Apoio prestado pela Universidade do Alabama em Birmingham Faculdade Programa de Desenvolvimento Grant.

<html> 1. Psicofisiologia O Biopac Systems, Inc. sistema de monitoramento fisiológico (ver Tabela de equipamentos específicos) é não-padrão em instalações de equipamentos mais imagens. Cronograma de 15-30 minutos antes da chegada participante para configurar o monitoramento fisiológico e outros equipamentos descritos neste protocolo (Figura 1). <ol><li> Conectar um controle operacional AcqKnowledge sala de informática (Biopac Systems, Inc.) software de monitorização fisiológica para o MP150 Biopac (MP150WSW) usando um padrão Ethernet cabo cruzado (CBLETH2). </li><li> Conecte o Biopac Isolated Digital Interface (STP100C) para um controle operacional Apresentação sala de informática (Sistemas Neurocomportamentais, Inc; Albany, CA) usando um software DB25 M / F cabo de fita. </li><li> Conecte o Biopac GSR amplificador (EDA-100C-MRI) para a interferência RF filtro (MRIRFIF) dentro da sala de controle usando um cabo de extensão blindado (MECMRI-3). </li><li> Conectar a interferência RF filtro (MRIRFIF) para um cabo de extensão blindado (MECMRI-1) dentro da câmara de ressonância magnética. </li><li> Conecte o cabo de extensão de cabos blindados para levar fibra de carbono (LEAD 108) que prendem a eletrodos de rádio translúcido (EL508). Nota: Torcendo o leva em uma espiral apertada reduz artefatos nos dados de condutância da pele que podem ser criados durante a digitalização. </li><li> Anexar eletrodos de rádio translúcido (EL508) para a falange distal dos dedos médio e anelar da mão esquerda do participante. </li><li> Devido à natureza da exploração do equipamento, temperatura ambiente MRI câmara são muitas vezes fixados abaixo de 21 ° C. Cobrir o participante com um cobertor para manter a temperatura da mão. </li></ol> 2. As respostas comportamentais (Joystick) <ol><li> Conectar um controle sala de informática software Presentation operacional (Sistemas Neurocomportamentais, Inc; Albany, CA) para o joystick de unidade de interface de FORP (atual Designs, Inc; Philadelphia, PA) usando um cabo USB-mini. </li><li> Conecte um cabo de fibra óptica para a unidade de interface de FORP dentro da sala de controle, em seguida, passar o cabo através de uma guia de onda dentro da câmara de ressonância magnética. </li><li> Conecte o cabo de fibra óptica para o MR-joystick compatível. </li><li> Participantes diretos para colocar o joystick em uma posição confortável e fácil de alcançar. </li></ol> 3. Apresentação de estímulo <ol><li> Conectar um controle sala de informática software Presentation operacional para as portas externas VGA e áudio do IFIS-SA consola quarto (Invivo Corp, Orlando, FL) controle (Figura 1). </li><li> Verifique as conexões de fibra óptica do cabo entre o console da sala de controle IFIS ea Unidade de Interface Periférica IFIS dentro da câmara de ressonância magnética, bem como as conexões entre a Unidade de Interface Periférica e da Unidade de exibição de áudio / visual. </li><li> Coloque a unidade de exibição de áudio / visual por trás da cabeça da bobina de forma que o participante pode ver o monitor através de um espelho preso à cabeça da bobina. </li><li> Conectar a caixa de unidade de exibição de áudio / visual da interface acústica para MR compatível com o sistema IFIS de fones de ouvido estéreo usando tubos de vinil. </li><li> Calibrar o volume de estímulos auditivos usando um medidor de nível de pressão sonora. </li></ol> 4. Procedimento Experimental <ol><li> Informar os participantes que 2 tons serão apresentadas várias vezes durante o estudo, e que o volume dos tons podem variar acima e abaixo do seu limiar de percepção (Figura 2). </li><li> Participantes diretos de apertar um botão na caixa de joystick imediatamente após ouvir tom ou, então, para atualizar sua expectativa de receber a UCS movendo o joystick para controlar a posição de uma barra de classificação numa escala de 0 a 100 (Figura 3). </li><li> Instruir os participantes para classificar a sua esperança UCS em uma escala contínua de 0 a 100. Informá-los que as classificações de 0 indicam que eles estão certos da UCS não será apresentado, as classificações de 50 indicam que eles são incertos se a UCS serão apresentados, e as classificações de 100 indicam que eles estão certos da UCS será apresentado. Participantes diretos de usar outros valores na escala intermediária para indicar expectativas. Então, permitir aos participantes a prática usando o joystick para fazer avaliações. </li><li> Expor os participantes a um procedimento de condicionamento diferencial medo de usar dois tons (700 e 1.300 Hz, duração 10s; 20s ITI) como estímulos condicionados (CS) e um alto