Gli autori desiderano ringraziare il Prof. ME Schwab, Dr. P. Freund, Dr. A. Wyss, Dr. S. Bashir, Dr. A. Mir, il Dr. J. Bloch, il Dr. JF Brunet, il Dr. J. Aebischer, Dr. A. Meszaros, Dr. V. Goetschman per il loro contributo agli esperimenti precedenti e analisi. Il set-up sperimentale sono stati costruiti da André Gaillard, Bernard Aebischer e Laurent Monney. Nella struttura animale, le scimmie sono stati posti sotto la sorveglianza dei custodi degli animali professionale: Josef Corpataux, Laurent Bossy e Jacques Maillard. I test comportamentali e analisi dei dati, così come istologia, sono state condotte con il contributo di grande valore di tecnici di laboratorio: Véronique Moret (anche web master), Françoise Tinguely, Christine Roulin, Monica Bennefeld, Christiane Marti e Georgette Fischer. Questo lavoro è stato sostenuto dal Fondo nazionale svizzero, non attribuisce alcun 31-61.857,00, 310000-110005, 31003A-132.465 (EMR), 310030-118357, 31003A-104061 (TW), 310030-120.411 (ABS), PZ00P3_121646 (ES), Novartis Foundation; Il Centro Nazionale di Competenza nella Ricerca (PRN) su &quot;plasticità neurale e riparazione&quot;.

<ol>
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M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+recovery+of+dexterity+following+unilateral+lesion+of+the+sensorimotor+cortex+in+adult+monkeys.">Mechanisms of recovery of dexterity following unilateral lesion of the sensorimotor cortex in adult monkeys.</a> Exp. Brain. Res. 128, 149-159 (1999).</li><li>Freund, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nogo-A-specific+antibody+treatment+enhances+sprouting+and+functional+recovery+after+cervical+lesion+in+adult+primates.">Nogo-A-specific antibody treatment enhances sprouting and functional recovery after cervical lesion in adult primates.</a> Nature. Med. 12, 790-792 (2006).</li><li>Freund, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anti-Nogo-A+antibody+treatment+promotes+recovery+of+manual+dexterity+after+unilateral+cervical+lesion+in+adult+primates--re-examination+and+extension+of+behavioral+data.">Anti-Nogo-A antibody treatment promotes recovery of manual dexterity after unilateral cervical lesion in adult primates--re-examination and extension of behavioral data.</a> Eur. J. Neurosci. 29, 983-996 (2009).</li><li>Kaeser, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+Unilateral+Motor+Cortex+Lesion+on+Ipsilesional+Hand's+Reach+and+Grasp+Performance+in+Monkeys:+Relationship+With+Recovery+in+the+Contralesional+Hand.">Effects of Unilateral Motor Cortex Lesion on Ipsilesional Hand's Reach and Grasp Performance in Monkeys: Relationship With Recovery in the Contralesional Hand.</a> J. Neurophysiol. 103, 1603-1645 (2010).</li><li>Kaeser, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Autologous+adult+cortical+cell+transplantation+enhances+functional+recovery+following+unilateral+lesion+of+motor+cortex+in+primates:+a+pilot+study.">Autologous adult cortical cell transplantation enhances functional recovery following unilateral lesion of motor cortex in primates: a pilot study.