Os autores gostariam de reconhecer o seguinte: Dr. Joseph Bronzino, Dr. Khamis Abu-Hassaballah, o Sr. Austin-RJ LaFrance, ea Sra. Jessica Koranda.

Este protocolo é apropriado para os ratinhos de 3 e 18 meses de idade e peso corporal aproximado de 30-50 g). Ratos podem ser obtidos no Laboratório Jackson (Bar Harbor, ME). Todos os protocolos cirúrgicos e experimentais foram aprovados pelo Comitê Animal Care e Use o Trinity College e estavam de acordo com o Guia do NIH para o Cuidado e Uso de Animais de Laboratório. 1. Preparação Animal e Procedimentos Cirúrgicos <ol><li> Prepare coquetel contendo anestesia de cetamina (25 mg / ml), xilazina (2,5 mg / mL) e acepromazina (0,5 mg / ml). Injectar 1 mL / kg de peso corporal no quadrante inferior direito do abdómen completada por injecções 0,2 ml / kg a cada 45 minutos depois disso a manter uma profundidade de anestesia estável utilizando o reflexo de retirada do pedal. </li><li> Prepare rato anestesiado para a cirurgia raspando a pele em seu crânio com um barbeador elétrico. A área raspada deve incluir no meio dos olhos para o meio das orelhas. Certifique-se de não fazer a barba a Whiskers como eles fazem parte do sistema sensorial barril mouse. </li><li> Use cotonetes para aplicar álcool isopropílico seguido de Betadine para a área raspada na cabeça. Em seguida, também usando cotonetes, aplicar uma pequena quantidade de óleo mineral sobre os olhos para evitar os globos oculares de secagem. </li><li> Use punhos da orelha de ratos infantis em vez de barras de ouvido regulares para montar a cabeça do animal sobre o aparelho quadro estereotáxico. Para fazer isso, a primeira montagem da orelha esquerda no punho da orelha esquerda. Então aliviar suavemente a orelha direita para a braçadeira direita, apoiando suavemente a cabeça do animal com uma mão e fazer avançar o manguito ouvido com a outra mão. Em seguida, coloque uma almofada de aquecimento sob o corpo do animal e configurá-lo para 86 ° F para manter a temperatura corporal. </li><li> Em seguida, verifique se há rigidez bilateral ao tentar mover o lado da cabeça para o outro. O animal está correctamente montada na armação estereotáxica quando a cabeça é rígido e não pode ser movida de lado a lado, mas pode facilmente rodar para cima e para baixo. </li><li> Delicadamente, coloque ofocinho de rato na barra de nariz / dente montagem certificando-se que os incisivos estão descansando suavemente, mas de forma segura dentro da abertura da lâmina da barra nariz / dente. </li><li> Usando um bisturi estéril, fazer uma incisão na linha média a partir de meados dos olhos para o meio das orelhas. Certifique-se que o bisturi é ser realizada em um ângulo de 45 ° e aplicar pressão suficiente para cortar através da fáscia subjacente, mas não através do crânio como que vai causar sangramento excessivo. </li><li> Use cotonetes para separar a fáscia e expor o crânio. Crânio marcos bregma e lambda deve ser claramente visível (Figura 1A). Assegure-se que o animal se encontra na posição crânio plana, ajustando a barra de nariz / dente de tal modo que bregma e lambda são marcos na mesma posição do dorso-ventral (DV) (+ 0,1 mm). </li><li> Aplicar uma pequena quantidade de soro fisiológico estéril para a área exposta e usar cotonetes para limpar suavemente o crânio. Então espere 2-3 min para o crânio completamente o ar secoantes de prosseguir. </li><li> Montar uma agulha no braço estereotáxico esquerda para medir a posição de DV bregma e lambda. O posicionamento DV destes dois pontos de referência deve estar dentro de 0,1 milímetros um do outro. Se não, a barra de nariz da estrutura estereotáxica pode ser ajustado para cima e para baixo para alcançar este objectivo. </li><li> Posicione a agulha no bregma para gravar seu ântero-posterior (AP), lateral (LAT) e medições dorsoventral (DV). Verifique se a agulha toca apenas o crânio e não penetrá-la. </li><li> Use um mouse atlas do cérebro, tais como &quot;O Rato do cérebro em estereotáxica Coordenadas&quot; 1, para determinar as coordenadas para as estruturas-alvo: medial via perfurante (MPP) no feixe angular e giro denteado (DG) do hipocampo, em relação a lambda e bregma, respectivamente (ver também Tabela 1). Em seguida, use uma caneta de ponta fina ou lápis para marcar esses pontos no crânio. </li><li> Além disso, marcar mais dois pontos no lado contralateral do crânio. Estes pontos which servirá de terra (GND) e de referência (REF) deve ser cerca de 3 mm a partir da linha média e posicionada longitudinalmente em relação umas às outras (Figura 1A, também a Tabela 