDT. * P &lt;0,05; ** P &lt;0,001, CS + rispetto al CS. #p &lt;0.05, CS + PDT rispetto CS + estinzione. ^ P &lt;0.05, CS + estinzione contro CS + prova. Figura originariamente pubblicato in Chaudhri, Sahuque, e Janak (2008). Usato con il permesso di Elsevier. </p>

Watch this Scientific Journal Video about A Procedure to Observe Context-induced Renewal of Pavlovian-conditioned Alcohol-seeking Behavior in Rats at JoVE.com

A Procedure to Observe Context-induced Renewal of Pavlovian-conditioned Alcohol-seeking Behavior in Rats

<p class="jove_content"> Una procedura per studiare la capacità di un contesto ambientale di alcol associato per attivare il rinnovamento dei comportamenti alcol-seeking nei ratti è descritto. </p>

Una Procedura di osservare Context-indotta Rinnovo pavloviano condizionata alcol-seeking comportamento in Rats

Environmental contexts in which drugs of abuse are consumed can trigger craving, a subjective Pavlovian-conditioned response that can facilitate ...

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12.8K Views.  Concordia University.  Una procedura per studiare la capacità di un contesto ambientale di alcol associato per attivare il rinnovamento dei comportamenti alcol-seeking nei ratti è descritto. 

Video: A Procedure to Observe Context-induced Renewal of Pavlovian-conditioned Alcohol-seeking Behavior in Rats

Esterification

<p>Source: Lara Al Hariri at the University of Massachusetts Amherst, MA, USA</p>
<ol class="note-items">
	<li data-note-item="1" data-parent-collapse="">Preparation of the Laboratory<button class="expand">Expand</button>
	<p><em>Here, we show the laboratory preparation for 10 students working in pairs, with some excess. Please adjust quantities as needed.</em></p>
	<ul class="lab-items">
		<li data-sub-note-item="1-2">
		<div class="lab-step">First, put on a lab coat, safety glasses, and nitrile gloves.<strong> Note:</strong> Some of the reagents are volatile or have unpleasant odors, so you will handle them in a fume hood.</div>
		</li>
		<li data-sub-note-item="1-3">
		<div class="lab-step">Ensure that you have glass and organic waste containers and that each sink has paper towels.</div>
		</li>
		<li data-sub-note-item="1-4">
		<div class="lab-step">Follow the table to designate codes for the carboxylic acids and alcohols used in this lab. The carboxylic acids are coded A through C, and the alcohols are coded D through I. The students will use a 3:1 molar ratio of alcohol to carboxylic acid in the reaction.
		<table align="center" height="224" style="border-collapse: collapse; font-size: 17px; line-height: 24px;">
			<tbody>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Code</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Compound</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>MW<br />
					(g/mol) </strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Density at RT (g/cm<sup>3</sup>)</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Moles per vial</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Grams per vial</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>cm<sup>3</sup> per vial </strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Number of vials</strong></td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">A</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Acetic acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">60.05</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1.049</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.07</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">4.2035</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">4.0</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">3</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">B</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Butyric acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">88.11</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.953</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.07</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">6.1677</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">6.5</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">3</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">C</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Anthranilic acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">137.14</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">-</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.07</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">9.5998</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">-</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">D</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1-Propanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">60.10</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.803</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.21</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">12.6210</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">15.7</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">E</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Isoamyl alcohol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">88.15</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.810</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.21</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">18.512</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">22.8</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">F</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1-Butanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">74.12</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.810</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.21</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">15.565</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">19.2</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">G</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Ethanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">46.07</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.789</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.21</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">9.6747</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">12.3</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">H</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Methanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">32.04</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.791</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.21</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">6.7284</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">8.5</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">2</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">I</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1-Octanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">130.23</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.830</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">0.21</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">27.348</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">32.9</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1</td>
				</tr>
			</tbody>
		</table>
		</div>
		</li>
		<li data-sub-note-item="1-5">
		<div class="lab-step">Prepare the three carboxylic acids: acetic acid, butyric acid, and anthranilic acid. Each group will need 0.07 moles of their assigned carboxylic acid in a vial.</div>
		</li>
		<li data-sub-note-item="1-6">
		<div class="lab-step">Tare a 10-mL glass vial on an analytical balance in a fume hood. Use a Pasteur pipette to measure 4.2035 g of acetic acid, which will be about 4 mL. Cap the vial and label it &#8216;A&#8217;. Prepare two more labeled vials of acetic acid in the same way.</div>
		</li>
		<li data-sub-note-item="1-7">
		<div class="lab-step">Weigh 6.1677 g of butyric acid, which will be about 6.5 mL, in a tared 10-mL vial. Cap the vial and label it &#8216;B&#8217;. Prepare two more vials of butyric acid in the same way.</div>
		</li>
		<li data-sub-note-item="1-8">
		<div class="lab-step">Use a small spatula to weigh 9.5998 g of solid anthranilic acid in a tared 10-mL vial. Cap the vial and label it &#8216;C&#8217;. You should now have a total of seven vials of carboxylic acid.</div>
		</li>
		<li data-sub-note-item="1-9">
		<div class="lab-step">Prepare the 7 alcohols. Each group will need 0.21 moles of their assigned alcohol in a vial. Use the table as a guide to prepare and label one vial of each alcohol. Prepare and label two vials of methanol.</div>
		</li>
		<li data-sub-note-item="1-10">
		<div class="lab-step">You should now have a total of seven vials of alcohol. Keep the reagent vials in the instructor&#39;s hood for now and put away the stock bottles of the carboxylic acids and alcohols.</div>
		</li>
		<li data-sub-note-item="1-11">
		<div class="lab-step">Assign each student group one of these seven combinations of reagents.
		<table align="center" height="224" style="border-collapse: collapse; font-size: 17px; line-height: 24px;">
			<tbody>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Ester code</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Carboxylic acid </strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Alcohol</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Ester product</strong></td>
					<td style="border: 1px solid; padding: 0px; text-align: center;"><strong>Scent</strong></td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">AD</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Acetic acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1-Propanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Propyl acetate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Pear</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">AE</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Acetic acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Isoamyl alcohol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Isoamyl acetate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Banana</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">AI</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Acetic acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1-Octanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Octyl acetate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Orange</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">BF</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Butyric acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">1-Butanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Butyl butyrate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Pineapple</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">BG</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Butyric acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Ethanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Ethyl butyrate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Strawberry</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">BH</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Butyric acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Methanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Methyl butyrate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Apple</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; text-align: center;">CH</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Anthranilic acid</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Methanol</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Methyl anthranilate</td>
					<td style="border: 1px solid; padding: 0px; text-align: center;">Grape</td>
				</tr>
			</tbody>
		</table>
		</div>
		</li>
		<li data-sub-note-item="1-12">
		<div class="lab-step">Prepare or obtain seven vials of the fruit scents and label them with the fruit names as references.</div>
		</li>
		<li data-sub-note-item="1-13">
		<div class="lab-step">Set out the following glassware and equipment at each student lab station (we suggest that students work in pairs):</div>
		</li>
		<li data-sub-note-item="1-14">
		<div class="lab-step">Put bottles of saturated sodium bicarbonate and 3 M sulfuric acid in a dedicated fume hood, along with a roll of plastic paraffin film and scissors.</div>
		</li>
		<li data-sub-note-item="1-15">
		<div class="lab-step">Set out filter paper and Pasteur pipettes in a central area so that every student group can easily access them.
		<table align="center" height="224" style="border-collapse: collapse; font-size: 17px; line-height: 24px;">
			<tbody>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Lab stand</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;2&#160;&#160;&#160;&#160;3-prong clamps</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Stir plate</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Lab jack</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Heating mantle</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Temperature controller</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;2&#160;&#160;&#160;&#160;Pieces of tubing (for condenser)</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Joint clip</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Tube of vacuum grease</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Box of laboratory wipes</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;10-mL graduated cylinder</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;600-mL beaker</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;50-mL round-bottom flask and flask stand</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Standard condenser</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Small stir bar</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Glass rod</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;1&#160;&#160;&#160;&#160;Package of pH paper</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px;">&#160;2&#160;&#160;&#160;&#160;Pasteur pipette bulbs</td>
				</tr>
			</tbody>
		</table>
		</div>
		</li>
		<li data-sub-note-item="1-16">
		<div class="lab-step">Just before the lab, fill a large insulated cooler with crushed ice. Place the cooler and an ice scoop in a central area in the lab.</div>
		</li>
		<li data-sub-note-item="1-17">
		<div class="lab-step">When the students arrive, show them how to safely waft vapors before giving them their assigned reagents.</div>
		</li>
	</ul>
	</li>
</ol>

Esterification: Synthesis of Ester by Fischer Esterification - Prep

Esterificazione

Source: Lara Al Hariri at the University of Massachusetts Amherst, MA, USA

	Preparation of the LaboratoryExpand
	Here, we show the laboratory preparation ...

