This work was supported by a CIHR operating grant (MOP-102624) to Q. Y. We thank Dr Carolyn Harley for helpful discussions throughout the study, Dr. Qinlong Hou, Amin Shakhawat, and Andrea Darby-King for technical support.

Sono utilizzati Sprague Dawley di ratto (Charles River) cuccioli di entrambi i sessi. Cucciolate sono abbattuti a 12 su PD1 (nascita essendo PD0). Le dighe sono mantenuti su una 12 ore di luce / buio ciclo con ad libitum l&#39;accesso al cibo e all&#39;acqua. Procedure sperimentali sono stati approvati dal Comitato Istituzionale Animal Care del Memorial University. 1 Nose Plug costruzione NOTA:. Questa procedura è stato adattato e modificato da Cummings et al (1997) 35. <ol><li> Aquire polietilene-20 tubo e filo di sutura di seta 3-0. </li><li> Tagliare un piccolo pezzo di polietilene-20 tubo di circa 0,8 mm. </li><li> Discussione sutura di seta attraverso la tubazione disposta in modo tale che vi sia filo su entrambi i lati della sezione del tubo. </li><li> Su una estremità del filo esterno del tappo, un nodo nel filo. </li><li> Estrarre la sezione della tubazione giù sopra il nodo nel thread. Il nodo dovrebbe presentare all&#39;interno del tubo.</li><li> Tagliare le due estremità del filo in modo tale che ~ 2 mm filo sporge da un&#39;estremità del tubo (vedere Figura 1A). </li></ol> 2 Naris occlusione prima di Formazione <ol><li> Rimuovere pup dalla diga e mettere in un piatto coperto custodito con biancheria da letto regolari. </li><li> Utilizzare un&#39;applicazione punta di cotone per tamponare una gelatina anestetico locale, 2% Xylocaine, sulle narice da occluso. </li><li> Lasciare il cucciolo di riposare nel piatto per ~ 3 min. </li><li> Tenere il cucciolo delicatamente ma saldamente in mano non dominante. </li><li> Usando la mano dominante, prendere un tappo il naso e tamponare la stessa pappa anestetico locale intorno alla punta da cui il filo non sporga. Questo agirà sia come anestetico per eventuali dolori lievi associati con l&#39;inserimento spina e agirà come un lubrificante all&#39;interno della narice. NOTA: L&#39;effetto di Xylocaine inizia entro un paio di minuti e dura 20-30 minuti. Pups in generale mostrano una buona tolleranza per l&#39;inserimento spina dopo l&#39;applicazione gelatina Xylocaine (minimum lottare e vocalizzazione). </li><li> Inserire delicatamente la spina naso tenendo saldamente il cucciolo e ruotando lentamente la spina con spinte molto dolci fino a quando la spina sia completamente inserita e solo il thread 2 millimetri sporge dalla narice (vedi Figura 1B). Non ci dovrebbero essere sanguinamento da entrambi narice durante questo processo. Sono esclusi e restituiti dighe Pups con sanguinamento durante l&#39;inserimento spina naso </li><li> Consentire agli animali di riposare in questo piatto per 5 minuti in modo da abituare alla spina. </li><li> Rimuovere il cucciolo dal piatto assuefazione e iniziare il paradigma di condizionamento 24. </li></ol><img alt="Figura 1" fo:content-width="5in" src="/files/ftp_upload/51808/51808fig1highres.jpg" width="500" /> Figura 1 Costruzione di un tappo naso. A) Schema che mostra le fasi di fare una spina naso. Un filo è tirato attraverso il tubo di polietilene; un nodo è fatto e PULportato al centro del tubo per bloccarlo; due estremità sono tagliate con un residuo di 2 mm su un&#39;estremità del tubo. B) Vista frontale e laterale di un ratto con una spina naso in una narice. 3. Profumato Bedding Preparazione <ol><li> Indossando guanti nuovi e in una cappa aspirante per evitare la contaminazione odori, inserire 500 ml di biancheria da letto di trucioli di legno in un sacchetto di plastica. </li><li> Utilizzare una siringa per elaborare 0,3 ml di estratto di menta piperita, e spruzzare questo sopra la biancheria nel sacchetto di plastica. </li><li> Legare la borsa chiusa, agitare vigorosamente il sacchetto, e lasciare la biancheria da letto per riposare in borsa per 5 min. </li><li> Posizionare la biancheria profumata in modo chiaro, poco profondo, scatola acrilica di formazione (20 x 20 x 5 cm 3, Figura 2A) scoperto in una cappa aspirante per 5 minuti prima dell&#39;uso. Una volta che la biancheria da letto viene preparato, scartare questi guanti, e non permettere questi guanti vengano a contatto con gli animali. </li><li> Posizionare la biancheria profumata in una scatola di plastica trasparente identici, e enAssicurarsi che non entri in contatto con la biancheria profumata o guanti utilizzati. </li></ol> 4. Odore condizionata Paradigm (Vedi immagine in figura 2A) Pups subiscono una singola sessione di condizionamento, il PD 6, o sessioni di prove multiple (una seduta al giorno, PD 3-6). <ol><li> Posizionare il cucciolo abituato a biancheria da letto profumato. Per il controllo degli odori solo (O / S -) cuccioli, lasciare questi cuccioli sulla biancheria da letto per 10 minuti, quindi passare al punto 4.5. Per la corsa sperimentale odore + (O / S +) cuccioli, continuano i seguenti passaggi in questa sezione. </li><li> Stroke il cucciolo per 30 secondi con un piccolo pennello. Usa movimenti circolari rapidi principalmente intorno alla regione posteriore del cucciolo. </li><li> Lasciare il cucciolo a riposare per 30 sec. </li><li> Ripetere i punti 4.2 e 4.3 per un totale di 10 min (cioè 10 coppie di carezze + odore). </li><li> Rimuovere pup dalla casella di