</a> Neurosurgery. 68, 1405-1417 (2011).</li><li>Bashir, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Short-term+effects+of+unilateral+lesion+of+the+primary+motor+cortex+(M1)+on+Ipsilesional+hand+dexterity+in+adult+macaque+monkeys.">Short-term effects of unilateral lesion of the primary motor cortex (M1) on Ipsilesional hand dexterity in adult macaque monkeys.</a> Brain Structure and Function. , (2011).</li><li>Hamadjida, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+anti-Nogo-A+treatment+on+the+reorganization+of+callosal+connectivity+of+the+premotor+cortical+areas+following+unilateral+lesion+of+primary+motor+cortex+(M1)+in+adult+macaque+monkeys.">Influence of anti-Nogo-A treatment on the reorganization of callosal connectivity of the premotor cortical areas following unilateral lesion of primary motor cortex (M1) in adult macaque monkeys.</a> , (2011).</li><li>Jenny, A. B., Inukai, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Principles+of+motor+organization+of+the+monkey+cervical+spinal+cord.">Principles of motor organization of the monkey cervical spinal cord.</a> J. Neurosci. 3, 567-575 (1983).</li><li>Schmidlin, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Progressive+plastic+changes+in+the+hand+representation+of+the+primary+motor+cortex+parallel+incomplete+recovery+from+a+unilateral+section+of+the+corticospinal+tract+at+cervical+level+in+monkeys.">Progressive plastic changes in the hand representation of the primary motor cortex parallel incomplete recovery from a unilateral section of the corticospinal tract at cervical level in monkeys.</a> Brain Research. 1017, 172-183 (2004).</li><li>Schmidlin, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reduction+of+the+hand+representation+in+the+ipsilateral+primary+motor+cortex+following+unilateral+section+of+the+corticospinal+tract+at+cervical+level+in+monkeys.">Reduction of the hand representation in the ipsilateral primary motor cortex following unilateral section of the corticospinal tract at cervical level in monkeys.</a> BMC Neuroscience. 6, 56-56 (2005).</li><li>Prescott, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Refinement+of+the+use+of+food+and+fluid+control+as+motivational+tools+for+macaques+used+in+behavioural+neuroscience+research:+report+of+a+Working+Group+of+the+NC3Rs.">Refinement of the use of food and fluid control as motivational tools for macaques used in behavioural neuroscience research: report of a Working Group of the NC3Rs.</a> J. Neurosci. Methods. 193, 167-188 (2010).</li><li>Darling, 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=Volumetric+effects+of+motor+cortex+injury+on+recovery+of+dexterous+movements.">Volumetric effects of motor cortex injury on recovery of dexterous movements.</a> Exp. Neurol. 220, 90-108 (2009).</li><li>Darling, W. 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Nessun conflitto di interessi dichiarati.