ruído branco (100 dB, 500ms) como a UCS. </li><li> Apresentar 60 ensaios do CS + (coterminating com o UCS) e 60 ensaios do CS-(apresentada sem a UCS) em uma ordem pseudo tal que não mais de 2 ensaios do CS mesmo são apresentados consecutivamente. </li><li> Contrabalançar os tons que servem como + CS e CS-entre os participantes. </li><li> Modular o volume da + CS e CS-independente. Ajustar o volume CS sobre o julgamento posterior com a mesma CS. Diminuir o volume de 5dB CS se pressionar um botão é feita (ou seja, após um julgamento percebida). Aumentar o volume de 5dB se pressionar um botão não é feita (ou seja, após um julgamento despercebido). </li></ol><html> ve_title &quot;Procedimento&gt; 5. Scanning <ol><li> Coletar padrão de alta resolução T1 estruturais (por exemplo MPRAGE) para servir como uma referência anatômica para dados funcionais. </li><li> Coletar fMRI BOLD de todo o cérebro durante o processo de condicionamento. Trinta e seis, 4mm de espessura deve ser suficiente para cobrir o cérebro com parâmetros de imagem relativamente padrão (por exemplo, TR = 2000ms matriz, TE = 30ms, FOV = 24 cm, 64x64). Sincronizar a aquisição fMRI com apresentação do estímulo usando uma caixa de gatilho fMRI. </li></ol> 6. SCR de Aquisição de Dados e Análise <ol><li> Condutância da pele em amostra de 2.000 Hz usando AcqKnowledge software eo MR-compatível Biopac sistema de monitoramento fisiológico descrito na seção 1. </li><li> Aplicar a 1 Hz Infinite Impulse Response (IIR) filtro passa-baixa digital para os dados de condutância da pele para reduzir artefatos produzidos durante o exame (veja Figura 4). </li><li> Resample os dados de condutância da pele em 250 Hz. </li><li> Calcule SCR como a diferença de nível da pele condutância do início de resposta ao pico de resposta. </li><li> Dados SCR pode ser transformado de raiz quadrada para normalizar a distribuição de amplitudes de resposta antes da análise estatística. </li></ol> 7. Expectativa UCS Aquisição de Dados e Análise <ol><li> Amostra (40 Hz) e registro de dados expectativa UCS usando software de apresentação. </li><li> Calcular a expectativa de UCS como a resposta (amostra 1s) média durante o último segundo de apresentação CS. </li></ol> 8. Funcional de Aquisição de Dados e Análise de ressonância magnética <ol><li> Pré-processamento padrão completo de dados de imagem cerebral (por exemplo, correção de sincronismo fatia, registro de imagens, suavização espacial), utilizando um pacote de software de imagem funcional de análise (por exemplo, AFNI 18). </li><li> Criar incômodo padrão (movimento, por exemplo) e estímulo baseada regressores para ensaios percebida e despercebida do CS e CS +, bem como a UCS. </li><li> Criar uma resposta baseada em motor de forma de onda de referência para servir como um regressor incômodo para dar conta do motor actividade relacionada com as respostas pressione o botão. <ol class="lalpha"><li> Crie uma função vara que codifica para o tempo de respostas pressione o botão. </li><li> Convolve o botão de função vara de imprensa com a função de resposta hemodinâmica canônica (HRF). </li></ol></li><li> Criar resposta com base em forma de onda do motor de referência para servir como um regressor incômodo para dar conta do motor actividade relacionada com as respostas joystick. <ol class="lalpha"><li> Crie uma função vara que codifica para o momento de mudanças na inclinação (por exemplo, valor da inclinação absoluto&gt; 10) de ratings expectativa UCS. </li><li> Convolve o joystick função de inclinação vara com a HRF canônico. </li></ol></li><li> Realizar análises de primeiro nível com todos os regressores estímulo-based e incômodo. </li><li> Realizar um segundo nível de medidas repetidas ANOVA para identificar regiões em que a ativação mostra um efeito principal do CS, tipo um efeito principal da percepção, ou um tipo de interação percepção CS X. </li></ol> 9. Resultados representativos: A metodologia apresentada aqui normalmente resulta em avaliações expectativa relativamente alta UCS durante percebida CS + testes e avaliações baixas durante percebida CS-ensaios (Figura 5) 10,15,19. Esses resultados indicam os participantes estão cientes de CS-UCS contingências. Em ensaios despercebido, avaliações expectativa UCS normalmente permanecem inalterados desde a pré-CS ratings. UCS expectativas sobre esses unperceived + CS e CS-ensaios geralmente caem perto de 50 participantes, indicando não tem certeza se o UCS será apresentado 10,15,19 (Figura 5). Esta incapacidade de produzir ratings diferencial expectativa UCS para o unperceived CS + e CS-indica que os participantes são incapazes de expressar a sua consciência de contingência em ensaios condicionado despercebido (Figura 6). Em