1). </li><li> Retire o marcador agulha do braço estereotáxico esquerda e substituí-lo por uma broca dental elétrico equipado com um monte estereotáxica. Posicione a broca sobre cada ponto marcado e fazer pequenos furos rebarba de aproximadamente 0,5 mm de diâmetro. Quando a perfuração, use um movimento para cima e para baixo repetitivo e verificar com freqüência se a broca penetrou no crânio para expor a superfície do cérebro. </li><li> Delicadamente, mas com firmeza, dirigir um eletrodo parafuso (# 0-80 1/8 em parafusos de aço inoxidável, com fenda cabeça oval) em cada um dos buracos contralateral (GND e REF) de tal forma que apenas tocar a superfície cortical, sem penetrá-lo (Figura 1A ). Estes dois eletrodos parafuso servir como base e referência. </li><li> Mount estimulantes e gravação eletrodos em electrode titulares nos braços estereotáxica esquerdo e direito, respectivamente. </li><li> Conecte os terminais do eletrodo estimulando a um estimulador eletrofisiológico com isolamento atual e do terminal de eletrodo de registro a um amplificador diferencial eletrofisiológico (ganho = 1.000, filtro de banda = 1 Hz-3 kHz) e, em seguida, para um osciloscópio digital para inspeção visual de respostas evocadas. Além disso, conecte GND e terminais do eletrodo REF para o amplificador diferencial. </li><li> Suavemente e abaixe lentamente tanto estimulante e eletrodos de registro em incrementos de 0,5 mm enquanto monitorando visualmente a resposta evocada a um choque de estímulo 600-800 uA. Continue a baixar cada um dos eléctrodos, até que eles atinjam a sua posição respectiva DV-alvo e um sinal estereotipado é observado no osciloscópio (Figura 1B). </li><li> Depois disso, use cimento acrílico dental para fazer uma tampa para manter os terminais do eletrodo no lugar, qualquer cimento odontológico que toca a pele deve ser apagadoimediatamente. Uma vez que o cimento dentário é completamente curado transferir o animal a partir do quadro estereotáxico para uma gaiola limpa roedor. Use uma lâmpada de aquecimento ou uma almofada para manter a temperatura do núcleo. </li><li> Monitorar o animal a cada hora até que ele recupere a consciência. Uma injecção de flunixina pode ser administrado como um analgésico. </li></ol> 2. LTP Indução <ol><li> Permitir que o animal de 5-7 dias para se recuperar da cirurgia. Colocar o animal no meio de gravação que consiste de uma gaiola de Faraday (122 centímetros x 43 cm x 43 cm) equipadas com material de isolamento acústico e de um comutador rotativo 5-canal que liga o eléctrodo leva à configuração estimulante / gravação. </li><li> Permitir que o animal se adapte para 1-2 horas no ambiente de gravação antes de conectar os eletrodos para os instrumentos de estímulo e de gravação (consulte a etapa 1.17). </li><li> Defina os controles de estimulador para a saída de 400 uA e registrar a amplitude de uma média de 10 respostas evocadas certificando-them pelo menos 10 seg decorrer entre estímulos. Usar o método mostrado na Figura 1C para quantificar a resposta evocada. Repita este procedimento para 600, 800, 1.000, 1.200, e 1.400 uA. </li><li> Construir uma curva de entrada / saída traçando a amplitude média da resposta evocada versus intensidade de estímulo. A partir deste gráfico determinar a intensidade de estímulo que corresponde a 50% da amplitude máxima de medida. Utilizar a intensidade de 50% para o restante da experiência. </li><li> Em seguida, utilizando a intensidade do estímulo de 50%, se obter uma linha de base através da gravação da amplitude média de 5 respostas evocadas a cada minuto durante 15 min. Mais uma vez certifique-se de que pelo menos 10 seg decorrer entre estímulos. </li><li> Posteriormente entregar a estimulação tetânica composto por 10 rajadas de 10 pulsos entregues a 400 Hz com uma taxa de disparo de 5 Hz para a via perfurante medial. Monitorar o animal de perto para sinais de apreensão, incluindo shakes cachorro molhado. Se o animal tem uma convulsão a experiment deve ser encerrado e todos os dados de que animal deve ser excluída dos resultados do estudo. </li><li> Em seguida, continuar a gravar a amplitude média de 5 respostas evocadas a cada minuto por 30 min posttetanization. Depois disso, comparar esta amplitude à amplitude da linha de base obtido no passo 2.5 calculando a percentagem de alteração a partir da linha de base (Fig. 2). </li><li> Após a conclusão de experimentos, o animal é sacrificado por meio de inalação de isoflurano, um método que é consistente com as orientações sobre eutanásia da American Veterinary Medical Association. </li></ol>