Didattica

JoVE Lab Manual

Chimica

Lab Manual Sub Articles

Identifying Alcohols

<p>Source: Lara Al Hariri and Ahmed Basabrain at the University of Massachusetts Amherst, MA, USA</p>
<ol class="note-items">
	<li data-note-item="1" data-parent-collapse="">Ferric Chloride Test for Phenols<button class="expand">Expand</button>
	<p>In this lab, you will identify an unknown alcohol using the ferric chloride test, the Jones test, and the Lucas test. You&#39;ll test known alcohols alongside the unknown alcohol as examples of positive and negative results for each test.</p>
	<p>The four known alcohols are 1-butanol, a primary alcohol, 2-butanol, a secondary alcohol, 2-methyl-2-propanol, a tertiary alcohol, and phenol. The four possible unknown alcohols are 1-propanol, 2-propanol, 2-methyl-2-butanol, and para-chlorophenol.</p>
	<p>You&#39;ll start with the ferric chloride test for the presence of phenols. Iron(III) chloride, or ferric chloride, forms a brown complex when it&#39;s dissolved in water. If you add a phenol to this solution, a purple iron(III)-phenol complex will form instead. This effect is not observed with non-aromatic or aliphatic alcohols.</p>
	<ul class="lab-items">
		<li data-sub-note-item="1-4">
		<div class="lab-step">Put on a lab coat, safety glasses, and gloves before you begin. <strong>Note:</strong> The alcohols that you will test are toxic and flammable, and the reagents that you will use are toxic and corrosive, so you will perform this lab in a fume hood.</div>
		</li>
		<li data-sub-note-item="1-5">
		<div class="lab-step">Now, get your assigned unknown alcohol from your instructor and bring it to your fume hood. Label a 250-mL beaker &#8216;aqueous metal waste&#8217;.</div>
		</li>
		<li data-sub-note-item="1-6">
		<div class="lab-step">Next, put five small test tubes in a rack and label them &#8216;1-butanol&#8217;, &#8216;2-butanol&#8217;, &#8216;2-methyl-2-propanol&#8217;, &#8216;phenol&#8217;, and &#8216;unknown&#8217;. Bring the rack to the analytical balance to obtain phenol.</div>
		</li>
		<li data-sub-note-item="1-7">
		<div class="lab-step">If your unknown is a solid, bring it with you. Weigh about 0.2 g of phenol and pour it into the corresponding tube. Do the same for your unknown if it is a solid and then return to your hood.</div>
		</li>
		<li data-sub-note-item="1-8">
		<div class="lab-step">Now label three small vials &#8216;1-butanol&#8217;, &#8216;2-butanol&#8217;, and &#8216;2-methyl-2-propanol&#8217;.</div>
		</li>
		<li data-sub-note-item="1-9">
		<div class="lab-step">Bring the vials to the solvent hood and use clean Pasteur pipettes to fill each vial with about 1 mL of the matching alcohol.<strong> Note: </strong>Do not reuse pipettes.</div>
		</li>
		<li data-sub-note-item="1-10">
		<div class="lab-step">Next, obtain at least four Pasteur pipettes and a 250-mL beaker. Get more pipettes whenever you need them, and don&#39;t put used pipettes back in the beaker.</div>
		</li>
		<li data-sub-note-item="1-11">
		<div class="lab-step">In your fume hood, pipette 3 &#8211; 4 drops of 1-butanol, 2-butanol, and 2-methyl-2-propanol into the corresponding tubes.</div>
		</li>
		<li data-sub-note-item="1-12">
		<div class="lab-step">To the phenol tube, add several drops of deionized water and shake to mix. This will provide better visual results for the ferric test.</div>
		</li>
		<li data-sub-note-item="1-13">
		<div class="lab-step">If your unknown alcohol is a liquid, use a clean pipette to place 3 &#8211; 4 drops of it in the unknown test tube.</div>
		</li>
		<li data-sub-note-item="1-14">
		<div class="lab-step">Now, obtain the bottle of iron(III) chloride solution and add 2 &#8211; 3 drops to each test tube.</div>
		</li>
		<li data-sub-note-item="1-15">
		<div class="lab-step">Stir each mixture well with a glass rod. Use deionized water to rinse off residual alcohol before mixing the contents of the next tube. Make sure that the phenol dissolves completely along with the unknown if it is a solid.</div>
		</li>
		<li data-sub-note-item="1-16">
		<div class="lab-step">Wait 5 min before comparing your unknown to the known solutions. <strong>Note:</strong> If your unknown solution is purple, like the phenol solution, then your unknown is a phenol derivative. If it looks like the other three tubes, it is not a phenol derivative. Record the results of the test in your lab notebook.
		<h4><strong>Table 1: Identifying Unknown Alcohol</strong></h4>
		<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
			<tbody>
				<tr>
					<td style="text-align: center;"><strong>Alcohol</strong></td>
					<td style="text-align: center;"><strong>Ferric Chloride Test</strong></td>
					<td style="text-align: center;"><strong>Jones Test</strong></td>
					<td style="text-align: center;"><strong>Lucas Test</strong></td>
				</tr>
				<tr>
					<td>1-butanol</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td>2-butanol</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td>2-methyl-2-propanol</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td>Phenol</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td>Unknown</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
			</tbody>
		</table>
		<a href="https://www.jove.com/CDNSource/lm/labs/71/Table_1._Identifying_unknown_alcohol.xlsx" target="_blank">Click Here to download Table 1</a></div>
		</li>
		<li data-sub-note-item="1-17">
		<div class="lab-step">The next tests don&#39;t work with phenols, so you&#39;ll only test aliphatic alcohols for the rest of the lab. If your unknown is a phenol derivative, skip the instructions for the unknown and test only the three known aliphatic alcohols.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="2" data-parent-collapse="">The Jones Test for Aliphatic Primary and Secondary Alcohols<button class="expand">Expand</button>
	<p>In this section, you&#39;ll perform the Jones test for primary and secondary alcohols. Jones&#39; reagent is made with chromium trioxide and sulfuric acid in water, which forms chromic acid (H<sub>2</sub>CrO<sub>4</sub>)<em> in situ</em>. This powerful reagent oxidizes secondary alcohols to ketones, primary alcohols to aldehydes, which after forming an aldehyde hydrate, are further reduced to carboxylic acids.</p>
	<p>The oxidation state of chromium is the key to this test. Chromium is in the +6 oxidation state in the Jones&#39; reagent. The Cr(VI) complexes in the reagent give it its bright reddish, orange color.</p>
	<p>When Cr(VI) oxidizes an alcohol, chromium is reduced to the +3 oxidation state. First, the alcohol and chromic acid form a chromate ester. Then, a base (H<sub>2</sub>O) cleaves the C-H bond of the alcohol, forming the carbonyl group while reducing Cr(VI) to Cr(IV). Because the carbon of the alcohol undergoes a 2-electron oxidation, and Cr(VI) a 2-electron reduction, this step is a reduction-oxidation step. Cr(IV) participates in further oxidation steps and is eventually reduced to Cr(III). Cr(III) is often present as hexaaquachromium(III) ions&#160;&#8212; [Cr(H<sub>2</sub>O)<sub>6</sub>]<sup>3+</sup> &#8212; and Cr(III) complexes, where H<sub>2</sub>O molecules are replaced by one or more sulphate ions, [Cr(H<sub>2</sub>O)<sub>5</sub>(SO<sub>4</sub>)]<sup>+</sup>. These complexes give Cr(III) the characteristic green color. So, when you mix Jones&#39; reagent with a primary or secondary alcohol in acetone, the orange solution will turn green.</p>
	<p>Jones oxidation of alcohols doesn&#39;t work with tertiary alcohols because the -OH group is already bonded to three carbon atoms and cannot form an extra C-O bond. Thus, you won&#39;t see a color change if you combine Jones&#39; reagent with a tertiary alcohol because chromium isn&#39;t reduced.</p>
	<ul class="lab-items">
		<li data-sub-note-item="2-5">
		<div class="lab-step">Label four clean test tubes &#8216;1-butanol&#8217;, &#8216;2-butanol&#8217;, &#8216;2-methyl-2-propanol&#8217;, and &#8216;unknown&#8217; and place 10 drops of acetone in each. Acetone is the preferred solvent for Jones oxidation.</div>
		</li>
		<li data-sub-note-item="2-6">
		<div class="lab-step">Then, use clean pipettes to add 3 &#8211; 4 drops of the known aliphatic alcohols and your unknown alcohol to the corresponding tubes.</div>
		</li>
		<li data-sub-note-item="2-7">
		<div class="lab-step">Stir each solution well with the glass rod and rinse the rod with water after mixing each one.</div>
		</li>
		<li data-sub-note-item="2-8">
		<div class="lab-step">Now, obtain the bottle of Jones&#39; reagent and carefully add 2 &#8211; 3 drops to each tube. The reagent is strongly acidic and corrosive, so be careful with it and tightly cap the bottle when you&#39;re done. Then, mix each solution well, cleaning the glass rod after each one.</div>
		</li>
		<li data-sub-note-item="2-9">
		<div class="lab-step">Let the solutions react for 10 min, and then compare your unknown to the knowns. <strong>Note:</strong> If the unknown solution turned green, like 1-butanol and 2-butanol, the test is positive for a primary or secondary alcohol. If it stayed orange, like 2-methyl-2-propanol, the test is negative.</div>
		</li>
		<li data-sub-note-item="2-10">
		<div class="lab-step">Record the results in your lab notebook.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="3" data-parent-collapse="">The Lucas Test for Classifying Small Saturated Alcohols<button class="expand">Expand</button>
	<p>The Jones test doesn&#39;t distinguish between primary and secondary alcohols, so you&#39;ll use the Lucas test to identify which is which.</p>
	<p>Lucas&#39; reagent, which is a mixture of zinc chloride and hydrochloric acid, converts secondary and tertiary alcohols to chloroalkanes at room temperature. Chloroalkanes are nearly insoluble in water, so a positive result appears as the mixture separates into a cloudy chloroalkane-containing layer over a clear layer.</p>
	<p>The rate-determining step of the reaction involves converting the alcohol to a carbocation, so the speed of the reaction depends on how stable the carbocation is.</p>
	<p>Tertiary carbocations are very stable, so tertiary alcohols give a positive reaction almost immediately. Secondary carbocations are less stable, so secondary alcohols give a positive result after a few seconds to a few minutes. Primary carbocations are too unstable for this reaction, so primary alcohols give a negative result.</p>
	<ul class="lab-items">
		<li data-sub-note-item="3-5">
		<div class="lab-step">Now, label for clean test tubes as &#8216;1-butanol&#8217;, &#8216;2-butanol&#8217;, &#8216;2-methyl-2-propanol&#8217;, and &#8216;unknown&#8217;.</div>
		</li>
		<li data-sub-note-item="3-6">
		<div class="lab-step">Obtain the bottle of Lucas&#39; reagent and use a graduated cylinder to measure 2 mL of the reagent for each tube.</div>
		</li>
		<li data-sub-note-item="3-7">
		<div class="lab-step">Next, add 2 drops of 2-methyl-2-propanol to the appropriate tube. Immediately, stir the mixture for a few seconds and place it in the rack. The mixture will promptly separate into a clear lower layer and a cloudy upper layer.</div>
		</li>
		<li data-sub-note-item="3-8">
		<div class="lab-step">Thoroughly rinse the glass rod and record the immediate positive result in your lab notebook.</div>
		</li>
		<li data-sub-note-item="3-9">
		<div class="lab-step">Mix the remaining alcohols with Lucas&#39; reagent in the same way, using a fresh pipette for each alcohol and rinsing the glass rod each time. <strong>Note:</strong> Don&#39;t add too much alcohol or jostle the tubes because that can slow down the reaction.</div>
		</li>
		<li data-sub-note-item="3-10">
		<div class="lab-step">Watch carefully after mixing the unknown alcohol with Lucas&#39; reagent to ensure that you can distinguish an immediate positive result from a positive result after a brief delay.</div>
		</li>
		<li data-sub-note-item="3-11">
		<div class="lab-step">Check on the tubes every few minutes and record positive results as you see them. Note approximately how long it took for delayed positive results to appear.</div>
		</li>
		<li data-sub-note-item="3-12">
		<div class="lab-step">Once the tubes have sat for at least 15 min, you can assume that the results are final. Record the results for 1-butanol, 2-butanol, and for your unknown if applicable.</div>
		</li>
		<li data-sub-note-item="3-13">
		<div class="lab-step">Now, let&#39;s clean up. Your instructor will collect the Cr(VI) solutions, so leave any tubes of orange solution from the Jones test in your hood.</div>
		</li>
		<li data-sub-note-item="3-14">
		<div class="lab-step">Empty the other test tubes into the waste beaker and rinse them with deionized water. Pour the waste into the aqueous metal waste container using a funnel and rinse the beaker with deionized water.</div>
		</li>
		<li data-sub-note-item="3-15">
		<div class="lab-step">Now, wash your glassware as usual and dispose of used pipettes in the glass waste. Lastly, clean your hood and throw out any trash in the lab trash container.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="4" data-parent-collapse="">Results<button class="expand">Expand</button>
	<p>Identify your unknown alcohol. The four possibilities are 1-propanol, 2-propanol, 2-methyl-2-butanol, and para-chlorophenol.</p>
	<ul class="lab-items">
		<li data-sub-note-item="4-2">
		<div class="lab-step">Consider the results of the ferric chloride test first. If the mixture with your unknown alcohol turned purple, like phenol did, your alcohol must be para-chlorophenol. If you saw a negative result, your unknown is one of the three aliphatic alcohols.</div>
		</li>
		<li data-sub-note-item="4-3">
		<div class="lab-step">Look at the results of the Jones test. If the mixture with the unknown alcohol turned green, the unknown could be primary or secondary. If the mixture with your unknown alcohol stayed orange, your unknown alcohol is tertiary. Therefore, a negative Jones result means that your alcohol must be 2-methyl-2-butanol.</div>
		</li>
		<li data-sub-note-item="4-4">
		<div class="lab-step">Lastly, look at the results of the Lucas test. If you had a negative result in the Jones test, you should also have seen an immediate positive result in the Lucas test. This confirms that your alcohol was tertiary. If your unknown had a positive result after a short delay, then your unknown alcohol is secondary, and therefore, must be 2-propanol. Finally, if your unknown had a negative result in the Lucas test, your alcohol is primary, and therefore, must be 1-propanol.</div>
		</li>
	</ul>
	</li>
</ol>