contraste, a aprendizagem relacionados com mudanças no SCR têm sido observados durante os ensaios condicionado percebida e despercebida 10,15,19. Especificamente, SCRs eram maiores para o CS + percebida do que a percebida CS. Da mesma forma, SCRs maiores foram demonstrados durante CS unperceived + do que despercebido CS-ensaios 10,15,19 (Figura 6). Tomados em conjunto, estes dados comportamentais e autonômicas demonstrar o condicionamento do medo com a consciência de contingência em ensaios percebido, e condicionamento do medo sem consciência de contingência em ensaios despercebido. A pesquisa de imagem funcional utilizando esta metodologia demonstrou-aprendizagem relacionados com a ativação do hipocampo em percebida, mas não despercebido ensaios condicionado 15 (Figura 7). Em contraste, a atividade da amígdala diferencial foi observado em ambos os ensaios condicionado percebidas e despercebidas 15. Essas descobertas são consistentes com a visão de que o hipocampo suporta os processos referentes à sensibilização de contingência, enquanto a amígdala suporta expressão CR com e sem consciência. / 3083/3083fig1.jpg &quot;alt =&quot; Figura 1 &quot;/&gt; Figura 1. Diagrama de equipamentos básicos para a apresentação do estímulo e monitoramento da resposta comportamental / psicofisiológicas. Software de apresentação é usado para apresentar os estímulos áudio-visual e avaliações expectativa de monitorar UCS feitas por mover um joystick com a mão direita. AcqKnowledge software e Biopac equipamentos são usados ​​para monitorar a condutância da pele da mão esquerda. Sólido (Biopac), único linhas tracejadas (IFIS Áudio-Visual), e dupla tracejada (joystick de fibra óptica) retratam cabos para apresentação do estímulo distintos e sistemas de monitoramento de resposta. Setas pretas indicam a direção do fluxo de informações. <img src="/files/ftp_upload/3083/3083fig2.jpg" alt="Figura 2"/> Figura 2. Estímulos condicionados. Apresentar o + CS e CS-em uma ordem pseudo tal que não mais de 2 ensaios do CS mesmo são apresentados consecutivamente. Variar o volume do + CS e CS-independente. Se um CS é percebido (indicado por pressionar um botão), diminuição do volume CS 5dB sobre o julgamento posterior do CS mesmo. Se um CS é despercebido (indicado por não pressionar o botão), aumentar o volume CS 5dB sobre o julgamento posterior com a mesma CS. <img src="/files/ftp_upload/3083/3083fig3.jpg" alt="Figura 3"/> Figura 3. UCS escala de classificação expectativa. Instruir os participantes para classificar a sua expectativa de apresentação UCS em uma escala de 0 a 100. Pontuações de 0 indicam certeza a UCS não será apresentado, as classificações de 100 indicam certeza a UCS serão apresentados, e as classificações de 50 reflectir a incerteza sobre se a UCS será apresentado. Avaliações intermediárias devem ser usadas para indicar gradações na expectativa de UCS. <img src="/files/ftp_upload/3083/3083fig4.jpg" alt="Figura 4"/> Figura 4. Comparação de dados de condutância primas e filtrada pele. a) dados de condutância da pele Raw coletados durante fMRI. b) os dados de condutância da pele após a aplicação de um filtro passa-1Hz IIR baixo. <img src="/files/ftp_upload/3083/3083fig5.jpg" alt="Figura 5"/> Figura 5. Ratings expectativa UCS. Participantes normalmente relatam altas expectativas na UCS percebida CS + ensaios e expectativas pouco percebida CS-julgamentos. Expectativas UCS em unperceived + CS e CS-ensaios não diferem. <img src="/files/ftp_upload/3083/3083fig6.jpg" alt="Figura 6"/> Figura 6. UCS expectativa e SCR. As diferenças na esperança UCS são tipicamente observado em percebida CS + e CS-ensaios participantes indicando estão cientes das contingências de estímulo. Em ensaios despercebido, avaliações expectativa UCS, normalmente, não diferem indicando os participantes são incapazes de expressar a sua consciência de contingência. Em contraste, as diferenças de SCRs condicionado são geralmente observados em ambos os ensaios condicionado percebida e despercebida. Tais constatações refletem expressão com medo aprendido (ou seja, ensaios percebido) e sem (ou seja, ensaios despercebido) a consciência de contingência. <img src="/files/ftp_upload/3083/3083fig7.jpg" alt="Figura 7"/> Figura 7. Ressonância magnética funcional do hipocampo e amígdala. Respostas do hipocampo são geralmente maiores do que + CS CS-on percebido, mas não despercebido ensaios condicionado. Respostas diferenciais amígdala são tipicamente observado em ambos os ensaios condicionado percebida e despercebida. Essas descobertas são consistentes com a visão de que o hipocampo suporta os processos referentes à sensibilização de contingência, enquanto a amígdala suporta expressão com temor e sem consciência.