<ol>
	<li>Paxinos, G., Franklin, K. B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The mouse brain in stereotaxic coordinates. , (2004).</li><li>Bliss, T. V. P., Lomo, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-lasting+potentiation+of+synaptic+transmission+in+the+dentate+area+of+the+anesthetized+rabbit+following+stimulation+of+the+perforant+path.">Long-lasting potentiation of synaptic transmission in the dentate area of the anesthetized rabbit following stimulation of the perforant path.</a> J. Physiol. 232, 331-356 (1973).</li><li>Blaise, J. H., Bronzino, J. D. <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+stimulus+frequency+and+age+on+bidirectional+synaptic+plasticity+in+the+dentate+gyrus+of+freely+moving+rats.">Effects of stimulus frequency and age on bidirectional synaptic plasticity in the dentate gyrus of freely moving rats.</a> Exp. Neurol. 182, 497-506 (2003).</li><li>Blaise, J. H., Koranda, J. L., Chow, U., Haines, K. E., Dorward, E. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neonatal+isolation+stress+alters+bidirectional+long-term+synaptic+plasticity+in+amygdalo-hippocampal+synapses+in+freely+behaving+adult+rats.">Neonatal isolation stress alters bidirectional long-term synaptic plasticity in amygdalo-hippocampal synapses in freely behaving adult rats.</a> Brain Res. 1193, 25-33 (2008).</li><li>Koranda, J. L., Masino, S. A., Blaise, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bidirectional+synaptic+plasticity+in+the+dentate+gyrus+of+the+awake+freely+behaving+mouse.">Bidirectional synaptic plasticity in the dentate gyrus of the awake freely behaving mouse.</a> J. Neurosci. Methods. 167, 160-166 (2008).</li><li>Cooke, S. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Autophosphorylation+of+alphaCaMKII+is+not+a+general+requirement+for+NMDA+receptor-dependent+LTP+in+the+adult+mouse.">Autophosphorylation of alphaCaMKII is not a general requirement for NMDA receptor-dependent LTP in the adult mouse.</a> J. Physiol. 574, 805-818 (2006).</li><li>Davis, S., Bliss, T. V., Dutrieux, G., Laroche, S., Errington, M. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+and+duration+of+long-term+potentiation+in+the+hippocampus+of+the+freely+moving+mouse.">Induction and duration of long-term potentiation in the hippocampus of the freely moving mouse.</a> J. Neurosci. Methods. 75, 75-80 (1997).</li><li>Jones, M. W., Peckham, H. M., Errington, M. L., Bliss, T. V., Routtenberg, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+plasticity+in+the+hippocampus+of+awake+C57BL/6+and+DBA/2+mice:+interstrain+differences+and+parallels+with+behavior.">Synaptic plasticity in the hippocampus of awake C57BL/6 and DBA/2 mice: interstrain differences and parallels with behavior.</a> Hippocampus. 11, 391-396 (2001).</li><li>Zhang, T. A., Tang, J., Pidoplichko, V. I., Dani, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Addictive+nicotine+alters+local+circuit+inhibition+during+the+induction+of+in+vivo+hippocampal+synaptic+potentiation.">Addictive nicotine alters local circuit inhibition during the induction of in vivo hippocampal synaptic potentiation.</a> J. Neurosci. 30, 6443-6453 (2010).</li><li>Tang, J., Dani, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dopamine+enables+in+vivo+synaptic+plasticity+associated+with+the+addictive+drug+nicotine.">Dopamine enables in vivo synaptic plasticity associated with the addictive drug nicotine.</a> Neuron. 63, 673-682 (2009).</li></ol>

Autores têm nada a revelar.

Neste protocolo, demonstramos um método simples e confiável para o estudo de LTP em DG em ratos comportando livremente. Enquanto muitos estudos de LTP em ratos acordados foram realizados 3,4, muito poucos têm sido realizados em ratos acordados, principalmente devido à complexidade técnica colocado pela imobiliária craniana limitada em ratos eo peso do headstages eletrodo em relação ao peso médio de camundongos 5. Os poucos estudos que demonstraram LTP na DG em comportar livremente camundon...

O desenvolvimento da tecnologia para manipular genes produziu modelos de ratos transgênicos e knockout de quase todas as doenças neurodegenerativas e neurológicos. Isso exigiu a tradução de técnicas de pesquisa eletrofisiológicos anteriormente utilizados em espécies de roedores maiores para o modelo animal mouse. Uma tal técnica de investigação neurofisiológicos é o uso de potenciação de longo prazo (LTP), para testar a eficácia das conexões sinápticas nas redes neuronais envolvidas em várias doenças neuropatológicas. Este protocolo descreve técnicas para a investigação eletrofisiológica confiável de LTP em comportar livremente ratos. A vantagem deste protocolo sobre os outros é que é simples e fácil de implementar, é também um pouco menos dispendioso, uma vez que não requer nem a utilização de sistemas de micro-unidade caros controlados por computador nem transistor de efeito de campo headstages, e, para o nosso conhecimento, é o primeiro protocolo de vídeo de registros eletrofisiológicos crônicas to estudo LTP em comportar livremente ratos. Para este fim, descrevemos no artigo metodologias simples para estudar a potenciação a longo prazo em murganhos comportando livremente. Estas metodologias podem ser facilmente traduzido para modelos transgênicos e knockout rato de distúrbios neuropatológicos.