Nonostante i compiti attuali di comportamento sono state considerate finora nel nostro laboratorio, nel contesto di studi relativi alla lesione del midollo cervicale o lesione della corteccia motoria, con l&#39;obiettivo di testare i vari trattamenti (vedi 14,15,17 e <a href="http://www.unifr.ch/neuro/rouiller">http:/ / www.unifr.ch / neuro / Rouiller</a> &gt; selezionare &quot;ricerca&quot; nella barra dei menu in alto, quindi&gt; &quot;motore del sistema&quot; &quot;recupero dopo lesione&quot;&gt;), possono anche avere una più ampia applicazione, come destrezza manuale è anche un aspetto di prendere in considerazione in altre patologie, come il morbo di Parkinson (scimmie MPTP) o in caso di de-sensoriale afferentation propriocezione interessano e / o il senso del tatto (in particolare la prova 2 in assenza di controllo visivo). I test comportamentali proposti sono adatti per indagare il controllo motorio dei movimenti distali dell&#39;arto anteriore, in quanto coinvolti nella destrezza manuale. La specificità del test è dimostrato dall&#39;assenza di deficit (ad eccezione di un paio di giorni) in caso di una lesione che non pregiudica componenti rilevanti del sistema di controllo: infatti, in caso di una lesione posto più caudale rispetto al controllo dei motoneuroni muscoli della mano, non c&#39;era deficit. La pertinenza della prova 1 può essere apprezzata dal confronto su sequenze video presi in due punti l&#39;ora del post-lesione curva di recupero, che si differenziano da un miglioramento del recupero funzionale del 25%, probabilmente in relazione a un trattamento di terapia cellulare 17. Nonostante qualche iniziale, fase di formazione relativamente brevi all&#39;inizio (generalmente della durata di 2-3 mesi), le prove comportamentali proposti qui sono relativamente &quot;naturale&quot; e semplice, rispetto al complesso (ad esempio condizionale) i compiti per i quali la formazione della scimmia potrebbe richiedere circa un anno o più. Il rinforzo positivo è basata su cibi solidi, che è meno sensibile dal punto di vista etico di privazione di acqua, di solito utilizzato in più comcompiti complessi 23. Non c&#39;è bisogno di privare le scimmie dal cibo per ottenere risultati stabili e coerenti. Il pellet ricevuti durante le attività rappresentano il primo accesso al cibo il giorno della sessione comportamentali (assumendo che la scimmia non mangiare durante la notte precedente, ma cibo supplementare può essere somministrato fino alla fine del pomeriggio del giorno precedente) . E &#39;fondamentale che ogni scimmia esegue la sessione di comportamento, allo stesso tempo della giornata, così come rispettare lo stesso ordine sequenziale tra le scimmie diverse formare un gruppo nella stanza alloggi. Come le scimmie sono sensibili a segnali inquietanti eventi, i compiti del comportamento deve essere condotto in presenza di musica di sottofondo, mascherando potenziali rumori molesti provenienti dalle camere vicine o laboratori. E &#39;fondamentale che durante tutta la durata dell&#39;esperimento (dalla formazione iniziale fino all&#39;ultimo giorno della sessione sperimentale, un periodo di diversi mesi se non anni), una scimmia dato èposta quotidianamente sotto la supervisione dello sperimentatore stesso. I test comportamentali presenti, utilizzato da diversi anni nel nostro laboratorio per quantificare la destrezza manuale, sono in qualche modo paragonabile ad altri test di destrezza manuale recentemente riportato in letteratura 24-28. C&#39;è comunque una necessità cruciale di standardizzare i test attraverso diversi laboratori (per una migliore comparazione), che è un obiettivo provvisorio della presente relazione. Su richiesta, le proprietà dettagliata della set-up illustrato qui per i test di 1-4 possono essere forniti dall&#39;autore corrispondente, al fine di replicare loro. Al di là della questione della medicina rigenerativa (recupero dalla lesione del midollo spinale o corteccia cerebrale), la gamma attuale dei test può essere adatto per affrontare in condizioni normali non umani temi primati dello sviluppo (corso a tempo ad esempio di sviluppo motorio dei movimenti abili), per indagare aspetti lateralizzazione (mano preferenza / dominio) e alle domande decifrare evolutivo da comparing le abilità motorie di diverse specie di primati, tra cui soggetti umani. Si noti tuttavia che le dimensioni degli apparati deve essere adattata in base alle dimensioni delle cifre &#39;(spessore e lunghezza) delle specie di primati, in quanto può influenzare il rendimento delle attività. Nel presente studio, i test sono stati condotti sulle scimmie Macaca fascicularis, da 2,5 a 8 anni e di peso compreso tra 2,5 e 8 kg. La lunghezza del dito indice (usato prima di manipolare il pellet) varia dal 32-35 mm, mentre la circonferenza della falange distale (punta) del dito indice è compreso tra 22 e 25 mm nelle scimmie incluso nei nostri studi. Come testato in esperimenti precedenti, gli stessi test sono adatti per afferrare Macaca mulatta pure. Tutti gli esperimenti sono stati condotti in conformità con la Guida per la cura e l&#39;uso di animali da laboratorio (1996) e approvato dal locale (Svizzera) le autorità veterinarie. Tutte le procedure sperimentali sulle scimmies, come pure le condizioni di detenzione degli animali nella struttura, sono stati descritti in dettaglio nei rapporti recenti del nostro laboratorio: vedere i riferimenti 12-18.