Identifying Alcohols: Ferric Chloride Test, Jones Test, and Lucas Test - Procedure

Identificazione degli alcoli

Source: Lara Al Hariri and Ahmed Basabrain at the University of Massachusetts Amherst, MA, USA

	Ferric Chloride Test for PhenolsExpand
	In this lab, you ...

Beer's Law

Beer-Lambert Law and Equilibrium Constant Determination - Procedure

Legge della birra

Source: Smaa Koraym at Johns Hopkins University, MD, USA

	Lab Setup for Beer&#8217;s LawExpand
	In this lab, you will combine aqueous solutions of FeCl3 ...

Concentration Dependence

<p>Source: Smaa Koraym at Johns Hopkins University, MD, USA</p>
<ol class="note-items">
	<li data-note-item="1" data-parent-collapse="">Concentration Dependence<button class="expand">Expand</button>
	<p>In this experiment, you will combine varying concentrations of aqueous HCl and sodium thiosulfate to make solid sulfur, which rapidly gathers into opaque yellow particles at a certain sulfur concentration. Since the reaction solution starts clear and colorless, you can easily tell when it reaches that concentration.</p>
	<p>You&#39;ll use the same test tube and total volume each time, so it will take the same amount of sulfur to make each solution completely opaque. Thus, you&#39;ll measure the reaction progress by timing how long that takes. You&#39;ll then use that data to estimate the reaction orders of the individual reactants and of the overall reaction.</p>
	<p>Before starting the lab, make a table listing the reactant concentrations, the time to solution opacity, and the solution temperature for the trials.</p>
	<h4><strong>Table 1: Reaction progress</strong></h4>
	<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
		<tbody>
			<tr>
				<td><strong>Trial</strong></td>
				<td><strong>[Na<sub>2</sub>S<sub>2</sub>O<sub>3</sub>] (M)</strong></td>
				<td><strong>[HCl] (M)</strong></td>
				<td><strong>Time (s)</strong></td>
				<td><strong>Temp (&#176;C)</strong></td>
			</tr>
			<tr>
				<td style="text-align: center;">1</td>
				<td style="text-align: center;">0.1 M</td>
				<td style="text-align: center;">3.0 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">2</td>
				<td style="text-align: center;">0.1 M</td>
				<td style="text-align: center;">3.0 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">3</td>
				<td style="text-align: center;">0.2 M</td>
				<td style="text-align: center;">3.0 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">4</td>
				<td style="text-align: center;">0.15 M</td>
				<td style="text-align: center;">3.0 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">5</td>
				<td style="text-align: center;">0.05 M</td>
				<td style="text-align: center;">3.0 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">6</td>
				<td style="text-align: center;">0.1 M</td>
				<td style="text-align: center;">6.0 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">7</td>
				<td style="text-align: center;">0.1 M</td>
				<td style="text-align: center;">4.5 M</td>
				<td></td>
				<td></td>
			</tr>
			<tr>
				<td style="text-align: center;">8</td>
				<td style="text-align: center;">0.1 M</td>
				<td style="text-align: center;">1.5 M</td>
				<td></td>
				<td></td>
			</tr>
		</tbody>
	</table>
	<p><a href="https://www.jove.com/CDNSource/lm/labs/50/Table_1._Reaction_progress.xlsx" target="_blank">Click Here to download Table 1</a></p>
	<p>We want to make sure that the reactions are all taking place at room temperature. You&#39;ll perform two benchmark trials, three trials with different sodium thiosulfate concentrations, and three trials with different HCl concentrations. You will vary the concentrations by diluting stock solutions of sodium thiosulfate and HCl, as shown in the following tables.</p>
	<h4><strong>Table 2: Sodium thiosulfate dilutions</strong></h4>
	<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
		<tbody>
			<tr>
				<td style="text-align: center;"><strong>Target Concentration</strong></td>
				<td style="text-align: center;"><strong>Volume of 0.2 M sodium thiosulfate</strong></td>
				<td style="text-align: center;"><strong>Volume of DI water</strong></td>
			</tr>
			<tr>
				<td style="text-align: center;">0.05 M</td>
				<td style="text-align: center;">5 mL</td>
				<td style="text-align: center;">15 mL</td>
			</tr>
			<tr>
				<td style="text-align: center;">0.10 M</td>
				<td style="text-align: center;">10 mL</td>
				<td style="text-align: center;">10 mL</td>
			</tr>
			<tr>
				<td style="text-align: center;">0.15 M</td>
				<td style="text-align: center;">15 mL</td>
				<td style="text-align: center;">5 mL</td>
			</tr>
			<tr>
				<td style="text-align: center;">0.20 M</td>
				<td style="text-align: center;">20 mL</td>
				<td style="text-align: center;">0 mL</td>
			</tr>
		</tbody>
	</table>
	<p><a href="https://www.jove.com/CDNSource/lm/labs/50/Table_2._Sodium_thiosulfate_dilutions.xlsx" target="_blank">Click Here to download Table 2</a></p>
	<h4><strong>Table 3: HCl Dilutions</strong></h4>
	<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
		<tbody>
			<tr>
				<td style="text-align: center;"><strong>Target Concentration</strong></td>
				<td style="text-align: center;"><strong>Volume of 6.0 M HCl</strong></td>
				<td style="text-align: center;"><strong>Volume of DI water</strong></td>
			</tr>
			<tr>
				<td style="text-align: center;">1.5 M</td>
				<td style="text-align: center;">2.5 mL</td>
				<td style="text-align: center;">7.5 mL</td>
			</tr>
			<tr>
				<td style="text-align: center;">3.0 M</td>
				<td style="text-align: center;">5.0 mL</td>
				<td style="text-align: center;">5.0 mL</td>
			</tr>
			<tr>
				<td style="text-align: center;">4.5 M</td>
				<td style="text-align: center;">7.5 mL</td>
				<td style="text-align: center;">2.5 mL</td>
			</tr>
			<tr>
				<td style="text-align: center;">6.0 M</td>
				<td style="text-align: center;">10 mL</td>
				<td style="text-align: center;">0 mL</td>
			</tr>
		</tbody>
	</table>
	<p><a href="https://www.jove.com/CDNSource/lm/labs/50/Table_3._HCl_Dilutions.xlsx" target="_blank">Click Here to download Table 3</a></p>
	<p>Remember to use caution when handling HCl, which is toxic and highly corrosive. Sulfur dioxide, which is a gaseous product of this reaction, is also toxic. You will leave the reaction waste in the fume hood overnight to let the sulfur dioxide escape harmlessly.</p>
	<ul class="lab-items">
		<li data-sub-note-item="1-6">
		<div class="lab-step">First, put on a lab coat, splash-proof safety glasses, and nitrile gloves.</div>
		</li>
		<li data-sub-note-item="1-7">
		<div class="lab-step">Fill a plastic wash bottle with deionized water and get a test tube marked with an X.</div>
		</li>
		<li data-sub-note-item="1-8">
		<div class="lab-step">Clamp the test tube over the center of a small stir plate. The label must be on the opposite side from you. Make sure that you can clearly see the X. Place a small magnetic stir bar in the test tube.</div>
		</li>
		<li data-sub-note-item="1-9">
		<div class="lab-step">Use a thermometer clamp to secure a glass thermometer in the lower third of the tube, keeping it clear of the X and the stir bar.</div>
		</li>
		<li data-sub-note-item="1-10">
		<div class="lab-step">Cut a sheet of about 20 squares of plastic paraffin film from a roll and bring the sheet back to your hood.</div>
		</li>
		<li data-sub-note-item="1-11">
		<div class="lab-step">Lay paper towels in the fume hood as a clean surface for labware to be reused.</div>
		</li>
		<li data-sub-note-item="1-12">
		<div class="lab-step">Label a 600-mL beaker as &#8216;waste&#8217; for the reaction, a 100-mL beaker as &#8216;0.2 M sodium thiosulfate&#8217;, and a 50-mL beaker as &#8216;6 M HCl&#8217;.</div>
		</li>
		<li data-sub-note-item="1-13">
		<div class="lab-step">Fill the 100-mL beaker with 0.2 M sodium thiosulfate from the stock bottle in the dispensing hood.</div>
		</li>
		<li data-sub-note-item="1-14">
		<div class="lab-step">After that, bring the 50-mL beaker and a watch glass to the acid-dispensing hood to obtain 50 mL of 6 M HCl. Cover the filled beaker with the watch glass and bring your portion of HCl back to your hood.</div>
		</li>
		<li data-sub-note-item="1-15">
		<div class="lab-step">Prepare 20 mL of 0.1 M sodium thiosulfate for the first benchmark trial. Use a 10-mL volumetric pipette to measure 10 mL of 0.2 M sodium thiosulfate and dispense it into a 20-mL volumetric flask. Fill the 20-mL volumetric flask with deionized water to the mark.</div>
		</li>
		<li data-sub-note-item="1-16">
		<div class="lab-step">Then, cut a square of plastic paraffin film, seal the flask, and invert it several times to thoroughly mix the solution.</div>
		</li>
		<li data-sub-note-item="1-17">
		<div class="lab-step">Remove the film and pour the solution into the test tube.</div>
		</li>
		<li data-sub-note-item="1-18">