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			<td height="21" style="height:21px;width:195px;">Ketamine (100 mg/ml)</td>
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			<td style="width:79px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:195px;">Xylazine (20 mg/ml)</td>
			<td style="width:104px;">Henry Schein</td>
			<td style="width:137px;">33197</td>
			<td style="width:79px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:195px;">Acepromazine (10 mg/ml)</td>
			<td style="width:104px;">Henry Schein</td>
			<td style="width:137px;">2177</td>
			<td style="width:79px;"></td>
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		<tr height="63">
			<td height="63" style="height:63px;width:195px;">Dental acrylic powder</td>
			<td style="width:104px;">Lang Dental Manufacturing Co.</td>
			<td style="width:137px;">1330CLR</td>
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			<td style="width:104px;">Lang Dental Manufacturing Co.</td>
			<td style="width:137px;">1306CLR</td>
			<td style="width:79px;"></td>
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		<tr height="63">
			<td height="63" style="height:63px;width:195px;">Tungsten wire (0.127 mm)</td>
			<td style="width:104px;">World Precision Instruments</td>
			<td style="width:137px;">TGW0515</td>
			<td style="width:79px;"></td>
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		<tr height="63">
			<td height="63" style="height:63px;width:195px;">Stainless Steel Hypodermic Tubing (0.286 mm)</td>
			<td style="width:104px;">World Precision Instruments</td>
			<td style="width:137px;">832400</td>
			<td style="width:79px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:195px;">Flunixin (50 mg/ml)</td>
			<td style="width:104px;">Henry Schein</td>
			<td style="width:137px;">14165</td>
			<td style="width:79px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:195px;">Epoxilyte</td>
			<td style="width:104px;">Superior Essex</td>
			<td style="width:137px;">EP 6001-M</td>
			<td style="width:79px;"></td>
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			<td height="63" style="height:63px;width:195px;">Stainless steel wire insert (0.2 mm)</td>
			<td style="width:104px;">World Precision Instruments</td>
			<td>792900</td>
			<td style="width:79px;"></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:195px;">Stereotaxic frame apparatus</td>
			<td style="width:95px;">Kopf Instruments</td>
			<td style="width:137px;">Model 902</td>
			<td style="width:79px;"></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:195px;">Ear cuffs (ear cups)</td>
			<td style="width:95px;">Kopf Instruments</td>
			<td style="width:137px;">Model 921&#160;</td>
			<td style="width:79px;"></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:195px;">Electrophysiological stimulator</td>
			<td style="width:95px;">Astro-Med, Inc.</td>
			<td style="width:137px;">S88</td>
			<td style="width:79px;"></td>
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		<tr height="63">
			<td height="63" style="height:63px;width:195px;">Digital oscilloscope</td>
			<td style="width:95px;">B &#38; K Precision Corp.</td>
			<td style="width:137px;">2542</td>
			<td style="width:79px;"></td>
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			<td height="42" style="height:42px;width:195px;">Current isolation unit</td>
			<td style="width:95px;">Astro-Med, Inc.</td>
			<td style="width:137px;">PSIU-6</td>
			<td style="width:79px;"></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:195px;">Differential amplifier</td>
			<td style="width:95px;">World Precision Instruments, Inc.</td>
			<td style="width:137px;">DAM-50</td>
			<td style="width:79px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:195px;">Commutator</td>
			<td style="width:95px;">Plastics One</td>
			<td style="width:137px;">SLC6</td>
			<td style="width:79px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:195px;">Dental drill</td>
			<td style="width:95px;">Stoelting</td>
			<td style="width:137px;">58650</td>
			<td style="width:79px;"></td>
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long-term potentiation of perforant pathway-dentate gyrus synapse in freely behaving mice

Estudos da potenciação de longa duração de eficácia sináptica, um fenômeno sináptica dependente de atividade que tem propriedades que o tornam atraente como um potencial mecanismo celular subjacente aprendizagem e armazenamento de informações, têm sido muito utilizados para elucidar a fisiologia de vários circuitos neuronais no hipocampo, amígdala e outras estruturas límbicas e corticais. Com isto em mente, os modelos de camundongos transgênicos de doenças neurológicas representam plataformas úteis para conduzir potenciação de longa duração (LTP) estudos para desenvolver uma maior compreensão do papel dos genes na comunicação sináptica normal e anormal em redes neuronais envolvidos na aprendizagem, emoção e informação processamento. Este artigo descreve as metodologias para indução de forma confiável LTP no rato comportando livremente. Estas metodologias podem ser utilizadas em estudos sobre transgênicos e knockout comportando livremente modelos do rato de doenças neurodegenerativas.