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A metodologia de medo condicionado descrito aqui fornece um meio para investigar os mecanismos neurais de processos conscientes e inconscientes da memória do medo. Este método tira vantagem do monitoramento simultâneo de dados comportamentais, autonômicas e fMRI. Monitoramento respostas comportamentais (ou seja, a expectativa UCS) e autonômicos (ie SCR) é um componente crítico deste método. Expectativa UCS oferece um meio para avaliar a consciência de contingência, enquanto SCR fornece um índice de expressão...

<table border="1"><tbody><tr><td> Equipamento </td><td> Companhia </td><td> Número do item </td></tr><tr><td> Sistema Integrado de Imagem Funcional (IFIS-SA) </td><td> Invivo Corp, Orlando, FL </td><td></td></tr><tr><td> Mestre Unidade de Controle (localizado na sala de controle) Unidade de Interface Periférica (localizado na câmara de ressonância magnética) Audio / Visual Display Unit (localizado na câmara de ressonância magnética), inclui: <ul><li> 6.4 &quot;(diagonal) monitor de vídeo LCD <ul><li> Resolução 640 x 480 e 15 ° de campo de visão </li></ul></li><li> caixa de interface acústica <ul><li> proporciona um som pneumático em estéreo </li></ul></li><li> MR compatível com fones de ouvido estéreo </li></ul></td><td></td><td></td></tr><tr><td> SISTEMA DE MONITORAMENTO FISIOLÓGICOS </td><td> Biopac Systems, Inc., Goleta, CA </td><td></td></tr><tr><td> Aquisição de Dados e Sistema de Análise para Windows (MP150) Isolado Digital Interface (Interface Digital) Resposta Galvânica da Pele Amplificador (GSR) MRI Cable / Filter System para definir Amplificador Transdutor, inclui: <ul><li> Cabo de extensão de MRI (Câmara de filtro) </li><li> Filtro de interferência de RF </li><li> Extensão MRI Cable (amplificador GSR para filtrar) </li></ul> Componentes adicionais: DB25 M / F cabo de fita Eletrodos descartáveis ​​radiotranslucent Fibra de carbono leva </td><td></td><td valign="top"> MP150WSW STP100C EDA100C-RM MECMRI-TRANS - MECMRI-1 - MRIRFIF - MECMRI-3 CBL110C EL508 LEAD108 </td></tr><tr><td> JOYSTICK </td><td> Designs atual, Inc., Philadelphia, PA </td><td></td></tr><tr><td> Joystick legado </td><td></td><td> HH-JOY-4 </td></tr><tr><td> Legado FORP interface </td><td></td><td> FIU-005 </td></tr></tbody></table>