		<div class="lab-step">Prepare 10 mL of 3 M HCl. Pipette 5 mL of 6 M HCl into a 10-mL graduated cylinder. Add deionized water to fill the graduated cylinder to the 10-mL line.</div>
		</li>
		<li data-sub-note-item="1-19">
		<div class="lab-step">Seal the top of the graduated cylinder with plastic paraffin film and thoroughly mix the HCl solution by inverting it several times.</div>
		</li>
		<li data-sub-note-item="1-20">
		<div class="lab-step">Now, start the stir motor and get a reset stopwatch. Confirm that you can see the X through the solution in the test tube.</div>
		</li>
		<li data-sub-note-item="1-21">
		<div class="lab-step">Simultaneously start the stopwatch and pour the 3 M HCl into the test tube.</div>
		</li>
		<li data-sub-note-item="1-22">
		<div class="lab-step">Watch the X through the solution, which will start becoming cloudy when enough sulfur has been produced to saturate it. Stop the stopwatch exactly when the solution becomes so opaque that the X disappears.</div>
		</li>
		<li data-sub-note-item="1-23">
		<div class="lab-step">Record the time in your lab notebook for benchmark trial 1 and then reset the stopwatch. Also, record the temperature of the solution.</div>
		</li>
		<li data-sub-note-item="1-24">
		<div class="lab-step">Now, turn off the stir motor, remove the thermometer, and rinse it with deionized water into the waste beaker.</div>
		</li>
		<li data-sub-note-item="1-25">
		<div class="lab-step">Then, empty the test tube into the waste beaker, retrieve the stir bar with long forceps, and rinse the test tube and stir bar with deionized water.</div>
		</li>
		<li data-sub-note-item="1-26">
		<div class="lab-step">Thoroughly clean the test tube with a test tube brush. Briefly rinse the volumetric flask and graduated cylinder with deionized water as well. Dry everything with paper towels.</div>
		</li>
		<li data-sub-note-item="1-27">
		<div class="lab-step">Repeat the trial in the exact same way for the second benchmark time. If the time is not within 3 - 5 s of the first trial, repeat the benchmark trial until you have two consistent trials.</div>
		</li>
		<li data-sub-note-item="1-28">
		<div class="lab-step">Follow the same overall procedure for each subsequent trial, changing the dilutions as indicated in your table and cleaning your glassware between each trial. Use a 5-mL volumetric pipette as needed when measuring the sodium thiosulfate stock solution in trials four and five.</div>
		</li>
		<li data-sub-note-item="1-29">
		<div class="lab-step">Once you have performed all eight trials, rinse your glassware and tools one last time and neutralize the reaction waste with baking soda. Use caution because sulfur dioxide is one of the reaction products.</div>
		</li>
		<li data-sub-note-item="1-30">
		<div class="lab-step">Transfer the neutralized reaction waste to the designated waste container. Then, pour any remaining sodium thiosulfate solution into its labeled waste container and rinse the beaker with deionized water.</div>
		</li>
		<li data-sub-note-item="1-31">
		<div class="lab-step">Neutralize any remaining HCl with baking soda before flushing it down the drain with tap water.</div>
		</li>
		<li data-sub-note-item="1-32">
		<div class="lab-step">Finally, clean and put away your labware according to your standard practices and throw out any trash remaining in your hood. Your instructor will dispose of the reaction waste later.</div>
		</li>
	</ul>
	</li>
	<li data-note-item="2" data-parent-collapse="">Results<button class="expand">Expand</button>
	<ul class="lab-items">
		<li data-sub-note-item="2-1">
		<div class="lab-step">Now, estimate the reaction order of sodium thiosulfate and HCl. Calculate the starting concentration of sodium thiosulfate in the reaction mixture for trials 1 through 5. Looking at the concentration of sodium thiosulfate and the reaction times, we see that as the concentration doubles, the time to opacity is halved.
		<h4><strong>Table 4: Estimating reaction order sodium thiosulfate</strong></h4>
		<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
			<tbody>
				<tr>
					<td style="text-align: center;"><strong>Trial</strong></td>
					<td style="text-align: center;"><strong>[Na<sub>2</sub>S<sub>2</sub>O<sub>3</sub>] added</strong></td>
					<td style="text-align: center;"><strong>Na<sub>2</sub>S<sub>2</sub>O<sub>3</sub> volume (mL)</strong></td>
					<td style="text-align: center;"><strong>Total volume (mL)</strong></td>
					<td style="text-align: center;"><strong>[Na<sub>2</sub>S<sub>2</sub>O<sub>3</sub>] in mixture </strong></td>
					<td style="text-align: center;"><strong>Time (s)</strong></td>
					<td style="text-align: center;"><strong>Inverse time (s<sup>-1</sup>)</strong></td>
				</tr>
				<tr>
					<td style="text-align: center;">1, 2</td>
					<td style="text-align: center;">0.1</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td style="text-align: center;">3</td>
					<td style="text-align: center;">0.2</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td style="text-align: center;">4</td>
					<td style="text-align: center;">0.15</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td style="text-align: center;">5</td>
					<td style="text-align: center;">0.05</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
			</tbody>
		</table>
		<a href="https://www.jove.com/CDNSource/lm/labs/50/Table_4._Estimating_reaction_order_sodium_thiosulfate.xlsx" target="_blank">Click Here to download Table 4</a></div>
		</li>
		<li data-sub-note-item="2-2">
		<div class="lab-step">Remember that each trial used the same volume, so it took the same amount of sulfur to make each solution opaque. Thus, the reaction is twice as fast when the sodium thiosulfate concentration is doubled.</div>
		</li>
		<li data-sub-note-item="2-3">
		<div class="lab-step">When the concentration increases by a factor of 4, the reaction rate also increases by a factor of 4. This one-to-one relationship suggests that the reaction was first order for sodium thiosulfate. If so, we would expect a plot of rate versus sodium thiosulfate concentration to be linear.</div>
		</li>
		<li data-sub-note-item="2-4">
		<div class="lab-step">We didn&#39;t measure how much sulfur it took to turn the solution opaque, but we assume that it was the same for each trial. This lets us use the inverse of time to opacity as a stand-in for the rate. The slope won&#39;t be equal to k, but that&#39;s OK for this lab.</div>
		</li>
		<li data-sub-note-item="2-5">
		<div class="lab-step">Plot inverse reaction time as a function of sodium thiosulfate concentration. This confirms the linear relationship between sulfur production and thiosulfate concentration. Thus, the reaction seems to be first order with respect to thiosulfate.</div>
		</li>
		<li data-sub-note-item="2-6">
		<div class="lab-step">Use the same method to estimate the reaction order of HCl. First, calculate the starting concentration of HCl in the benchmark trials and in trials 6 through 8.
		<h4><strong>Table 5: Estimating reaction order HCl</strong></h4>
		<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
			<tbody>
				<tr>
					<td style="text-align: center;"><strong>Trial</strong></td>
					<td style="text-align: center;"><strong>[HCl] added</strong></td>
					<td style="text-align: center;"><strong>HCl volume (mL) </strong></td>
					<td style="text-align: center;"><strong>Total volume (mL) </strong></td>
					<td style="text-align: center;"><strong>[HCl] in mixture </strong></td>
					<td style="text-align: center;"><strong>Time (s)</strong></td>
				</tr>
				<tr>
					<td style="text-align: center;">1, 2</td>
					<td style="text-align: center;">0.1</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td style="text-align: center;">3</td>
					<td style="text-align: center;">0.2</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td style="text-align: center;">4</td>
					<td style="text-align: center;">0.15</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
				<tr>
					<td style="text-align: center;">5</td>
					<td style="text-align: center;">0.05</td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
					<td style="text-align: center;"></td>
				</tr>
			</tbody>
		</table>
		<a href="https://www.jove.com/CDNSource/lm/labs/50/Table_5._Estimating_reaction_order_HCl.xlsx" target="_blank">Click Here to download Table 5</a></div>
		</li>
		<li data-sub-note-item="2-7">
		<div class="lab-step">Then, look at the concentration of HCl and the reaction times for these trials. The reaction times are nearly identical, so the HCl concentration has no effect on the reaction rate. This means that HCl seems to be zeroth order for this reaction.</div>
		</li>
		<li data-sub-note-item="2-8">
		<div class="lab-step">Lastly, add up the reaction orders of the reactants to get the overall reaction order.</div>
		</li>
	</ul>
	</li>
</ol>