A Tabela 1 mostra as coordenadas para DG e MPP, usado neste protocolo Figura 1A apresenta as marcações para as estruturas alvo no crânio;.. Também são indicados a localização da terra e eletrodos de referência Figura 1B ilustra representante resposta evocada traça tanto pre- e posttetanization no mesmo animal. Note-se que a resposta evocada posttetanization é maior do que a resposta pretetanization que é indicativa de indução de LTP 2. Fi...

Watch this Scientific Journal Video about Long-term Potentiation of Perforant Pathway-dentate Gyrus Synapse in Freely Behaving Mice at JoVE.com

Long-term Potentiation of Perforant Pathway-dentate Gyrus Synapse in Freely Behaving Mice

Modelos de camundongos transgênicos e knockout de doenças neurológicas são úteis para estudar o papel dos genes em neurofisiologia normal e anormal. Este artigo descreve metodologias que podem ser usados ​​para estudar a potenciação a longo prazo, um mecanismo celular que pode ser a base de aprendizagem e de memória, em transgénicos e knockout livremente comportando modelos de rato de neuropatologia.

Potenciação de longo prazo da Via Perfurante-giro denteado Synapse em Livremente Comportando Ratos

Studies of long-term potentiation of synaptic efficacy, an activity-dependent synaptic phenomenon having properties that make it attractive as a potential ...

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14.0K visualizações.  Trinity College.  Modelos de camundongos transgênicos e knockout de doenças neurológicas são úteis para estudar o papel dos genes em neurofisiologia normal e anormal. Este artigo descreve metodologias que podem ser usados ​​para estudar a potenciação a longo prazo, um mecanismo celular que pode ser a base de aprendizagem e de memória, em transgénicos e knockout livremente comportando modelos de rato de neuropatologia. 

Vídeo: Long-term Potentiation of Perforant Pathway-dentate Gyrus Synapse in Freely Behaving Mice

Mouse Short- and Long-term Locomotor Activity Analyzed by Video Tracking Software

Locomotor activity (LMA) is a simple and easily performed measurement of behavior in mice and other rodents. Improvements in video tracking software (VTS) have allowed it to be coupled to LMA testing, dramatically improving specificity and sensitivity when compared to the line crossings method with manual scoring. In addition, VTS enables high-throughput experimentation. While similar to automated video tracking used for the open field test (OFT), LMA testing is unique in that it allows mice to remain in their home cage and does not utilize the anxiogenic stimulus of bright lighting during the active phase of the light-dark cycle. Traditionally, LMA has been used for short periods of time (mins), while longer movement studies (hrs-days) have often used implanted transmitters and biotelemetry. With the option of real-time tracking, long-, like short-term LMA testing, can now be conducted using videography. Long-term LMA testing requires a specialized, but easily constructed, cage so that food and water (which is usually positioned on the cage top) does not obstruct videography. Importantly, videography and VTS allows for the quantification of parameters, such as path of mouse movement, that are difficult or unfeasible to measure with line crossing and/or biotelemetry. In sum, LMA testing coupled to VTS affords a more complete description of mouse movement and the ability to examine locomotion over an extended period of time.

Locomotor activity (LMA) is a simple and easily performed measurement of behavior in mice. Coupling of video tracking software (VTS) and LMA allows for the improvement of specificity and sensitivity, especially when compared with the manual, line crossing method of LMA analysis. Additionally VTS allows long-term tracking of mouse LMA.

Rato de curto e longo prazo da atividade locomotora analisados ​​pelo software Rastreamento Vídeo

Locomotor  activity (LMA) is a simple and easily performed measurement of behavior in mice  and other rodents. Improvements in video tracking software ...

Comportamento

Construction of Microdrive Arrays for Chronic Neural Recordings in Awake Behaving Mice

State-of-the-art electrophysiological recordings from the brains of freely behaving animals allow researchers to simultaneously examine local field potentials (LFPs) from populations of neurons and action potentials from individual cells, as the animal engages in experimentally relevant tasks. Chronically implanted microdrives allow for brain recordings to last over periods of several weeks. Miniaturized drives and lightweight components allow for these long-term recordings to occur in small mammals, such as mice. By using tetrodes, which consist of tightly braided bundles of four electrodes in which each wire has a diameter of 12.5 &mu;m, it is possible to isolate physiologically active neurons in superficial brain regions such as the cerebral cortex, dorsal hippocampus, and subiculum, as well as deeper regions such as the striatum and the amygdala. Moreover, this technique insures stable, high-fidelity neural recordings as the animal is challenged with a variety of behavioral tasks. This manuscript describes several techniques that have been optimized to record from the mouse brain. First, we show how to fabricate tetrodes, load them into driveable tubes, and gold-plate their tips in order to reduce their impedance from M&Omega; to K&Omega; range. Second, we show how to construct a custom microdrive assembly for carrying and moving the tetrodes vertically, with the use of inexpensive materials. Third, we show the steps for assembling a commercially available microdrive (Neuralynx VersaDrive) that is designed to carry independently movable tetrodes. Finally, we present representative results of local field potentials and single-unit signals obtained in the dorsal subiculum of mice. These techniques can be easily modified to accommodate different types of electrode arrays and recording schemes in the mouse brain.