condizionamento, rimuovere il tappo il naso e tornare il cucciolo alla diga. </li></ol> &lt;p class = &quot;jove_title&quot;&gt; 5. Lateralizzato Preferenza Odore Testing (Vedi immagine in figura 2B) Test avviene in vari momenti (ad esempio, 24 o 48 ore) a seguito della sessione di allenamento finale. Il test viene effettuato in una camera di acciaio inossidabile test (30 x 20 x 18 cm 3), che è posto sulla parte superiore di due scatole di formazione (training casella è descritto in 3.4), separate da una zona neutra 2 cm. Una scatola di formazione contiene biancheria da letto di menta profumata, mentre l&#39;altra scatola contiene pulita, biancheria da letto inodore. Il pavimento della camera di prova è una griglia metallica, che viene poi ricoperto da un foglio rimovibile di rete plastica (Figura 2B). <ol><li> Preparare una menta piperita e una biancheria profumata ai sensi dell&#39;articolo 3, e porre ogni casella sotto lati opposti della camera di prova, 2 cm l&#39;uno dall&#39;altro. Inserire la rete plastica sul pavimento metallo-griglia della camera di prova. </li><li> Rimuovere il cucciolo dalla diga e mettere una piccola quantità di grasso inodore ferma al silicone sulle nariceche è occluso durante l&#39;allenamento. Re-applicare il grasso durante la prima procedura di test come necessario. NOTA: naris occlusione casuale durante l&#39;allenamento e le prove possono essere considerate per evitare distorsioni. </li><li> Posizionare pup in zona neutra della camera di prova. </li><li> Lasciare il cucciolo di esplorare la camera per 1 minuto, registrando per quanto tempo il cucciolo ha speso sopra i due lati della camera (cioè più di menta piperita o di letti neutro profumato). </li><li> Lasciare il cucciolo a riposare per 1 minuto in una camera di contenimento di plastica coperto. </li><li> Ripetere i punti 5.2 e 5.3 per un totale di 10 min (cioè 5 prove di laboratorio, separati da 5 studi di riposo) commutazione l&#39;orientamento iniziale del cucciolo nella camera per controllare per le preferenze di direzione. </li><li> Subito dopo il test, rimuovere il grasso dalla narice. </li><li> Inserire una noseplug polietilene nella narice opposta di cui al punto 2 e consentire agli animali di riposare per 10 min. </li><li> Testare il cucciolo ancora una volta come in<html> 5.3 -5.6, rimuovere il tappo, e tornare il cucciolo alla diga. Rimuovere e pulire la rete di plastica della camera di test con etanolo al 95% e che il liquido evapori. Posizionare la maglia indietro prima di testare il prossimo cucciolo. NOTA: Applicare grasso al silicone al primo occlusione narice durante i test previene il rischio di sanguinamento e di stress associato con l&#39;inserimento spina naso. </li></ol><img alt="Figura 2" fo:content-width="5in" src="/files/ftp_upload/51808/51808fig2highres.jpg" width="500" /> Figura 2 odore precoce preferenza di formazione e test. A) la formazione degli odori precoce preferenza utilizzando odore + accarezzare paradigma. B) Due scelta odore test preferenza con biancheria da letto di menta piperita su un lato, il controllo di letti inodore sul lato opposto. Una zona neutra 2 cm è messo in mezzo. 6. testare l&#39;efficacia del singolo Naris Occlusione nt &quot;&gt; Questo esperimento viene eseguito per determinare se l&#39;occlusione singola narice porta all&#39;attivazione lateralizzato del sistema olfattivo. <ol><li> Eseguire occlusioni Naris unilaterali sul PD 6 o 7 cuccioli come descritto nella sezione 2. </li><li> Dopo ~ 5 min assuefazione, mettere il cucciolo in un contenitore di plastica coperto ed esporlo a 30 ml di olio di menta piperita puro imbevuti di un pezzo di tessuto per 10 min. </li><li> Subito dopo l&#39;esposizione odore menta piperita, iniettare il cucciolo per via intraperitoneale (ip) con cloralio idrato (400 mg / kg) come anestetico generale o pentobarbitol, (150 mg / kg). </li><li> Una volta completamente anestetizzato (mostrando alcuna risposta alla coda o il piede pizzico), transcardiaca profumato il cucciolo con le soluzioni di ghiaccio-freddo di soluzione fisiologica (0,9%) per ~ 1 min, seguito da paraformaldeide (4%, disciolto in soluzione tampone fosfato 0.1 M, PBS ). </li><li> Dopo 10 min di paraformaldeide perfusione, raccogliere il cervello e metterlo in paraformaldeide notte a 4 ° C, poi transfer il cervello di una soluzione di saccarosio (20% in PBS) per altre 24 ore. </li><li> Tagliare le fette di cervello coronali a 30 micron di spessore, con un criostato. Raccogliere OB e PC fette e montare su vetrini rivestiti di gelatina, seguita da colorazione immunoistochimica standard per pCREB anticorpi 21,25,30. </li></ol> 7 Verifica del Reversibilità del singolo Naris Occlusione Questo esperimento test se l&#39;effetto di blocco è reversibile a 24 ore dopo la rimozione del tappo naso. <ol><li> Eseguire occlusioni Naris unilaterali sul PD 6 o 7 cuccioli come descritto nella sezione 2. </li><li> Dopo 15 min (equivalente alla durata dell&#39;occlusione narice durante l&#39;allenamento - 5 min assuefazione + 10 min di formazione), rimuovere il tappo naso, e restituire il cucciolo alla diga. </li><li> 24 ore più tardi, esporre il cucciolo per l&#39;odore di menta piperita in un contenitore di plastica coperto per 10 minuti come descritto al punto 6.2. </li><li> Seguire le stesse operazioni in settaioni 6,3-6,6. </li></ol>