behavioral assessment of manual dexterity in non-human primates

(CS) tratto corticospinale è il supporto anatomico delle capacità motorie squisita di manipolare abilmente piccoli oggetti, una prerogativa principalmente dei primati 1. In caso di lesioni che colpiscono il sistema di proiezione CS alla sua origine (lesione di aree motorie corticali) o lungo la sua traiettoria (lesione del midollo cervicale), vi è una drammatica perdita di destrezza manuale (paralisi mano), come si è visto in alcuni pazienti tetraplegici o emiplegica . Anche se c&#39;è qualche recupero spontaneo funzionale dopo lesione, rimane molto limitata nell&#39;adulto. Diverse strategie terapeutiche sono attualmente proposti (terapia cellulare per esempio, la neutralizzazione delle molecole inibitorie della crescita assonale, l&#39;applicazione di fattori di crescita, ecc), che sono per lo più sviluppata nei roditori. Tuttavia, prima applicazione clinica, è spesso raccomandata per testare la fattibilità, l&#39;efficacia e la sicurezza del trattamento in primati non umani. Ciò è particolarmente vero quando l&#39;obiettivo è quello di ripristinare la destrezza manuale dopouna lesione del sistema nervoso centrale, come l&#39;organizzazione del sistema motorio dei roditori è diverso da quello dei primati 1,2. Macachi sono illustrati qui come un modello idoneo a quantificare comportamentale manualità nei primati, per riflettere il passivo derivante dalla lesione della corteccia motoria o midollo cervicale, per esempio, misurare il grado di recupero funzionale e spontanea, quando un trattamento sia stato effettuato, valutare quanto può migliorare il recupero funzionale. La valutazione del comportamento di destrezza manuale si basa su quattro distinti, complementari, attività di raggiungere e afferrare manuale (uso della presa di precisione a cogliere pellet), che richiede una formazione iniziale di macachi adulti. La preparazione degli animali è dimostrata, così come il posizionamento rispetto al comportamento di set-up. Le prestazioni di una scimmia tipico è illustrato per ogni attività. La raccolta e l&#39;analisi dei parametri rilevanti che riflettono manipolazione precisa manozione, così come il controllo della forza, è spiegato e dimostrato con i risultati rappresentativi. Questi dati vengono inseriti poi in un contesto più ampio, mostrando come i dati comportamentali possono essere sfruttate per studiare l&#39;impatto di una lesione del midollo spinale o di una lesione della corteccia motoria e in quale misura un trattamento può migliorare il recupero funzionale spontaneo, mettendo a confronto diversi gruppi di scimmie (trattati versus sham trattati per esempio). Vantaggi e limiti dei test comportamentali sono discussi. L&#39;attuale approccio comportamentale è in linea con le precedenti relazioni sottolineando la pertinenza del non-umano modello primati nel contesto delle malattie del sistema nervoso 2,3.

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Behavioral Assessment of Manual Dexterity in Non-Human Primates

Come manualità è una prerogativa soprattutto dei primati, i compiti comportamentali sono stati sviluppati in macachi. Quattro raggiungere e afferrare i compiti prensione, che misura la capacità di manipolazione mano e la forza, permettono di stabilire il recupero funzionale dopo una lesione del sistema nervoso centrale e per testare l&#39;effetto di un trattamento.

La valutazione del comportamento di destrezza manuale in primati non umani

The corticospinal (CS) tract is the anatomical support of the exquisite motor ability to skillfully manipulate small objects, a prerogative mainly of ...

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22.2K Views.  University of Fribourg. Come manualità è una prerogativa soprattutto dei primati, i compiti comportamentali sono stati sviluppati in macachi. Quattro raggiungere e afferrare i compiti prensione, che misura la capacità di manipolazione mano e la forza, permettono di stabilire il recupero funzionale dopo una lesione del sistema nervoso centrale e per testare l'effetto di un trattamento.

Video: Behavioral Assessment of Manual Dexterity in Non-Human Primates

A General Method for Evaluating Incubation of Sucrose Craving in Rats

For someone on a food-restricted diet, food craving in response to food-paired cues may serve as a key behavioral transition point between abstinence and relapse to food taking 1. Food craving conceptualized in this way is akin to drug craving in response to drug-paired cues. A rich literature has been developed around understanding the behavioral and neurobiological determinants of drug craving; we and others have been focusing recently on translating techniques from basic addiction research to better understand addiction-like behaviors related to food 2-4.
As done in previous studies of drug craving, we examine sucrose craving behavior by utilizing a rat model of relapse. In this model, rats self-administer either drug or food in sessions over several days. In a session, lever responding delivers the reward along with a tone+light stimulus. Craving behavior is then operationally defined as responding in a subsequent session where the reward is not available. Rats will reliably respond for the tone+light stimulus, likely due to its acquired conditioned reinforcing properties 5. This behavior is sometimes referred to as sucrose seeking or cue reactivity. In the present discussion we will use the term "sucrose craving" to subsume both of these constructs.
In the past decade, we have focused on how the length of time following reward self-administration influences reward craving. Interestingly, rats increase responding for the reward-paired cue over the course of several weeks of a period of forced-abstinence. This "incubation of craving" is observed in rats that have self-administered either food or drugs of abuse 4,6. This time-dependent increase in craving we have identified in the animal model may have great potential relevance to human drug and food addiction behaviors.
Here we present a protocol for assessing incubation of sucrose craving in rats. Variants of the procedure will be indicated where craving is assessed as responding for a discrete sucrose-paired cue following extinction of lever pressing within the sucrose self-administration context (Extinction without cues) or as responding for sucrose-paired cues in a general extinction context (Extinction with cues).