Concentration Dependence and Reaction Order - Procedure

Dipendenza dalla concentrazione

Source: Smaa Koraym at Johns Hopkins University, MD, USA

	Concentration DependenceExpand
	In this experiment, you will combine varying concentrations of ...

Optimal Foraging

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Simulating Foraging with Artificial Habitats and Prey
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: The foragers will hunt for prey represented by pinto beans in four buckets of rice with varying prey densities. Without knowing what these densities are, foragers must obtain as many prey items as possible in as little time as possible. 
</div></li><li data-sub-note-item="1-2"><div class="lab-step">HYPOTHESES: In this experiment, the experimental hypothesis could be that foragers will catch the most prey in the higher prey density bucket and also spend the most time foraging there.  The null hypothesis would then be that foragers will catch an equal number of prey in each bucket and that foragers will spend an equal amount of time foraging in each patch. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">To begin the foraging activity, record the names and role of each participant at the top of the student data collection sheet.  NOTE: Before starting the experiment, the forager is allowed to become familiar with the foraging process &#8211; the following steps are a practice round. 
</div></li><li data-sub-note-item="1-4"><div class="lab-step">The forager should kneel or sit in front of the average density bucket. 
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Once the timer says start, the forager may begin looking for the pinto beans representing the prey. The forager can spend as much time as they like foraging in this bucket. 
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Once a prey item is located, the forager must signal that one has been found and place the prey in the cup next to the bucket.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Once they believe they have acquired most of the prey in the patch, they should tell the timer to stop the time and the practice round is over. 
</div></li><li data-sub-note-item="1-8"><div class="lab-step">To begin the simulation proper, the forager should sit or kneel in front of bucket A. 
</div></li><li data-sub-note-item="1-9"><div class="lab-step">The timer must be ready to start the stopwatch. 
</div></li><li data-sub-note-item="1-10"><div class="lab-step">The recorder should be ready to record the time each prey item is located, in addition to the time that the forager arrived at and left each bucket. 
</div></li><li data-sub-note-item="1-11"><div class="lab-step">When all parties are ready, the timer should start the stopwatch and say &#8220;start&#8221;,  and the forager should begin foraging. NOTE: The stopwatch will run through the rest of the round for buckets B and C.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">As the forager signals finding pinto beans and places them in the cup next to the bucket, the timer should tell the recorder the time.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">The recorder should then write the time in the student data collection sheet.  <a href="https://www.jove.com/CDNSource/lm/labs/34/Table_1_Optimal_foraging.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Once the forager has decided to move on to the next bucket, they should tell the timer &#8220;done&#8221;, and begin moving to the next patch. 
</div></li><li data-sub-note-item="1-15"><div class="lab-step">The recorder should record the time the forager left and the time they arrived at bucket B in the student data collection sheet.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Repeat the same procedure for buckets B and C. Once the forager has completed the activity, it is time for the next forager to begin using the same procedure. 
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Before performing any data analysis, review the data collection sheet and compile your individual data for captured prey, foraging time, and travel time for each bucket in your student calculations table. <a href="https://www.jove.com/CDNSource/lm/labs/34/Table_2_Optimal_foraging.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="2-2"><div class="lab-step">After this, calculate your individual capture rate for each bucket by dividing the number of prey captured by the sum of the foraging time and travel time and record these values in your table. Individual capture rate =  Number of prey captured / (foraging time + travel time)
</div></li><li data-sub-note-item="2-3"><div class="lab-step">To calculate your Giving-Up-Time or a GUT for each bucket, subtract the time that the last prey item was acquired from the time you left the bucket.  GUT = time forager left bucket  -  time last prey item acquired
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Next, calculate the mean GUT for your foraging group and record the values in your table.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Finally, calculate your scaled GUT for each bucket by dividing the bucket GUT by your group&#39;s mean GUT. Scaled GUT =  Bucket GUT / Group mean GUT NOTE: The scaled GUT will allow class data to be pooled later on without differences in individual foragers skewing the data. 
</div></li><li data-sub-note-item="2-6"><div class="lab-step">The instructor should now compile the data into the classroom spreadsheet.  <a href="https://www.jove.com/CDNSource/lm/labs/34/Table_3_Optimal_foraging.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 3</a>
</div></li><li data-sub-note-item="2-7"><div class="lab-step">Plot a bar graph of the average number of prey collected in each bucket. On the x-axis, A should represent the low prey density bucket, B should represent the medium prey density bucket, and C the high prey density bucket. Plot the y-axis as the number of prey items caught. 
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Next, plot the graph of the class average scaled GUT, where the x-axis represents the buckets with varying levels of prey density and the y-axis shows the values for the average scaled GUT. 
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Plot a third graph of the average time spent foraging in each bucket. 
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Now review the Marginal Value Theorem (MVT) from the concepts video and examine the three graphs that you generated from your class data. Take note of whether the graphs conform to the predictions of MVT. 
</div></li></ul></li></ol>

Optimal Foraging: Foraging with Artificial Habitats and Prey - Procedure

Foraggiamento ottimale

Simulating Foraging with Artificial Habitats and Prey
ExpandNOTE: The foragers will hunt for prey represented by pinto beans in four buckets of rice with ...