<ol>
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W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=What+a+nostril+knows:+olfactory+nerve-evoked+AMPA+responses+increase+while+NMDA+responses+decrease+at+24-h+post-training+for+lateralized+odor+preference+memory+in+neonate+rat.">What a nostril knows: olfactory nerve-evoked AMPA responses increase while NMDA responses decrease at 24-h post-training for lateralized odor preference memory in neonate rat.</a> Learn Mem. 19, 50-53 (2012).</li><li>Schwob, J. E., Price, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+development+of+axonal+connections+in+the+central+olfactory+system+of+rats.">The development of axonal connections in the central olfactory system of rats.</a> J Comp Neurol. 223, 177-202 (1984).</li><li>Kucharski, D., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+routes+to+early+memories.">New routes to early memories.</a> Science. 238, 786-788 (1987).</li><li>Kucharski, D., Johanson, I. B., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unilateral+olfactory+conditioning+in+6-day-old+rat+pups.">Unilateral olfactory conditioning in 6-day-old rat pups.</a> Behav Neural Biol. 46, 472-490 (1986).</li><li>Cummings, D. M., Henning, H. E., Brunjes, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+bulb+recovery+after+early+sensory+deprivation.">Olfactory bulb recovery after early sensory deprivation.</a> J Neurosci. 17, 7433-7440 (1997).</li><li>Kucharski, D., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+change+in+the+access+to+olfactory+memories.">Developmental change in the access to olfactory memories.</a> Behav Neurosci. 102, 340-348 (1988).</li><li>Brunjes, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unilateral+odor+deprivation:+time+course+of+changes+in+laminar+volume.">Unilateral odor deprivation: time course of changes in laminar volume.</a> Brain Res Bull. 14, 233-237 (1985).</li><li>Kass, M. D., Pottackal, J., Turkel, D. J., McGann, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Changes+in+the+neural+representation+of+odorants+after+olfactory+deprivation+in+the+adult+mouse+olfactory+bulb.">Changes in the neural representation of odorants after olfactory deprivation in the adult mouse olfactory bulb.</a> Chem Senses. 38, 77-89 (2013).</li><li>Kim, H. H., Puche, A. C., Margolis, F. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Odorant+deprivation+reversibly+modulates+transsynaptic+changes+in+the+NR2B-mediated+CREB+pathway+in+mouse+piriform+cortex.">Odorant deprivation reversibly modulates transsynaptic changes in the NR2B-mediated CREB pathway in mouse piriform cortex.</a> J Neurosci. 26, 9548-9559 (2006).</li><li>Korol, D. L., Brunjes, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+changes+in+2-deoxyglucose+uptake+and+amino+acid+incorporation+following+unilateral+odor+deprivation:+a+laminar+analysis.">Rapid changes in 2-deoxyglucose uptake and amino acid incorporation following unilateral odor deprivation: a laminar analysis.</a> Brain Res Dev Brain Res. 52, 75-84 (1990).</li><li>Leung, C. H., Wilson, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trans-neuronal+regulation+of+cortical+apoptosis+in+the+adult+rat+olfactory+system.">Trans-neuronal regulation of cortical apoptosis in the adult rat olfactory system.</a> Brain Res. 984, 182-188 (2003).</li></ol>

The authors have nothing to disclose.