Responding for food or drug-paired cues increases over the course of a period of abstinence and this may relate to an increased susceptibility to relapse behaviors. Here we detail a procedure for evaluating this "incubation of craving" in rats that have self-administered sucrose.

Un metodo generale per la valutazione di incubazione della brama di saccarosio in ratti

For someone on a food-restricted diet, food craving in response to food-paired cues may serve as a key behavioral transition point between abstinence and ...

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Neuroscienze

Recording and Analysis of Circadian Rhythms in Running-wheel Activity in Rodents

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, including rats, hamsters, and mice, are active during the night and relatively inactive during the day. Many other behavioral and physiological measures also exhibit daily rhythms, but in rodents, running-wheel activity serves as a particularly reliable and convenient measure of the output of the master circadian clock, the suprachiasmatic nucleus (SCN) of the hypothalamus. In general, through a process called entrainment, the daily pattern of running-wheel activity will naturally align with the environmental light-dark cycle (LD cycle; e.g. 12 hr-light:12 hr-dark). However circadian rhythms are endogenously generated patterns in behavior that exhibit a ~24 hr period, and persist in constant darkness. Thus, in the absence of an LD cycle, the recording and analysis of running-wheel activity can be used to determine the subjective time-of-day. Because these rhythms are directed by the circadian clock the subjective time-of-day is referred to as the circadian time (CT). In contrast, when an LD cycle is present, the time-of-day that is determined by the environmental LD cycle is called the zeitgeber time (ZT).
Although circadian rhythms in running-wheel activity are typically linked to the SCN clock6-8, circadian oscillators in many other regions of the brain and body9-14 could also be involved in the regulation of daily activity rhythms. For instance, daily rhythms in food-anticipatory activity do not require the SCN15,16 and instead, are correlated with changes in the activity of extra-SCN oscillators17-20. Thus, running-wheel activity recordings can provide important behavioral information not only about the output of the master SCN clock, but also on the activity of extra-SCN oscillators. Below we describe the equipment and methods used to record, analyze and display circadian locomotor activity rhythms in laboratory rodents.

Circadian rhythms in voluntary wheel-running activity in mammals are tightly coupled to the molecular oscillations of a master clock in the brain. As such, these daily rhythms in behavior can be used to study the influence of genetic, pharmacological, and environmental factors on the functioning of this circadian clock.

Registrazione e analisi dei ritmi circadiani in Running-ruota Attività nei roditori

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, ...

Assessment of Dendritic Arborization in the Dentate Gyrus of the Hippocampal Region in Mice

Dendritic arborization has been shown to be a reliable marker for examination of structural and functional integrity of neurons. Indeed, the complexity and extent of dendritic arborization correlates well with the synaptic plasticity in these cells. A reliable method for assessment of dendritic arborization is needed to characterize the deleterious effects of neurological disorders on these structures and to determine the effects of therapeutic interventions. However, quantification of these structures has proven to be a formidable task given their complex and dynamic nature. Fortunately, sophisticated imaging techniques can be paired with conventional staining methods to assess the state of dendritic arborization, providing a more reliable and expeditious means of assessment. Below is an example of how these imaging techniques were paired with staining methods to characterize the dendritic arborization in wild type mice. These complementary imaging methods can be used to qualitatively and quantitatively assess dendritic arborization that span a rather wide area within the hippocampal region.