investigating the neural mechanisms of aware and unaware fear memory with fmri

Condicionamento pavloviano medo é muitas vezes utilizado em combinação com ressonância magnética funcional (fMRI) em seres humanos para investigar os substratos neurais da aprendizagem associativa 1-5. Nesses estudos, é importante para fornecer provas de condicionamento comportamental para verificar que as diferenças na atividade cerebral estão aprendendo relacionados e correlacionados com o comportamento humano. Estudos de condicionamento do medo, muitas vezes acompanhar as respostas autonômicas (por exemplo, resposta de condutância da pele; SCR) como um índice de aprendizagem e memória 6-8. Além disso, outras medidas comportamentais podem fornecer informações valiosas sobre o processo de aprendizagem e / ou outras funções cognitivas que influenciam condicionado. Por exemplo, o impacto do estímulo incondicionado (UCS) têm expectativas sobre a expressão da resposta condicionada (CR) e resposta incondicionada (UCR) tem sido um tema de interesse em vários estudos recentes 9-14. SCR e medidas de expectativa de UCS têm sido usados ​​em conjunto com fMRI para investigar o substrato neural de aprendizado do medo consciente e inconsciente e processos de memória 15. Embora estes processos cognitivos podem ser avaliados em algum grau após a sessão de condicionamento, pós-condicionado avaliações não podem medir as expectativas em uma base experimental para tentativa-e são suscetíveis à interferência e esquecimento, bem como outros fatores que podem distorcer os resultados 16,17 . Monitoramento respostas autonômicas e comportamentais simultaneamente com fMRI fornece um mecanismo pelo qual os substratos neurais que medeiam as relações complexas entre processos cognitivos e respostas comportamentais / autônomo pode ser avaliado. No entanto, o monitoramento respostas autonômicas e comportamentais no ambiente MRI apresenta uma série de problemas práticos. Especificamente, 1) equipamentos de monitoramento padrão comportamental e fisiológica é construído de material ferroso que não pode ser utilizado com segurança perto do scanner de ressonância magnética, 2), quando este equipamento é colocado fora da câmara de ressonância magnética, os cabos de projetar para o assunto pode levar ruído RF que produz artefatos em imagens do cérebro, 3) os artefatos podem ser produzidos dentro do sinal de condutância da pele, alternando gradientes durante a digitalização, 4) o sinal de fMRI produzidos pelas exigências do motor de respostas comportamentais podem precisar de ser distinguida da actividade relacionada com os processos cognitivos de interesse . Cada um destes problemas podem ser resolvidos com modificações para a instalação de equipamento de monitorização fisiológica e procedimentos de análise de dados adicionais. Aqui apresentamos uma metodologia para monitorar simultaneamente respostas autonômicas e comportamentais durante o fMRI, e demonstrar o uso destes métodos para investigar os processos de memória consciente e inconsciente durante o condicionamento do medo.