L&#39;apprendimento e la memoria odore modello lateralizzato in cuccioli di ratto all&#39;interno di una finestra temporale critica è stata fondata da Hall e colleghi. In una serie di studi 33,34,36, hanno mostrato che una memoria preferenza odore potrebbe essere lateralizzato da odori + latte abbinamenti a una narice al PD 6 cuccioli di ratto. Memoria preferenza è stata forte quando la stessa narice era aperto durante l&#39;allenamento e test, ma non osserva quando la narice occlusa era sbloccato e testato...

L&#39;olfatto è la modalità sensoriale primaria nei roditori, senza il quale non sarebbero in grado di navigare con successo o di sopravvivere nel loro ambiente. E &#39;particolarmente critico per cuccioli neonati, che non possono né vedere, né udire durante la prima settimana post-natale, per usare l&#39;olfatto al fine di individuare la loro madre per alimentare 1. Come risultato, ratti neonatali possono essere condizionati a preferire gli odori con semplici manipolazioni sperimentali. Una varietà di stimoli sono stati utilizzati come stimolo incondizionato (UCS) per indurre risposte condizionate a nuovi odori (stimolo condizionato, CS) nei neonati, tra cui l&#39;ambiente di nidificazione 2,3, latte lattante 4-6, accarezzando o stimolazione tattile 7- 12, tail pinch 13, saliva materna 13, lieve scossa piede 14-18, e la stimolazione cerebrale intracranica 19. Il presente studio si avvale di un odore precoce preferenza paradigma consolidato in cui un odore, in questo caso menta piperita, is combinata con stimolazione tattile per produrre una preferenza per piperita 24 ore dopo 10,11,20. Questi odori ricordi dipendono dal circuito olfattivo intatto, tra cui in primo luogo il bulbo olfattivo (OB) 21-23 e la corteccia piriforme anteriore (APC) 24,25. Indagini sperimentali dei primi preferenza odore di apprendimento hanno approfondito e ampliato la nostra comprensione delle basi molecolari e fisiologiche di una memoria mammiferi. Questo modello mammiferi ha diversi vantaggi nello studio dei meccanismi della memoria. In primo luogo, sono state individuate le sorgenti neurali del segnale UCS. Vari stimoli come detto sopra stimolano locus coeruleus noradrenalina rilascio 26, che a sua volta attiva più adrenocettori nel OB e APC, causando effetti cellulari e fisiologici che sostengono l&#39;apprendimento 22,27,28. In secondo luogo, i meccanismi di memoria portante avvengono in strutture neurali laminari ben definite. Ilsemplicità dei circuiti olfattiva nei ratti neonati fornisce ai ricercatori con il quadro ideale con cui scoprire i processi complessi legati alla plasticità sinaptica. Neuroni sensoriali olfattivi (OSN) nel progetto epitelio olfattivo sulla mitrale / cellule tufted in OB e questi mitrale / cellule tufted in progetto di girare ipsilaterally a corteccia piriforme (PC) attraverso il tratto olfattivo laterale (LOT), tra le altre strutture 29. Entrambe le sinapsi OSN della OB 30,31 e la sorte sinapsi 24,25 in aPC sono stati identificati come loci critici per cambiamenti sinaptici che supportano l&#39;apprendimento e la memoria. In terzo luogo, in tenera età nei ratti, ingressi olfattive possono essere facilmente lateralizzato. Ogni APC ha accesso alle informazioni bilaterale odore attraverso la commessura anteriore una volta che questa sostanza bianca è completamente formato in post-natale giorno 12 (PD12) 32. Prima di PD 12, ingresso odore può essere isolato a ipisilateral OB e APC attraverso un&#39;unica narice occlusione 24,25,31,33,34 &lt;/ Sup&gt;. Singola narice occlusione permette la formazione della memoria odore dalla narice aperta ed impedire la stessa memoria dei narice occluse prima di PD 12 33. Memoria odore è isolato per l&#39;emisfero ipsilaterale compreso sia OB e APC. Pertanto, ogni pup ratto può essere il proprio controllo per l&#39;apprendimento e alla base della fisiologia. Nel presente studio, è introdotto l&#39;odore primi protocollo apprendimento preferenza lateralizzato. Questo metodo serve come un potente strumento per studiare i meccanismi neurali alla base dell&#39;apprendimento odore, fornendo un controllo intra-animale 24,25,31, riducendo in tal modo sia il numero di animali necessari e la variazione generale. Naris occlusione è reversibile in quanto il tappo di grasso o naso può essere applicato e rimosso con il minimo stress o danni per l&#39;animale. Qui, in primo luogo, vengono descritte le procedure dettagliate di odore precoce preferenza formazione e test, con un focus sul protocollo lateralizzante con occlusione singola narice con un nose spina. Poi i risultati vengono presentati per dimostrare l&#39;efficacia del singolo occlusione narice a isolare ingresso odore e la produzione di memoria odore lateralizzato. Infine, le potenzialità di utilizzo di questo modello di apprendimento lateralizzato per studiare i cambiamenti fisiologici nel sistema olfattivo che entrambi generano l&#39;apprendimento e la memoria supporto di espressione sono discussi.