The design and assembly of microdrives for in vivo electrophysiological recordings of brain signals from the mouse is described. By attaching microelectrode bundles to sturdy driveable carriers, these techniques allow for long-term and stable neural recordings. The lightweight design allows for unrestricted behavioral performance by the animal following drive implantation.

Construção de Matrizes Microdrive para gravações Neurais crônica na Awake Mice comportam

State-of-the-art electrophysiological recordings from the brains of freely behaving animals allow researchers to simultaneously examine local field ...

Trace Fear Conditioning in Mice

In this experiment we present a technique to measure learning and memory. In the trace fear conditioning protocol presented here there are five pairings between a neutral stimulus and an unconditioned stimulus. There is a 20 sec trace period that separates each conditioning trial. On the following day freezing is measured during presentation of the conditioned stimulus (CS) and trace period. On the third day there is an 8 min test to measure contextual memory. The representative results are from mice that were presented with the aversive unconditioned stimulus (shock) compared to mice that received the tone presentations without the unconditioned stimulus. Trace fear conditioning has been successfully used to detect subtle learning and memory deficits and enhancements in mice that are not found with other fear conditioning methods. This type of fear conditioning is believed to be dependent upon connections between the medial prefrontal cortex and the hippocampus. One current controversy is whether this method is believed to be amygdala-independent. Therefore, other fear conditioning testing is needed to examine amygdala-dependent learning and memory effects, such as through the delay fear conditioning.

In the following experiment we describe a protocol for trace fear conditioning in mice. This type of associative memory includes a trace period that separates the neutral stimulus and the unconditioned stimulus.

Traçar medo condicionado em ratos

In this experiment we present a technique to measure learning and memory. In the trace fear conditioning protocol presented here there are five pairings ...

The Modified Hole Board - Measuring Behavior, Cognition and Social Interaction in Mice and Rats

This protocol describes the modified hole board (mHB), which combines features from a traditional hole board and open field and is designed to measure multiple dimensions of unconditioned behavior in small laboratory mammals (e.g., mice, rats, tree shrews and small primates). This paradigm is a valuable alternative for the use of a behavioral test battery, since a broad behavioral spectrum of an animal&#8217;s behavioral profile can be investigated in one single test.
The apparatus consists of a box, representing the &#8216;protected&#8217; area, separated from a group compartment. A board, on which small cylinders are staggered in three lines, is placed in the center of the box, representing the &#8216;unprotected&#8217; area of the set-up. The cognitive abilities of the animals can be measured by baiting some cylinders on the board and measuring the working and reference memory. Other unconditioned behavior, such as activity-related-, anxiety-related- and social behavior, can be observed using this paradigm. Behavioral flexibility and the ability to habituate to a novel environment can additionally be observed by subjecting the animals to multiple trials in the mHB, revealing insight into the animals&#8217; adaptive capacities.
Due to testing order effects in a behavioral test battery, na&#239;ve animals should be used for each individual experiment. By testing multiple behavioral dimensions in a single paradigm and thereby circumventing this issue, the number of experimental animals used is reduced. Furthermore, by avoiding social isolation during testing and without the need to food deprive the animals, the mHB represents a behavioral test system, inducing if any, very low amount of stress.

This protocol describes the modified hole board, which is a behavioral test set-up that comprises the characteristics of an open field and a traditional hole board. This set-up enables the differential analysis of unconditioned behavior of small laboratory mammals as well as the analysis of cognitive abilities.

The Hole Board Modificado - Medir Comportamento, Cognição e Interação Social em camundongos e ratos

This protocol describes the modified hole board (mHB), which combines features from a traditional hole board and open field and is designed to measure ...

A Novel Experimental and Analytical Approach to the Multimodal Neural Decoding of Intent During Social Interaction in Freely-behaving Human Infants

Understanding typical and atypical development remains one of the fundamental questions in developmental human neuroscience. Traditionally, experimental paradigms and analysis tools have been limited to constrained laboratory tasks and contexts due to technical limitations imposed by the available set of measuring and analysis techniques and the age of the subjects. These limitations severely limit the study of developmental neural dynamics and associated neural networks engaged in cognition, perception and action in infants performing &#8220;in action and in context&#8221;. This protocol presents a novel approach to study infants and young children as they freely organize their own behavior, and its consequences in a complex, partly unpredictable and highly dynamic environment. The proposed methodology integrates synchronized high-density active scalp electroencephalography (EEG), inertial measurement units (IMUs), video recording and behavioral analysis to capture brain activity and movement non-invasively in freely-behaving infants. This setup allows for the study of neural network dynamics in the developing brain, in action and context, as these networks are recruited during goal-oriented, exploration and social interaction tasks.