We describe two methods for visualization and quantification of dendritic arborization in the hippocampus of mouse models: real-time and extended depth of field imaging. While the former method allows sophisticated topographical tracing and quantification of the extent of branching, the latter allows speedy visualization of the dendritic tree.

Valutazione della dendritiche arborizzazione nel giro dentato della Regione ippocampale nei topi

Dendritic arborization has been shown to be a reliable marker for examination of structural and functional integrity of neurons. Indeed, the complexity ...

Quantification of Filamentous Actin (F-actin) Puncta in Rat Cortical Neurons

Filamentous actin protein (F-actin) plays a major role in spinogenesis, synaptic plasticity, and synaptic stability. Changes in dendritic F-actin rich structures suggest alterations in synaptic integrity and connectivity. Here we provide a detailed protocol for culturing primary rat cortical neurons, Phalloidin staining for F-actin puncta, and subsequent quantification techniques. First, the frontal cortex of E18 rat embryos are dissociated into low-density cell culture, then the neurons grown in vitro for at least 12-14 days. Following experimental treatment, the cortical neurons are stained with AlexaFluor 488 Phalloidin (to label the dendritic F-actin puncta) and microtubule-associated protein 2 (MAP2; to validate the neuronal cells and dendritic integrity). Finally, specialized software is used to analyze and quantify randomly selected neuronal dendrites. F-actin rich structures are identified on second order dendritic branches (length range 25-75 &#181;m) with continuous MAP2 immunofluorescence. The protocol presented here will be a useful method for investigating changes in dendritic synapse structures subsequent to experimental treatments.

Quantification of Filamentous Actin F-actin Puncta in Rat Cortical Neurons

Filamentous actin (F-actin) plays an important role in spinogenesis, synaptic plasticity, and synaptic stability. Quantification of F-actin puncta is therefore a useful tool to study the integrity of synaptic structures. This protocol describes the procedures of quantifying F-actin puncta labeled with Phalloidin in low-density primary cortical neuronal cultures.

La quantificazione di filamentosa Actina (F-actina) Puncta nel ratto neuroni corticali

Filamentous actin protein (F-actin) plays a major role in spinogenesis, synaptic plasticity, and synaptic stability. Changes in dendritic F-actin rich ...

Identification of Dopamine D1-Alpha Receptor Within Rodent Nucleus Accumbens by an Innovative RNA In Situ Detection Technology

In the central nervous system, the D1-alpha subtype receptor (Drd1&#945;) is the most abundant dopamine (DA) receptor, which plays a vital role in regulating neuronal growth and development. However, the mechanisms underlying Drd1&#945; receptor abnormalities mediating behavioral responses and modulating working memory function are still unclear. Using a novel RNA in situ hybridization assay, the current study identified dopamine Drd1&#945; receptor and tyrosine hydroxylase (TH) RNA expression from DA-related circuitry in the nucleus accumbens (NAc) area and substantia nigra region (SNR), respectively. Drd1&#945; expression in the NAc shows a &#34;discrete dot&#34; staining pattern. Clear sex differences in Drd1&#945; expression were observed. In contrast, TH shows a &#34;clustered&#34; staining pattern. Regarding TH expression, female rats displayed a higher signal expression per cell relative to male animals. The methods presented here provide a novel in situ hybridization technique for investigating changes in dopamine system dysfunction during the progression of central nervous system diseases.

Identification of Dopamine D1-Alpha Receptor Within Rodent Nucleus Accumbens by an Innovative RNA In Situ Detection Technology

Identification of dopamine D1-alpha receptor in the nucleus accumbens is critical for clarifying D1 receptor dysfunction during a central nervous system disease. We performed a novel RNA in situ hybridization assay to visualize single RNA molecules in a specific brain area.

Identificazione della dopamina D1-alfa del ricevitore all'interno del roditore Nucleus Accumbens da un innovativo RNA In Situ Detection Technology

In the central nervous system, the D1-alpha subtype receptor (Drd1&#945;) is the most abundant dopamine (DA) receptor, which plays a vital role in ...