<table><tbody><tr><td>Polythylene 20 tubing</td><td>Intramedic</td><td>427406</td><td>Non-radiopaque, Non-toxic</td></tr><tr><td>3-0 Silk suture thread</td><td>Syneture</td><td>Sofsilk</td><td>Non-absorbent</td></tr><tr><td>Silicone grease</td><td>Warner Instrument</td><td>64-0378</td><td>Odorless</td></tr><tr><td>2% Xylocaine gel</td><td>AstraZeneca</td><td>Prod. No 061</td><td>Lidocaine hydrochloride jelly, purchased at local pharmacy</td></tr><tr><td>Paint brush</td><td>Dynasty</td><td>206R</td><td>Similar size/other brands work too</td></tr><tr><td>Peppermint extract</td><td>Sigma-Aldrich</td><td>W284807</td><td>Other brands should be okay too</td></tr><tr><td>Training box</td><td>Custom-made</td><td>N/A</td><td>Acrylic box (20 x 20 x 5 cm&lt;sup&gt;3&lt;/sup&gt;), see &lt;b&gt;Figure 2A&lt;/b&gt;. Parameters and material for the box are not critical and can be modified. Material used should be odorless and does not absorb odors</td></tr><tr><td>Testing chamber</td><td>Custom-made</td><td>N/A</td><td>Stainless steel (30 x 20 x 18 cm&lt;sup&gt;3&lt;/sup&gt;), see &lt;b&gt;Figure 2B&lt;/b&gt;. Parameters and material for the chamber are not critical and can be modified. For example, an acrylic chamber instead of a stainless steel one can be used</td></tr><tr><td>pCREB antibody</td><td>Cell Signaling</td><td>9198</td><td>Ser 133 (87G3) Rabbit mAb</td></tr><tr><td>Chloral hydrate</td><td>Sigma-Aldrich</td><td>C8383</td><td>N/A</td></tr><tr><td>Paraformaldehype</td><td>Sigma-Aldrich</td><td>P6148</td><td>N/A</td></tr><tr><td>Sucrose</td><td>Sigma-Aldrich</td><td>S9378</td><td>N/A</td></tr></tbody></table>

a lateralized odor learning model in neonatal rats for dissecting neural circuitry underpinning memory formation

Rat pups during a critical postnatal period (&#8804; 10 days) readily form a preference for an odor that is associated with stimuli mimicking maternal care. Such a preference memory can last from hours, to days, even life-long, depending on training parameters. Early odor preference learning provides us with a model in which the critical changes for a natural form of learning occur in the olfactory circuitry. An additional feature that makes it a powerful tool for the analysis of memory processes is that early odor preference learning can be lateralized via single naris occlusion within the critical period. This is due to the lack of mature anterior commissural connections of the olfactory hemispheres at this early age. This work outlines behavioral protocols for lateralized odor learning using nose plugs. Acute, reversible naris occlusion minimizes tissue and neuronal damages associated with long-term occlusion and more aggressive methods such as cauterization. The lateralized odor learning model permits within-animal comparison, therefore greatly reducing variance compared to between-animal designs. This method has been used successfully to probe the circuit changes in the olfactory system produced by training. Future directions include exploring molecular underpinnings of odor memory using this lateralized learning model; and correlating physiological change with memory strength and durations.

Qui passiamo in rassegna alcuni dei risultati stabiliti in precedenza 24 per dimostrare l&#39;efficacia dell&#39;occlusione narice a isolare ingresso odore e imparare a un emisfero, e la reversibilità di questo metodo. Occlusione singola narice durante l&#39;allenamento precoce preferenza odore conduce ad una memoria odore lateralizzato 24. La memoria è limitata alla narice risparmiato (Figura 3). Quando i cuccioli sono testati per la preferenza odori...

Watch this Scientific Journal Video about A Lateralized Odor Learning Model in Neonatal Rats for Dissecting Neural Circuitry Underpinning Memory Formation at JoVE.com

A Lateralized Odor Learning Model in Neonatal Rats for Dissecting Neural Circuitry Underpinning Memory Formation

This protocol introduces lateralized early odor preference learning in rats using acute single naris occlusion. Lateralized learning permits the examination of behavioral outcomes and underpinning biological mechanisms within the same animals, reducing variance induced by between-animal designs. This protocol can be used to investigate molecular mechanisms underpinning early odor learning.