This protocol presents a novel methodology for the neural decoding of intent from freely-behaving infants during unscripted social interaction with an actor. Neural activity is acquired using non-invasive high-density active scalp electroencephalography (EEG). Kinematic data is collected with inertial measurement units and supplemented with synchronized video recording.

Uma nova abordagem experimental e analítico ao Neural Decodificação de Intenções Multimodal Durante Interação Social no Livremente-comportando bebês humanos

Understanding typical and atypical development remains one of the fundamental questions in developmental human neuroscience. Traditionally, experimental ...

A Pressure Injection System for Investigating the Neuropharmacology of Information Processing in Awake Behaving Macaque Monkey Cortex

The top-down modulation of feed-forward cortical information processing is functionally important for many cognitive processes, including the modulation of sensory information processing by attention. However, little is known about which neurotransmitter systems are involved in such modulations. A practical way to address this question is to combine single-cell recording with local and temporary neuropharmacological manipulation in a suitable animal model. Here we demonstrate a technique combining acute single-cell recordings with the injection of neuropharmacological agents in the direct vicinity of the recording electrode. The video shows the preparation of the pressure injection/recording system, including preparation of the substance to be injected. We show a rhesus monkey performing a visual attention task and the procedure of single-unit recording with block-wise pharmacological manipulations.

Here, we show the pressure injection of neuropharmacological substances during single-cell recording in an awake, behaving macaque monkey. This procedure allows pharmacological manipulation in the direct vicinity of a cortical recording site.

Um sistema de injeção de pressão para a investigação da neurofarmacologia de Processamento de Informação em Despertai Behaving Macaque Cortex

The top-down modulation of feed-forward cortical information processing is functionally important for many cognitive processes, including the modulation ...

Multi-layer Cortical Ca2+ Imaging in Freely Moving Mice with Prism Probes and Miniaturized Fluorescence Microscopy

In vivo circuit and cellular level functional imaging is a critical tool for understanding the brain in action. High resolution imaging of mouse cortical neurons with two-photon microscopy has provided unique insights into cortical structure, function and plasticity. However, these studies are limited to head fixed animals, greatly reducing the behavioral complexity available for study. In this paper, we describe a procedure for performing chronic fluorescence microscopy with cellular-resolution across multiple cortical layers in freely behaving mice. We used an integrated miniaturized fluorescence microscope paired with an implanted prism probe to simultaneously visualize and record the calcium dynamics of hundreds of neurons across multiple layers of the somatosensory cortex as the mouse engaged in a novel object exploration task, over several days. This technique can be adapted to other brain regions in different animal species for other behavioral paradigms.

Multi-layer Cortical Ca2+ Imaging in Freely Moving Mice with Prism Probes and Miniaturized Fluorescence Microscopy

Here, we present a procedure for performing large-scale Ca2+ imaging with cellular-resolution across multiple cortical layers in freely moving mice. Hundreds of active cells can be observed simultaneously using a miniature, head-mounted microscope coupled with an implanted prism probe.

Multi-layer Cortical Ca<sup&gt; 2+</sup&gt; Imaging em ratos com movimento livre com sondas de prisma e microscopia de fluorescência miniaturizada

In vivo circuit and cellular level functional imaging is a critical tool for understanding the brain in action. High resolution imaging of mouse cortical ...

A Wireless, Bidirectional Interface for In Vivo Recording and Stimulation of Neural Activity in Freely Behaving Rats

In vivo electrophysiology is a powerful technique to investigate the relationship between brain activity and behavior at a millisecond and micrometer scale. However, current methods mostly rely on tethered cable recordings or only use unidirectional systems, allowing either recording or stimulation of neural activity, but not at the same time or same target. Here, a new wireless, bidirectional device for simultaneous multichannel recording and stimulation of neural activity in freely behaving rats is described. The system operates through a single portable head stage that both transmits recorded activity and can be targeted in real-time for brain stimulation using a telemetry-based multichannel software. The head stage is equipped with a preamplifier and a rechargeable battery, allowing stable long-term recordings or stimulation for up to 1 h. Importantly, the head stage is compact, weighs 12 g (including battery) and thus has minimal impact on the animal&#180;s behavioral repertoire, making the method applicable to a broad set of behavioral tasks. Moreover, the method has the major advantage that the effect of brain stimulation on neural activity and behavior can be measured simultaneously, providing a tool to assess the causal relationships between specific brain activation patterns and behavior. This feature makes the method particularly valuable for the field of deep brain stimulation, allowing precise assessment, monitoring, and adjustment of stimulation parameters during long-term behavioral experiments. The applicability of the system has been validated using the inferior colliculus as a model structure.