Biologia

Ecologia

Animal Behavior

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Making Choice Chambers
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Before beginning the experiments, use one of the Sharpies from the lab table to trace a line around the circumference of the bottom of both bottles. One line should be 1 cm from the base of the bottle and the other should be 1.5 cm from the base of the bottle.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Use a razor blade to make a small cut in each bottle at the mark. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Then, insert the scissors and carefully cut along the line until the bottoms of the bottles are removed completely.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Fit one bottle bottom into the other.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Tape the bottles together to make a secure chamber.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Use the razor blade and scissors to poke a hole into the center of the chamber. 
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Take a piece of tape and fold the edge onto itself to make a tab and place the tape over the hole.  Note: The tab helps with easy removal and reattachment of the tape to the chamber.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Use the Sharpie to label one side of the chamber A and the other side B.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Upon receiving the colony of <em>Drosophila</em>, perform a count to make sure there are 40 living individuals in the vial.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Geotaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">To perform a geotaxis assay, first attach a clean, empty bottle cap to each end of the choice chamber and turn the fly vial upside down onto its lid.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Open the tape flap in the center of the choice chamber.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Insert the funnel into the hole and tap all of the flies into the chamber. HYPOTHESES: The alternate hypothesis is that more flies will be moving against gravity, or up, regardless of the orientation of the choice chamber. Alternatively, if the flies are positively geotactic, then more flies will move down, or with gravity. The null hypothesis is that the flies will have no preference with regard to gravity and will move randomly within the chamber. 
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Once the <em>Drosophila</em> are in the chamber, close the tape flap, rotate the chamber so that it stands vertically with side A on the bottom, and start the timer.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Immediately, count the number of <em>Drosophila</em> that are moving up and the number that are moving down. Record these data for time point 0 in the table. 
</div></li><li data-sub-note-item="2-6"><div class="lab-step">After 30 seconds, rotate the chamber so that side B is on the bottom and count the number of <em>Drosophila</em> that are moving up or down. Record these data for time point 30 in the table.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">After 60 seconds, rotate the chamber so that side A is on the bottom again and record the number of moving flies for time point 60.  <a href="https://www.jove.com/CDNSource/lm/labs/15/Table_1_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Hold the chamber over the fly vial and carefully open the cap to return all of the <em>Drosophila</em> back to their empty vial. 
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Phototaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">To perform a phototaxis analysis, first cover side A of the choice chamber with tin foil, taking care that the foil reaches to the exact center between the two sides. 
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Ensure the caps are secured on both ends of the chamber.  HYPOTHESES: In this exercise, the alternative hypothesis is that more flies will be found in the light side of the choice chamber, if the flues are attracted to light, or on the dark side if they are repelled by light. The null hypothesis is that <em>Drosophila</em> will have no preference with regard to either stimulus. Therefore, equal numbers of flies will be found on either side of the choice chamber.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Open the tape flap in the center of the choice chamber, insert the funnel into the hole, and tap all 40 flies into the chamber as previously described.
</div></li><li data-sub-note-item="3-4"><div class="lab-step">Set the timer to sound an alarm at 10 minutes.
</div></li><li data-sub-note-item="3-5"><div class="lab-step">When the alarm sounds, stop the timer, and count the number of flies in the open side B of the chamber.
</div></li><li data-sub-note-item="3-6"><div class="lab-step">Subtract that number from the total number of flies added to the choice chamber to calculate the number of flies in the covered side A.
</div></li><li data-sub-note-item="3-7"><div class="lab-step">Record these data in the table for time point 10.  <a href="https://www.jove.com/CDNSource/lm/labs/15/Table_2_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="3-8"><div class="lab-step">Carefully, return all of the flies back to their empty vial.
</div></li></ul></li><li data-note-item="4" data-parent-collapse="">Chemotaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="4-1"><div class="lab-step">HYPOTHESES: In this exercise, the alternative hypothesis is that more flies will be found on the side of the chamber with the apple cider vinegar if the flies are attracted to the odor. Or, if the flies are repelled by the vinegar, then more of them will be found on the side of the chamber presenting the odorless water. The null hypothesis is that <em>Drosophila</em> will have no preference with regard to either stimulus and will move randomly in the chamber.  Use an individual disposable plastic pipette to dispense two drops of the apple cider vinegar onto one cotton ball for side A of the chamber. 
</div></li><li data-sub-note-item="4-2"><div class="lab-step">Use an individual disposable plastic pipette to dispense two drops of the distilled water onto a cotton ball for side B of the chamber. 
</div></li><li data-sub-note-item="4-3"><div class="lab-step">Fold two pieces of tape over themselves to create individual pieces of double-sided tape and use the tape to attach each cotton ball to the inside of the single bottle cap.
</div></li><li data-sub-note-item="4-4"><div class="lab-step">Screw the bottle caps with the cotton balls onto their corresponding sides of the chamber.
</div></li><li data-sub-note-item="4-5"><div class="lab-step">Transfer all of the flies into the chamber and set the timer to sound an alarm after 10 minutes. 
</div></li><li data-sub-note-item="4-6"><div class="lab-step">When the alarm sounds, stop the timer and record the number of flies in each side of the chamber. <a href="https://www.jove.com/CDNSource/lm/labs/15/Table_3_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 3</a>
</div></li><li data-sub-note-item="4-7"><div class="lab-step">At the end of the final experiment, funnel the flies into a morgue tube containing 70% ethanol. 
</div></li><li data-sub-note-item="4-8"><div class="lab-step">Dismantle the choice chambers for recycling.  NOTE: Since all of the chemicals used in these experiments are lab safe, the liquids can be disposed of down the sink, the chambers rinsed, and the solids can be thrown away in the trash. 
</div></li></ul></li><li data-note-item="5" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="5-1"><div class="lab-step">Gather all of the data collected for the geotaxis, phototaxis, and chemotaxis experiments.
</div></li><li data-sub-note-item="5-2"><div class="lab-step">Plot these values as bar graphs for different conditions and different time points.
</div></li><li data-sub-note-item="5-3"><div class="lab-step">Use these data to draw inferences about the behavior of <em>drosophila melanogaster</em>. 
</div></li></ul></li></ol>

Animal Behavior: Studying Taxis Behavior of Drosophila - Procedure

Comportamento animale

Making Choice Chambers
ExpandBefore beginning the experiments, use one of the Sharpies from the lab table to trace a line around the circumference of the ...

Extinction

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Diversity and Extinction during the Paleogene Epoch
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Begin this procedure by reading the extinction reading supplement that has been printed out for you. <br /><strong>NOTE:</strong> Alternatively, you may download the reading supplement at the link below.&nbsp; <a href="https://www.jove.com/CDNSource/lm/labs/24/Lab_20_Reading Supplement.docx" style="display:block;margin:20px;" target="_blank">Click here to download the extinction reading supplement</a>
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Using the information contained in the extinction reading supplements pertaining to the taxa of interest and the Chicxulub impact event, make predictions for each taxon listed in Table 1. Consider whether diversity in each taxon increased, decreased, or stayed the same immediately following the Cretaceous-Paleogene boundary and later on in the Paleogene epoch. <a href="https://www.jove.com/CDNSource/lm/labs/24/Table_1_Extinction.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Then, fill out the reasons for each of your predictions in the Contributing Factors column. HYPOTHESES: The experimental hypothesis is that many of the different taxa will show changes in diversity after the Cretaceous-Paleogene event, and again at the end of the Paleogene, with some increasing in diversity, while others decrease. The null hypothesis is that there will be no changes in diversity between taxa following the Cretaceous-Paleogene event, or at the end of the Paleogene.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Gather diversity data by going to the Paleobiology Database: <a href="http://www.paleobiodb.org" target="_blank">www.paleobiodb.org</a>.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">In the center of the screen, click the yellow box labeled Download Data. Under the italicized heading, &#8220;What do you want to download?&#8221;, select Diversity over time on the left side and Comma-separated values on the right side.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Scroll down to the heading &#8220;Select by taxonomy&#8221;.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">In the first box labeled 'Taxon or taxa to include', type in the name of the taxon that data is being gathered on.  NOTE: Ensure the spelling is correct.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Now, go to the dropdown menu right below that box labeled &#8220;Taxonomic resolution&#8221;, and select genera.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Under the next heading, Select by time, type Maastrichtian into the first box and Chattian in the box directly below. Make sure the dropdown menu labeled Time rule is set to major and the Temporal resolution is set to stage.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">Select &#8220;Assume extant taxa range through to present&#8221;. Scroll back up to the middle of the page and click download. A save window will open.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">Save the .csv file to a safe and accessible place and rename it to make it easy to identify.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Open the file and scroll down to the main dataset under Records.  NOTE: The data are arranged in reverse time order by geological stage. The second column, interval_name, gives the accepted name of the stage in question. The next two columns give the appropriate dates of the stage in millions of years before the present. The final column, number of occurrences, gives a total count of the genera extant during that geological stage as determined from occurrence of fossils.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">To more easily work with this data, copy and paste each column into a new sheet. Be sure to label this sheet with the taxon in question, the data and time the data were obtained, and the data source.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Repeat this process for each of the taxa in Table 1 (steps 4 &#8211; 12). NOTE: Data from multiple taxa can be kept on the same sheet, so long as they are kept distinct and labeled appropriately.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">To create line plots for each of the taxa showing the change in diversity over time, select the max_ma column and the number of occurrences column simultaneously.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">In Excel, select Insert from the top toolbar and find the section labeled Charts. Select the bottom center icon, Insert Scatter Chart.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">Then, select the Scatter with Straight Lines and Markers and a graph should appear. NOTE: Since the axis of this graph is in units of millions of years ago, the axis is in the reverse direction of time when put in numerical order.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">To fix this, click on the green plus sign in the top-right corner of the graph and hover over Axes. This should reveal a small black arrow. Click the arrow to open up a small menu and select More Options.
</div></li><li data-sub-note-item="1-19"><div class="lab-step">This will open the Axis Options menu. Under Axis Options, click the box next to Values in reverse order. NOTE: It is also possible to change the axis scales from this menu, which can be used to eliminate the excess time that is not being analyzed in this lab. Keep any changes consistent between different graphs.
</div></li><li data-sub-note-item="1-20"><div class="lab-step">Double-click on the title at the top of the graph and change it to the taxon being represented.
</div></li><li data-sub-note-item="1-21"><div class="lab-step">Make a separate graph for each of the taxa in Table 1. Keep them organized and in one location. A separate sheet for graphs is recommended.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">For each graph, identify the location of the Cretaceous-Paleogene boundary and draw a vertical line through the graph at this point.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Using the information from the graphs, fill out Table 2. In some cases, it may be helpful to go back to the original dataset and compare actual values, particularly when they are small. <a href="https://www.jove.com/CDNSource/lm/labs/24/Table_2_Extinction.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Compare these values to the predictions made in Table 1 and discuss if the original contributing factors are still likely candidates.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Using the dataset, calculate the percentage of total taxa that went extinct over the Cretaceous-Paleogene boundary using this equation.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">There is an event in the middle of the Paleogene known as the Paleocene-Eocene Thermal Maximum about 55.5 million years ago. This represents a high point in temperature during the Paleogene epoch at approximately 8 &#176;C higher than today&#39;s average temperature. Does this point seem to have an effect on diversity for any of the taxa? If yes, then which ones?
</div></li></ul></li></ol>

Extinction: Diversity and Extinction during the Paleogene Epoch - Procedure

Estinzione

Diversity and Extinction during the Paleogene Epoch
ExpandBegin this procedure by reading the extinction reading supplement that has been printed out for ...