The Power of Interstimulus Interval for the Assessment of Temporal Processing in Rodents

Temporal processing deficits have been implicated as a potential elemental dimension of higher-level cognitive processes, commonly observed in neurocognitive disorders. Despite the popularization of prepulse inhibition (PPI) in recent years, many current protocols promote using a percent of control measure, thereby precluding the assessment of temporal processing. The present study used cross-modal PPI and gap prepulse inhibition (gap-PPI) to demonstrate the benefits of employing a range of interstimulus intervals (ISIs) to delineate effects of sensory modality, psychostimulant exposure, and age. Assessment of sensory modality, psychostimulant exposure, and age reveals the utility of an approach varying the interstimulus interval (ISI) to establish the shape of the ISI function, including increases (sharper curve inflections) or decreases (flattening of the response amplitude curve) in startle amplitude. Additionally, shifts in peak response inhibition, suggestive of a differential sensitivity to the manipulation of ISI, are often revealed. Thus, the systematic manipulation of ISI affords a critical opportunity to evaluate temporal processing, which may reveal the underlying neural mechanisms involved in neurocognitive disorders.

Temporal processing, a preattentive process, may underlie deficits in higher-level cognitive processes, including attention, commonly observed in neurocognitive disorders. Using prepulse inhibition as an exemplar paradigm, we present a protocol for manipulating interstimulus interval (ISI) to establish the shape of the ISI function to provide an assessment of temporal processing.

Il potere di Interstimulus intervallo per la valutazione di elaborazione temporale in roditori

Temporal processing deficits have been implicated as a potential elemental dimension of higher-level cognitive processes, commonly observed in ...

Assessment of Long-term Depression Induction in Adult Cerebellar Slices

Synaptic plasticity provides a mechanism for learning and memory. For cerebellar motor learning, long-term depression (LTD) of synaptic transmissions from parallel fibers (PF) to Purkinje cells (PC) is considered the basis for motor learning, and deficiencies of both LTD and motor learning are observed in various gene-manipulated animals. Common motor learning sets, such as adaptation of the optokinetic reflex (OKR), the vestibular-ocular reflex (VOR), and rotarod test were used for evaluation of motor learning ability. However, results obtained from the GluA2-carboxy terminus modified knock-in mice demonstrated normal adaptation of the VOR and the OKR, despite lacking PF-LTD. In that report, induction of LTD was only attempted using one type of stimulation protocol at room temperature. Thus, conditions to induce cerebellar LTD were explored in the same knock-in mutants using various protocols at near physiological temperature. Finally, we found stimulation protocols, by which LTD could be induced in these gene-manipulated mice. In this study, a set of protocols are proposed to evaluate LTD-induction, which will more accurately allow examination of the causal relationship between LTD and motor learning. In conclusion, experimental conditions are crucial when evaluating LTD in gene-manipulated mice.

In some gene-manipulated animals, using a single protocol may fail to induce LTD in cerebellar Purkinje cells, and there may be a discrepancy between LTD and motor learning. Multiple protocols are necessary to assess LTD-induction in gene-manipulated animals. Standard protocols are shown.

Valutazione dell'induzione della depressione a lungo termine nelle fette di cerebellar adulto

Synaptic plasticity provides a mechanism for learning and memory. For cerebellar motor learning, long-term depression (LTD) of synaptic transmissions from ...

Ballistic Labeling of Pyramidal Neurons in Brain Slices and in Primary Cell Culture

It has been reported that the size and shape of dendritic spines is related to their structural plasticity. To identify the morphological structure of pyramidal neurons and dendritic spines, a ballistic labeling technique can be utilized. In the present protocol, pyramidal neurons are labeled with DilC18(3) dye and analyzed using neuronal reconstruction software to assess neuronal morphology and dendritic spines. To investigate neuronal structure, dendritic branching analysis and Sholl analysis are performed, allowing researchers to draw inferences about dendritic branching complexity and neuronal arbor complexity, respectively. The evaluation of dendritic spines is conducted using an automatic assisted classification algorithm integral to the reconstruction software, which classifies spines into four categories (i.e., thin, mushroom, stubby, filopodia). Furthermore, an additional three parameters (i.e., length, head diameter, and volume) are also chosen to assess alterations in dendritic spine morphology. To validate the potential of wide application of the ballistic labeling technique, pyramidal neurons from in vitro cell culture were successfully labeled. Overall, the ballistic labeling method is unique and useful for visualizing neurons in different brain regions in rats, which in combination with sophisticated reconstruction software, allows researchers to elucidate the possible mechanisms underlying neurocognitive dysfunction.