A Wireless, Bidirectional Interface for In Vivo Recording and Stimulation of Neural Activity in Freely Behaving Rats

A wireless, bidirectional system for multi-channel neural recordings and stimulation in freely behaving rats is introduced. The system is light and compact, thus having minimal impact on the animal&#180;s behavioral repertoire. Moreover, this bidirectional system provides a sophisticated tool to assess causal relationships between brain activation patterns and behavior.

Uma rede sem fio, Interface bidirecional para gravação na Vivo e estimulação da atividade Neural em livremente, comportando-se ratos

In vivo electrophysiology is a powerful technique to investigate the relationship between brain activity and behavior at a millisecond and micrometer ...

Automated, Long-term Behavioral Assay for Cognitive Functions in Multiple Genetic Models of Alzheimer's Disease, Using IntelliCage

Multiple factors&#8212;such as aging and genes&#8212;are frequently associated with cognitive decline. Genetically modified mouse models of cognitive decline, such as Alzheimer&#39;s disease (AD), have become a promising tool to elucidate the underlying mechanisms and promote the therapeutic advances. An important step is the validation and characterization of expected behavioral abnormality in the models, in the case of AD, cognitive decline. The long-term behavioral investigations of laboratory animals to study the effect of aging demand substantial efforts from researchers. The IntelliCage system is a high-throughput and cost-effective test battery for mice that eliminates the need for daily human handling. Here, we describe how the system is utilized in the long-term phenotyping of a genetic Alzheimer&#39;s disease model, specifically focusing on the cognitive functions. The experiment employs repeated battery of tests that assess spatial learning and executive functions. This cost-effective age-dependent phenotyping allows us to identify the transient and/or permanent effects of genes on various cognitive aspects.

Automated, Long-term Behavioral Assay for Cognitive Functions in Multiple Genetic Models of Alzheimer&#039;s Disease, Using IntelliCage

This paper describes a protocol for cognitive assessments for genetic models of the Alzheimer&#39;s disease using the IntelliCage system, which is a high throughput automated behavioral monitoring system with operant conditioning.

Automatizada, a longo prazo ensaio comportamental para funções cognitivas em vários modelos genéticos da doença de Alzheimer, usando IntelliCage

Multiple factors&#8212;such as aging and genes&#8212;are frequently associated with cognitive decline. Genetically modified mouse models of cognitive ...

Simultaneous Recordings of Cortical Local Field Potentials and Electrocorticograms in Response to Nociceptive Laser Stimuli from Freely Moving Rats

Electrocortical responses, elicited by laser heat pulses that selectively activate nociceptive free nerve endings, are widely used in many animal and human studies to investigate the cortical processing of nociceptive information. These laser-evoked brain potentials (LEPs) consist of several transient responses that are time-locked to the onset of laser stimuli. However, the functional properties of the LEP responses are still largely unknown, due to the lack of a sampling technique that can simultaneously record neural activities at the surface of the cortex (i.e., electrocorticogram [ECoG] and scalp electroencephalogram [scalp EEG]) and inside the brain (i.e., local field potential [LFP]). To address this issue, we present here an animal protocol using freely moving rats. This protocol is composed of three main procedures: (1) animal preparation and surgical procedures, (2) a simultaneous recording of ECoG and LFP in response to nociceptive laser stimuli, and (3) data analysis and feature extraction. Specifically, with the help of a 3D-printed protective shell, both ECoG and LFP electrodes implanted on the rat&#39;s skull were securely held together. During data collection, laser pulses were delivered on the rat&#39;s forepaws through gaps in the bottom of the chamber when the animal was in spontaneous stillness. Ongoing white noise was played to avoid the activation of the auditory system by the laser-generated ultrasounds. As a consequence, only nociceptive responses were selectively recorded. Using the standard analytical procedures (e.g., band-pass filtering, epoch extraction, and baseline correction) to extract stimulus-related brain responses, we obtained results showing that LEPs with a high signal-to-noise ratio were simultaneously recorded from ECoG and LFP electrodes. This methodology makes the simultaneous recording of ECoG and LFP activities possible, which provides a bridge of electrocortical signals at the mesoscopic and macroscopic levels, thereby facilitating the investigation of nociceptive information processing in the brain.

We developed a technique that simultaneously records both electrocorticography and local field potentials in response to nociceptive laser stimuli from freely moving rats. This technique helps establish a direct relationship of electrocortical signals at the mesoscopic and macroscopic levels, which facilitates the investigation of nociceptive information processing in the brain.

Gravações simultâneas de potenciais de campo corticais Local e Electrocorticograms em resposta a estímulos nociceptivos Laser de ratos movimentando-se livremente

Electrocortical responses, elicited by laser heat pulses that selectively activate nociceptive free nerve endings, are widely used in many animal and ...