Un laterizzazione Odore Learning Modello in ratti neonati per dissezione Formazione Neural circuiti Sostenere la memoria

Rat pups during a critical postnatal period (&#8804; 10 days) readily form a preference for an odor that is associated with stimuli mimicking maternal ...

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8.9K Views.  Faculty of Medicine, Memorial University. This protocol introduces lateralized early odor preference learning in rats using acute single naris occlusion. Lateralized learning permits the examination of behavioral outcomes and underpinning biological mechanisms within the same animals, reducing variance induced by between-animal designs. This protocol can be used to investigate molecular mechanisms underpinning early odor learning.

Video: A Lateralized Odor Learning Model in Neonatal Rats for Dissecting Neural Circuitry Underpinning Memory Formation

The Double-H Maze: A Robust Behavioral Test for Learning and Memory in Rodents

Spatial cognition research in rodents typically employs the use of maze tasks, whose attributes vary from one maze to the next. These tasks vary by their behavioral flexibility and required memory duration, the number of goals and pathways, and also the overall task complexity. A confounding feature in many of these tasks is the lack of control over the strategy employed by the rodents to reach the goal, e.g., allocentric (declarative-like) or egocentric (procedural) based strategies. The double-H maze is a novel water-escape memory task that addresses this issue, by allowing the experimenter to direct the type of strategy learned during the training period. The double-H maze is a transparent device, which consists of a central alleyway with three arms protruding on both sides, along with an escape platform submerged at the extremity of one of these arms.
Rats can be trained using an allocentric strategy by alternating the start position in the maze in an unpredictable manner (see protocol 1; &#167;4.7), thus requiring them to learn the location of the platform based on the available allothetic cues. Alternatively, an egocentric learning strategy (protocol 2; &#167;4.8) can be employed by releasing the rats from the same position during each trial, until they learn the procedural pattern required to reach the goal. This task has been proven to allow for the formation of stable memory traces.
Memory can be probed following the training period in a misleading probe trial, in which the starting position for the rats alternates. Following an egocentric learning paradigm, rats typically resort to an allocentric-based strategy, but only when their initial view on the extra-maze cues differs markedly from their original position. This task is ideally suited to explore the effects of drugs/perturbations on allocentric/egocentric memory performance, as well as the interactions between these two memory systems.

The goal of this protocol is to investigate spatial cognition in rodents. The double-H water maze is a novel test, which is particularly useful to elucidate the different components of learning, consolidation and memory, as well as the interplay of memory systems.

The Double-H labirinto: una prova comportamentale robusto per l&#39;apprendimento e la memoria nei roditori

Spatial cognition research in rodents typically employs the use of maze tasks, whose attributes vary from one maze to the next. These tasks vary by their ...

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A Method for Remotely Silencing Neural Activity in Rodents During Discrete Phases of Learning

This protocol describes how to temporarily and remotely silence neuronal activity in discrete brain regions while animals are engaged in learning and memory tasks. The approach combines pharmacogenetics (Designer-Receptors-Exclusively-Activated-by-Designer-Drugs) with a behavioral paradigm (sensory preconditioning) that is designed to distinguish between different forms of learning. Specifically, viral-mediated delivery is used to express a genetically modified inhibitory G-protein coupled receptor (the Designer Receptor) into a discrete brain region in the rodent. Three weeks later, when designer receptor expression levels are high, a pharmacological agent (the Designer Drug) is administered systemically 30 min prior to a specific behavioral session. The drug has affinity for the designer receptor and thus results in inhibition of neurons that express the designer receptor, but is otherwise biologically inert. The brain region remains silenced for 2-5 hr (depending on the dose and route of administration). Upon completion of the behavioral paradigm, brain tissue is assessed for correct placement and receptor expression. This approach is particularly useful for determining the contribution of individual brain regions to specific components of behavior and can be used across any number of behavioral paradigms.

This protocol describes how to temporarily and remotely silence neuronal activity in discrete brain regions while rats are engaged in learning and memory tasks. The approach combines pharmacogenetics (Designer-Receptors-Exclusively-Activated-by-Designer-Drugs) with a behavioral paradigm (sensory preconditioning) that is designed to distinguish between different components of learning.

Un metodo per Remotely Silencing Neural Activity in Roditore Durante Fasi discreti di apprendimento

This protocol describes how to temporarily and remotely silence neuronal activity in discrete brain regions while animals are engaged in learning and ...