We present a protocol to label and analyze pyramidal neurons, which is critical for evaluating potential morphological alterations in neurons and dendritic spines that may underlie neurochemical and behavioral abnormalities.

Etichettatura balistica dei neuroni piramidali nelle sezioni cerebrali e nella cultura cellulare primaria

It has been reported that the size and shape of dendritic spines is related to their structural plasticity. To identify the morphological structure of ...

A Hydrophobic Tissue Clearing Method for Rat Brain Tissue

Hydrophobic tissue clearing methods are easily adjustable, fast, and low-cost procedures that allows for the study of a molecule of interest in unaltered tissue samples. Traditional immunolabeling procedures require cutting the sample into thin sections, which restricts the ability to label and examine intact structures. However, if brain tissue can remain intact during processing, structures and circuits can remain intact for the analysis. Previously established clearing methods take significant time to completely clear the tissue, and the harsh chemicals can often damage sensitive antibodies. The iDISCO method quickly and completely clears tissue, is compatible with many antibodies, and requires no special lab equipment. This technique was initially validated for the use in mice tissue, but the current protocol adapts this method to image hemispheres of control and transgenic rat brains. In addition to this, the present protocol also makes several adjustments to preexisting protocol to provide clearer images with less background staining. Antibodies for Iba-1 and tyrosine hydroxylase were validated in the HIV-1 transgenic rat and in F344/N control rats using the present hydrophobic tissue clearing method. The brain is an interwoven network, where structures work together more often than separately of one another. Analyzing the brain as a whole system as opposed to a combination of individual pieces is the greatest benefit of this whole brain clearing method.

Here we present a hydrophobic tissue clearing method that allows for the viewing of target molecules as part of intact brain structures. This technique has now been validated for F344/N control and HIV-1 transgenic rats of both sexes.

Un metodo di compensazione del tessuto idrofobico per il tessuto cerebrale del ratto

Hydrophobic tissue clearing methods are easily adjustable, fast, and low-cost procedures that allows for the study of a molecule of interest in unaltered ...

A Rat Model of EcoHIV Brain Infection

It has been well studied that the EcoHIV infected mouse model is of significant utility in investigating HIV associated neurological complications. Establishment of the EcoHIV infected rat model for studies of drug abuse and neurocognitive disorders, would be beneficial in the study of neuroHIV and HIV-1 associated neurocognitive disorders (HAND). In the present study, we demonstrate the successful creation of a rat model of active HIV infection using chimeric HIV (EcoHIV). First, the lentiviral construct of EcoHIV was packaged in cultured 293 FT cells for 48 hours. Then, the conditional medium was concentrated and titered. Next, we performed bilateral stereotaxic injections of the EcoHIV-EGFP into F344/N rat brain tissue. One week after infection, EGFP fluorescence signals were detected in the infected brain tissue, indicating that EcoHIV successfully induces an active HIV infection in rats. In addition, immunostaining for the microglial cell marker, Iba1, was performed. The results indicated that microglia were the predominant cell type harboring EcoHIV. Furthermore, EcoHIV rats exhibited alterations in temporal processing, a potential underlying neurobehavioral mechanism of HAND as well as synaptic dysfunction eight weeks after infection. Collectively, the present study extends the EcoHIV model of HIV-1 infection to the rat offering a valuable biological system to study HIV-1 viral reservoirs in the brain as well as HAND and associated comorbidities such as drug abuse.

Here, we present a protocol to establish a new rat model of active HIV infection using chimeric HIV (EcoHIV), which is critical for enhancing our understanding of HIV-1 viral reservoirs in the brain and offering a system to study HIV-associated neurocognitive disorders and associated comorbidities (i.e., drug abuse).

Un modello di ratto di infezione cerebrale EcoHIV

It has been well studied that the EcoHIV infected mouse model is of significant utility in investigating HIV associated neurological complications. ...