Group Behavior

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">The Hawk-Dove Game
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: In this activity, you will play the hawk and dove game with a single partner and then with multiple partners to observe hypothetical costs and benefits of the antagonist or cooperative behavior respectively. 
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Before the activity starts, review the cost-benefit table so that you understand how to score the game. <a href="https://www.jove.com/CDNSource/lm/labs/33/Table_1_Group_behavior.xlsx" style="display:block;margin:20px;" target="_blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="1-3"><div class="lab-step">HYPOTHESES: The experimental hypothesis for this exercise could be that the dove strategy will become more frequent between repeated partners, and the hawk strategy will become more frequent when interactions are with a variety of partners. The null hypothesis could be that there will be no change in strategy over time with repeat or different partner combinations. NOTE: Benefit can be determined by looking at the left column and choosing the row that corresponds to the card you played. The box that corresponds to the choice of your partner can be found in the top row. For the purposes of this game, B equals four and C equals three. So depending on the cards played, player one here, which is you, would gain a score of two, four, zero or one half. 
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Collect a set of two game cards. 
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Take out the cards labeled hawk and dove and place them face down on the table or conceal them in your hands. 
</div></li><li data-sub-note-item="1-6"><div class="lab-step">When the instructor gives the signal show either of the two cards to the student sitting across from you. Try to strategize how to receive the highest benefit from every interaction on your own but do not communicate with your partner. 
</div></li><li data-sub-note-item="1-7"><div class="lab-step">In the Single Partner Recording Table (Table 2) provided by your instructor, note the cards played as well as the benefit you received for that round.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Repeat showing cards to the same partner and determining benefit on your instructor&#39;s signal for a total of 15 rounds.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">At the end of those rounds, the student with the highest total benefit value was determined to be the &#8220;winner&#8221;, or the most fit.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">As a class, determine the percentages of hawks and doves played as well as the average benefit received for each round, using this formula: Class average benefit=(sum total benefit)/(number of players)
</div></li><li data-sub-note-item="1-11"><div class="lab-step">For the second game, and then for each round of play, one row of students should rotate to their right, leaving all players with a new partner.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Continue playing for 15 rounds or until students end up back in their starting positions, recording the data in the Multiple Partner Recording Table (Table 3) given to you by your instructor.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">For each round, determine the percentages of hawks and doves played that round, as well as the average benefit received.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Cheating and Cooperation Exercise
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">NOTE: In this activity, you will exchange benefits, represented by colored or similar items, with your classmates. In this simulation, a cheater is defined as a person who gives fewer benefits than they receive. During this activity, you will observe the frequency of cheating behavior over time. 
</div></li><li data-sub-note-item="2-2"><div class="lab-step">HYPOTHESES: The experimental hypothesis is that cheaters will receive fewer benefits over time and cooperators will continue to provide equal benefits to one another. The null hypothesis is that the frequency of cheaters and cooperators does not change over time. 
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Each person should collect beads of one color to represent a tangible benefit, as well as one paper bag and a three-ring binder.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Get into a group of three so that all members have objects of the same color.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Now find another group with a different color to pair with. In each round, partners will exchange benefits without knowing what they are receiving from their partner and without discussion within their group.  NOTE: Benefits of a different color from your starting color are worth twice as much.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Turn face to face with a member from the other group while still being able to conceal the objects in your possession.  TIP:<strong> </strong>Setting up a screen using a notebook or sitting behind a desk can be helpful here.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">When given a signal from the instructor, place zero to two benefits into the paper bag and pass it to your partner.
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Simultaneously, receive the bag from your partner and then remove the benefits from the bag.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Make a note of how many benefits you gave away and how many you received in the Cheating and Cooperation Table (Table 4) provided by your instructor.
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Turn to a new member from the other group and repeat the exchange.
</div></li><li data-sub-note-item="2-11"><div class="lab-step">Rotate through all of the members of the other group for as many rounds as the instructor determines - which will not be revealed until the game ends so as not to alter player behavior.
</div></li><li data-sub-note-item="2-12"><div class="lab-step">Once the game has ended, note for each round in the cheating and cooperation table whether you gave more than your partner, received more from them or if you gave the same amount.
</div></li><li data-sub-note-item="2-13"><div class="lab-step">Now total up all of the benefits in your possession, with benefits you&#39;ve received from the other group being worth twice as much.
</div></li><li data-sub-note-item="2-14"><div class="lab-step">For each round, the whole class should total up the number of cheaters - with cheaters defined as those who gave less than they received.
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">For both the single partner hawk-dove game and the multiple partners hawk-dove game, use the whole class data to plot the percentages as two separate lines - one for the percentage of hawks, and one for the percentage of doves, for each of the 15 rounds.
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Take note of whether the cards played changed as more rounds were played.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Manipulate the values for B and C to see if you can find the point at which values of B and C would render the hawk strategy no longer effective (this would be the point when there is no benefit to choosing the hawk strategy over always choosing the dove strategy).
</div></li><li data-sub-note-item="3-4"><div class="lab-step">As a class, use the data from the cheating and cooperation activity to make a graph showing the percentage of rounds each individual player cheated versus the amount of benefits they ended with.
</div></li><li data-sub-note-item="3-5"><div class="lab-step">Consider whether it is better to be a cheater or a cooperator in this scenario and discuss as a class what makes this scenario different from the hawk-dove game.
</div></li><li data-sub-note-item="3-6"><div class="lab-step">Now, use the class data to graph the percentage of cheaters against the number of rounds, and look for any trends over time.
</div></li><li data-sub-note-item="3-7"><div class="lab-step">Consider which type of strategy would most likely evolve in a setting where partners interact repeatedly.
</div></li><li data-sub-note-item="3-8"><div class="lab-step">Then, note which tactic might be beneficial for a species that relies on cooperation. 
</div></li><li data-sub-note-item="3-9"><div class="lab-step">Discuss as a class what would likely happen if you increased or decreased the relative value of the benefits you receive.
</div></li></ul></li></ol>

Group Behavior: Hawk-Dove Game and Trading Exercise - Procedure

Comportamento di gruppo

The Hawk-Dove Game
ExpandNOTE: In this activity, you will play the hawk and dove game with a single partner and then with multiple partners to observe ...

Artificial Selection

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Observing Artificial Selection in Plants 
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">To begin the experiment, collect five two-week old grown plants from the generation one population that appear healthy. Note: Plants should appear green, not wilted, and not have damaged leaves. Experimental Hypothesis: The experimental hypothesis for this lab might be that the number of lead trichomes varies naturally, but the average number will increase over generations if only plants with the highest number of trichomes are interbred or decrease if those with the least are interbred. Null Hypothesis: The null hypothesis might be that the average number of trichomes will not change over subsequent generations after artificial selection.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Label each plant with a letter from A to E and with both partners&#8217; names.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Record any natural differences you see between your plant samples. Note: All of the plants are the same age, so any observed variation is due to expressed phenotype, not development.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Use a hand lens to count the number of trichomes found on the petiole of the lowest leaf, not including the round, first-grown cotyledons for each plant. Note: It can be helpful to shine a light onto the plant and to hold it against a dark background, such as a lab bench, during the trichome analysis. 
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Both partners will conduct counts on each plant independently and compare answers. If the counts are different, enlist a third counter or take the average of the two results.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">After this, determine the median number of trichomes for the entire class.  <a target="__blank" style="display:block;padding:20px;" href="https://www.jove.com/CDNSource/lm/labs/2/Lab_2_Table_A.xlsx">Click Here to download Table A</a>
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Determine the top and bottom 10% of the plants from the class trichome count. These groups will make up the two artificially selected lines: high hairiness and low hairiness, respectively.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">If more than 10% of the plants have no trichomes, select 10% of those plants randomly for the low hairiness plant line.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Separate the two groups of selected plants and label them clearly.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">Move the plants to a well-lit area and keep them well watered until they develop flowers. Note: Flowers typically develop around 14 days.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">When the flowers are fully developed, use a pollination wand or paintbrush to rub the stigma and anther of one plant&#8217;s flowers for a few seconds.
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Using the same wand or brush, rub the inside of each flower of every other plant in the same selection line.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">Using a different wand or brush, repeat steps 11 through 12 on the other line. Note: Take care not to cross the lines. Spread the pollen only among the appropriately selected plants.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Return the generation one plants to the growing areas. When the plants reach approximately 1 month old, they should produce mature seed pods.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">Allow the plants to dry out once the seed pods turn yellow.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Break the seed pods open and collect the seeds.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">Obtain new pots for the generation two seeds.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">Fill the pots with moist potting soil about halfway and add a fertilizer pellet to each pot.
</div></li><li data-sub-note-item="1-19"><div class="lab-step">Fill the pots with more soil to fill just below the lip.
</div></li><li data-sub-note-item="1-20"><div class="lab-step">Make a shallow depression in the soil of each pot with a probe.
</div></li><li data-sub-note-item="1-21"><div class="lab-step">Touch a clean toothpick to a gluestick and use the toothpick to transplant the seeds into the soil one by one.
</div></li><li data-sub-note-item="1-22"><div class="lab-step">Cover the seeds with a thin layer of soil.
</div></li><li data-sub-note-item="1-23"><div class="lab-step">Label the pots with their hairiness line and place them in the designated growing area.
</div></li><li data-sub-note-item="1-24"><div class="lab-step">Use a plastic dropper to water the plants as needed to ensure they do not dry out.
</div></li><li data-sub-note-item="1-25"><div class="lab-step">When the plants are two weeks old, collect five plants from each line and label them A to E.
</div></li><li data-sub-note-item="1-26"><div class="lab-step">Then, count the trichomes on these plants as previously described (steps 4- 5).
<a target="__blank" style="display:block;padding:20px;" href="https://www.jove.com/CDNSource/lm/labs/2/Lab_2_Table_B.xlsx">Click Here to download Table B</a>
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Data Analysis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Create a frequency graph based on the generation one class data with the trichome count on the x axis and the number of plants on the y axis. Label the mean. 
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Then, create frequency graphs for both of the selected hairiness lines.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Compare the frequency graphs for generation one and both hairiness lines.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Take note of phenotypic shifts observed in the selected hairiness lines.
</div></li></ul></li></ol>