Olfactory Context Dependent Memory: Direct Presentation of Odorants

Information is retrieved more effectively when the retrieval occurs in the same context as that in which the information was first encoded. This is termed context dependent memory (CDM). One category of cues that have been shown to effectively produce CDM effects are odors. However, it is unclear what the boundary conditions of these CDM effects are. In particular, given that olfaction has been called an implicit sense, it is possible that odors are only effective mnemonic cues when they are presented in the background. This assertion seems even more likely given that previous research has shown odors to be poor cues during paired associate memory tests, where odors are in the focus of attention as mnemonic cues for other information. In order to determine whether odors are only effective contextual mnemonic cues when presented outside the central focus of an observer, an olfactory CDM experiment was performed in which odorants were presented directly, rather than ambiently. Direct presentation was accomplished with the aid of an olfactometer. The olfactometer not only allows for direct presentation of odorants, but provides other methodological benefits, including the allowance of trial by trial manipulations of odorant presentations and, relatedly, time-specific releases of odorants. The presence of the same odor during both encoding and retrieval enhanced memory performance, regardless of whether the odor was presented ambiently or directly. This finding can serve as a basis for future olfactory CDM research which can utilize the benefits of direct presentation.

The use of an olfactometer for directly presenting odorants opens exciting opportunities for researchers of olfactory memory. The current paper discusses issues related to this methodology as related to a previously published experiment on olfactory context dependent memory.

Memoria dipendente dal contesto olfattivo: Presentazione diretta di odorizzazione

Information is retrieved more effectively when the retrieval occurs in the same context as that in which the information was first encoded. This is termed ...

Appetitive Associative Olfactory Learning in Drosophila Larvae

In the following we describe the methodological details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with genetic interference, provides a handle to analyze the neuronal and molecular fundamentals of specifically associative learning in a simple larval brain.
Organisms can use past experience to adjust present behavior. Such acquisition of behavioral potential can be defined as learning, and the physical bases of these potentials as memory traces1-4. Neuroscientists try to understand how these processes are organized in terms of molecular and neuronal changes in the brain by using a variety of methods in model organisms ranging from insects to vertebrates5,6. For such endeavors it is helpful to use model systems that are simple and experimentally accessible. The Drosophila larva has turned out to satisfy these demands based on the availability of robust behavioral assays, the existence of a variety of transgenic techniques and the elementary organization of the nervous system comprising only about 10,000 neurons (albeit with some concessions: cognitive limitations, few behavioral options, and richness of experience questionable)7-10.
Drosophila larvae can form associations between odors and appetitive gustatory reinforcement like sugar11-14. In a standard assay, established in the lab of B. Gerber, animals receive a two-odor reciprocal training: A first group of larvae is exposed to an odor A together with a gustatory reinforcer (sugar reward) and is subsequently exposed to an odor B without reinforcement 9. Meanwhile a second group of larvae receives reciprocal training while experiencing odor A without reinforcement and subsequently being exposed to odor B with reinforcement (sugar reward). In the following both groups are tested for their preference between the two odors. Relatively higher preferences for the rewarded odor reflect associative learning - presented as a performance index (PI). The conclusion regarding the associative nature of the performance index is compelling, because apart from the contingency between odors and tastants, other parameters, such as odor and reward exposure, passage of time and handling do not differ between the two groups9.

Appetitive Associative Olfactory Learning in Drosophila Larvae

Drosophila larvae are able to associate odor stimuli with gustatory reward. Here we describe a simple behavioral paradigm that allows the analysis of appetitive associative olfactory learning.

Appetitivo apprendimento associativo olfattiva in<em&gt; Drosophila</em&gt; Le larve

In the following we describe the methodological  details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with ...

Neuroscienze

Drosophila Adult Olfactory Shock Learning

Drosophila have been used in classical conditioning experiments for over 40 years, thus greatly facilitating our understanding of memory, including the elucidation of the molecular mechanisms involved in cognitive diseases1-7. Learning and memory can be assayed in larvae to study the effect of neurodevelopmental genes8-10 and in flies to measure the contribution of adult plasticity genes1-7. Furthermore, the short lifespan of Drosophila facilitates the analysis of genes mediating age-related memory impairment5,11-13. The availability of many inducible promoters that subdivide the Drosophila nervous system makes it possible to determine when and where a gene of interest is required for normal memory as well as relay of different aspects of the reinforcement signal3,4,14,16.
Studying memory in adult Drosophila allows for a detailed analysis of the behavior and circuitry involved and a measurement of long-term memory15-17. The length of the adult stage accommodates longer-term genetic, behavioral, dietary and pharmacological manipulations of memory, in addition to determining the effect of aging and neurodegenerative disease on memory3-6,11-13,15-21.
Classical conditioning is induced by the simultaneous presentation of a neutral odor cue (conditioned stimulus, CS+) and a reinforcement stimulus, e.g., an electric shock or sucrose, (unconditioned stimulus, US), that become associated with one another by the animal1,16. A second conditioned stimulus (CS-) is subsequently presented without the US. During the testing phase, Drosophila are simultaneously presented with CS+ and CS- odors. After the Drosophila are provided time to choose between the odors, the distribution of the animals is recorded. This procedure allows associative aversive or appetitive conditioning to be reliably measured without a bias introduced by the innate preference for either of the conditioned stimuli. Various control experiments are also performed to test whether all genotypes respond normally to odor and reinforcement alone.