Artificial Selection: Visualization in Plants and Data Analysis - Procedure

Selezione artificiale

Observing Artificial Selection in Plants 
ExpandTo begin the experiment, collect five two-week old grown plants from the generation one population that ...

Evoluzione

Natural Selection

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Simulating Natural Selection
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">To begin, cut the white, black, and gray pipe cleaners into quarters, which should produce a total of 100 pieces that are 3 inches long.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Then, cut the green pipe cleaners in the quantities and lengths specified in the video, taking care to minimize waste. This approximates a normal distribution of lengths and should result in an additional 100 pieces total. NOTE: One 12-inch pipe cleaner should yield multiple lengths, for example, a 5-inch piece and a 7-inch piece. Use this advantage to reduce waste.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Now, cut one sheet of black and one sheet of white construction paper into approximately 3-inch strips measured along the long edge of the paper.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Tape these strips down to cover the base of a dissecting tray to create an alternating pattern of black and white strips. NOTE: In the first simulation, you will use the white, gray, and black pipe cleaner segments against a black and white background to simulate different selection pressures due to predation on a population. You will select your prey using two different strategies, eyes-open and eyes-closed. For each of these strategies, you will perform four rounds consisting of 25 selections, alternating between two partners. After each round you will restore the population back to its original size using the proportions of phenotypes present after selection.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">To begin the first scenario, count out 12 white, 12 black, and 26 gray pipe cleaner segments with your partner. Hypotheses: Here the experimental hypothesis might be that different predation strategies, such as having the eyes open or closed, will result in distinct selection pressures on the populations. Further, different phenotypes may be positively selected in the alternate scenarios. The null hypothesis might be that different predation strategies will have no effect on the populations or phenotypes.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Mix the pipe cleaner segments together as best as possible. Then, sprinkle them into the papered tray. NOTE: Ideally, the pipe cleaners will be spread out. However, this may not occur due to their self-adhering nature.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Once the segments are in the tray, separate them as best as possible without moving them too far from where they landed. As few segments as possible should be touching.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Have the first partner close their eyes. When given an audible signal by the second partner, the first should open their eyes and grab the first pipe cleaner they see from the tray as quickly as possible.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Set the removed pipe cleaner to the side.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">Alternating partners, repeat this process a total of 25 times (steps 8 &#8211; 9).
</div></li><li data-sub-note-item="1-11"><div class="lab-step">Count the remaining pipe cleaners by color and record this in the data collection table for Scenario 1. <a href="https://www.jove.com/CDNSource/lm/labs/23/Lab_23_Tables_1_and_2.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Tables 1 and 2</a>
</div></li><li data-sub-note-item="1-12"><div class="lab-step">Then, restore the population to 50 by adding one pipe cleaner of the same color for each of the remaining pipe cleaners. Hold these to the side until they are all gathered.
</div></li><li data-sub-note-item="1-13"><div class="lab-step">Then, sprinkle the pipe cleaners into the tray as previously described (steps 6-7). Repeat this process for three more rounds consisting of 25 selections each and record the counts for each of the rounds in the data collection table for Scenario 1.
</div></li><li data-sub-note-item="1-14"><div class="lab-step">Remove all pipe cleaners from the tray after the fourth round.
</div></li><li data-sub-note-item="1-15"><div class="lab-step">For Scenario 2, again count out 12 white, 12 black, and 26 gray pipe cleaner segments and sprinkle them randomly into the tray.
</div></li><li data-sub-note-item="1-16"><div class="lab-step">Follow the same protocol as in Scenario 1. However, when given the signal, the grabbing partner should keep their eyes closed. To aid in accuracy, it may help to position the hand near to the tray before grabbing.
</div></li><li data-sub-note-item="1-17"><div class="lab-step">Repeat this process 25 times, switching partner roles, and record the remaining pipe cleaner amounts in the data collection table for Scenario 2.
</div></li><li data-sub-note-item="1-18"><div class="lab-step">Then, restore the population to 50 based on that count. Do this for all four rounds. NOTE: In the second simulation, you will be using the eyes-open and eyes-closed predation strategies as previously described to select pipe cleaners of various lengths.
</div></li><li data-sub-note-item="1-19"><div class="lab-step">For Scenario 3, gather all 100 of the pipe cleaners that have been cut to varying lengths to represent different phenotypes.
</div></li><li data-sub-note-item="1-20"><div class="lab-step">One partner should hold all the pipe cleaners upright and loosely in one hand resting on the tabletop, such that the bottom end of each pipe cleaner rests against the tabletop.
</div></li><li data-sub-note-item="1-21"><div class="lab-step">Now, the partner that is not holding the pipe cleaners should close their eyes. When given an audible signal, the partner should open their eyes and grab the first pipe cleaner they see from their partner&#39;s hand.
</div></li><li data-sub-note-item="1-22"><div class="lab-step">Repeat this process 50 times (step 21). However, the partners should not switch roles during the round this time. The partner holding the pipe cleaners should always be the one giving the signal and never grabbing from the bunch.
</div></li><li data-sub-note-item="1-23"><div class="lab-step">After the first round consisting of 50 selections, count the remaining pipe cleaners by length. Then, record this information in the data collection table for scenario three. <a href="https://www.jove.com/CDNSource/lm/labs/23/Lab_23_Tables_3_and_4.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Tables 3 and 4</a>
</div></li><li data-sub-note-item="1-24"><div class="lab-step">Add one pipe cleaner of the same length as each of the remaining ones to restore the population to 100. This will likely require more pipe cleaners than were originally cut. Try to use scraps from the class to reduce waste.
</div></li><li data-sub-note-item="1-25"><div class="lab-step">Repeat this for three more rounds, alternating which partner holds or grabs each round.
</div></li><li data-sub-note-item="1-26"><div class="lab-step">To perform Scenario 4, gather the varied-length pipe cleaners in their original proportions.
</div></li><li data-sub-note-item="1-27"><div class="lab-step">Now repeat the grabbing process, as described in Scenario 3. However, only remove 30 pipe cleaners instead of 50.
</div></li><li data-sub-note-item="1-28"><div class="lab-step">Divide the count at the end of each round by two for each phenotype 4 inches or below, rounding fractions down.
</div></li><li data-sub-note-item="1-29"><div class="lab-step">Then, record these values in the data recording table for Scenario 4.
</div></li><li data-sub-note-item="1-30"><div class="lab-step">Use the relative proportions of the phenotypes in the final population to restore the population to 100. When restoring the population to 100, it may be necessary to calculate relative proportions of phenotypes.
</div></li><li data-sub-note-item="1-31"><div class="lab-step">Repeat this for three more rounds, again alternating holding and grabbing the pipe cleaners.
</div></li><li data-sub-note-item="1-32"><div class="lab-step">For each scenario calculate the survival for each phenotype, that is the proportion of the phenotype surviving.
</div></li><li data-sub-note-item="1-33"><div class="lab-step">Then, calculate the relative fitness, or w, between each round. As reproductive rate does not vary in this simulation, this should always be considered equal, or 1.
</div></li><li data-sub-note-item="1-34"><div class="lab-step">Also, calculate the selection coefficients for each phenotype.
</div></li><li data-sub-note-item="1-35"><div class="lab-step">Now, use the relative frequencies of each phenotype to create histograms with appropriately labeled axes for each scenario showing the population before any selection took place and then alongside these in a different color after four rounds of selection.
</div></li><li data-sub-note-item="1-36"><div class="lab-step">Identify the type of selection taking place in each scenario and label the graphs. 
</div></li><li data-sub-note-item="1-37"><div class="lab-step">Develop a hypothesis to explain your observations. 
</div></li></ul></li></ol>

Natural Selection: Calculating Fitness and Selection - Procedure

Selezione naturale

Simulating Natural Selection
ExpandTo begin, cut the white, black, and gray pipe cleaners into quarters, which should produce a total of 100 pieces that ...