Drosophila Adult Olfactory Shock Learning

The method to measure adult Drosophila associative memory is described. The assay is based on the ability of the fly to associate an odor presented with a negative reinforcer (electric shock) and then recall this information at a later time, allowing memory to be measured.

Drosophila adulti olfattiva Shock Learning

Drosophila have been used in classical conditioning experiments for over 40 years, thus greatly facilitating our understanding of memory, including the ...

A Low-Cost, Odor-Reward Association Task for Tests of Learning and Memory

Robust and simple behavioral paradigms of appetitive, associative memory are crucial for researchers interested in cellular and molecular mechanisms of memory. In this paper, an effective and low-cost mouse behavioral protocol is described for examining the effects of physiological manipulation (such as the infusion of pharmacological agents) on the learning rate and duration of odor-reward memory. Representative results are provided from a study examining the differential role of tyrosine kinase receptor activity in short-term (STM) and long-term memory (LTM). Male mice were conditioned to associate a reward (sugar pellet) with one of the two odors, and their memory for the association was tested 2 or 48 h later. Immediately prior to the training, a tyrosine kinase (Trk) receptor inhibitor or vehicle infusions were delivered into the olfactory bulb (OB). Although there was no effect of the infusion on the learning rate, blockade of the Trk receptors in the OB selectively impaired LTM (48 h), and not short-term memory (STM; 2 h). The LTM impairment was attributed to the diminished odor selectivity as measured by the length of the digging time. The culmination of the results of this experiment showed that Trk receptor activation in the OB is the key in olfactory memory consolidation.

An odor-reward, associative learning task was used to investigate the differential effects of physiological manipulation on long-term and short-term memory.

Robust and simple behavioral paradigms of appetitive, associative memory are crucial for researchers interested in cellular and molecular mechanisms of ...

<meta charset="utf8"></head><body>Progettazione di un olfattometro flessibile ed economico per esperimenti olfattivi sui roditori

Constructing an Olfactometer for Rodent Olfactory Behavior Studies

The use of olfactometers to study rodent behavior and brain activity during olfactory tasks is crucial for understanding brain circuits. These sophisticated devices allow researchers to precisely control and deliver odor stimuli, enabling the investigation of complex olfactory processes in rodents. Although commercially available olfactometers are convenient, they present challenges when technical issues arise, often requiring costly assistance and potentially disrupting research timelines. This article details the construction of a custom olfactometer specifically designed for mouse olfactory behavior experiments, providing a comprehensive list of parts and step-by-step instructions. The olfactometer is controlled through MATLAB, offering a user-friendly interface for researchers. Importantly, the open-source code allows users to modify and adapt the system, tailoring behavioral tasks to meet specific experimental needs. Building a customized olfactometer empowers users with the knowledge and capability to perform custom experimental design and troubleshooting independently, saving both time and resources. This approach not only enhances research flexibility but also fosters a deeper understanding of the equipment's functionality, ultimately leading to more robust and reliable olfactory studies in rodents.

<meta charset="utf8"></head><body>Costruzione di un olfattometro per studi sul comportamento olfattivo dei roditori Spettroscopia nel vicino infrarosso Studio di iperscansione in consulenza psicolo

This protocol describes the construction of an olfactometer for go/no-go olfactory behavior experiments. Step-by-step instructions, along with images, are provided to ensure the successful construction of the olfactometer. Information for troubleshooting issues encountered during the process is also included.

Costruzione di un olfattometro per studi sul comportamento olfattivo dei roditori

The use of olfactometers to study rodent behavior and brain activity during olfactory tasks is crucial for understanding brain circuits. These ...

An Objective and Reproducible Test of Olfactory Learning and Discrimination in Mice

Olfaction is the predominant sensory modality in mice and influences many important behaviors, including foraging, predator detection, mating, and parenting. Importantly, mice can be trained to associate novel odors with specific behavioral responses to provide insight into olfactory circuit function. This protocol details the procedure for training mice on a Go/No-Go operant learning task. In this approach, mice are trained on hundreds of automated trials daily for 2&#8211;4 weeks and can then be tested on novel Go/No-Go odor pairs to assess olfactory discrimination, or be used for studies on how odor learning alters the structure or function of the olfactory circuit. Additionally, the mouse olfactory bulb (OB) features ongoing integration of adult-born neurons. Interestingly, olfactory learning increases both the survival and synaptic connections of these adult-born neurons. Therefore, this protocol can be combined with other biochemical, electrophysiological, and imaging techniques to study learning and activity-dependent factors that mediate neuronal survival and plasticity.

Here, we train mice on an associative learning task to test odor discrimination. This protocol also allows for studies on learning-induced structural changes in the brain.

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