Watch this Scientific Journal Video about Investigating the Neural Mechanisms of Aware and Unaware Fear Memory with fMRI at JoVE.com

Investigating the Neural Mechanisms of Aware and Unaware Fear Memory with fMRI

Uma metodologia para investigar os mecanismos neurais que suportam os processos de memória consciente e inconsciente durante o condicionamento do medo é descrito. Este método monitora o nível de oxigênio no sangue dependente (BOLD) a ressonância magnética funcional, a resposta de condutância da pele, ea expectativa de estímulo incondicionado durante o condicionamento pavloviano medo de avaliar os correlatos neurais dos processos de memória distintos.

Investigar os mecanismos neurais da memória Fear Consciente e Inconsciente com fMRI

Pavlovian fear conditioning is often used in combination with functional magnetic resonance imaging (fMRI) in humans to investigate the neural substrates ...

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Research

JoVE Journal

Neuroscience

12.8K visualizações.  University of Alabama at Birmingham. Uma metodologia para investigar os mecanismos neurais que suportam os processos de memória consciente e inconsciente durante o condicionamento do medo é descrito. Este método monitora o nível de oxigênio no sangue dependente (BOLD) a ressonância magnética funcional, a resposta de condutância da pele, ea expectativa de estímulo incondicionado durante o condicionamento pavloviano medo de avaliar os correlatos neurais dos processos de memória distintos.

Vídeo: Investigating the Neural Mechanisms of Aware and Unaware Fear Memory with fMRI

Simultaneous fMRI and Electrophysiology in the Rodent Brain

To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in which functional MRI data and in vivo intracortical recording can be performed simultaneously. The combination of MRI and electrical recording is technically challenging because the electrodes used for recording distort the MRI images and the MRI acquisition induces noise in the electrical recording. To minimize the mutual interference of the two modalities, glass microelectrodes were used rather than metal and a noise removal algorithm was implemented for the electrophysiology data. In our studies, two microelectrodes were separately implanted in bilateral primary somatosensory cortices (SI) of the rat and fixed in place. One coronal slice covering the electrode tips was selected for functional MRI. Electrode shafts and fixation positions were not included in the image slice to avoid imaging artifacts. The removed scalp was replaced with toothpaste to reduce susceptibility mismatch and prevent Gibbs ringing artifacts in the images. The artifact structure induced in the electrical recordings by the rapidly-switching magnetic fields during image acquisition was characterized by averaging all cycles of scans for each run. The noise structure during imaging was then subtracted from original recordings. The denoised time courses were then used for further analysis in combination with the fMRI data. As an example, the simultaneous acquisition was used to determine the relationship between spontaneous fMRI BOLD signals and band-limited intracortical electrical activity. Simultaneous fMRI and electrophysiological recording in the rodent will provide a platform for many exciting applications in neuroscience in addition to elucidating the relationship between the fMRI BOLD signal and neuronal activity.

We have developed a method for simultaneous functional magnetic resonance imaging and electrophysiological recording in the rodent brain, providing a platform for the investigation of the relationship between neural activity and the blood oxygenation level dependent (BOLD) MRI signal.

FMRI simultânea e Eletrofisiologia no cérebro de roedores

To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in ...

Neurociência

Extinction Training During the Reconsolidation Window Prevents Recovery of Fear

Fear is maladaptive when it persists long after circumstances have become safe. It is therefore crucial to develop an approach that persistently prevents the return of fear. Pavlovian fear-conditioning paradigms are commonly employed to create a controlled, novel fear association in the laboratory. After pairing an innocuous stimulus (conditioned stimulus, CS) with an aversive outcome (unconditioned stimulus, US) we can elicit a fear response (conditioned response, or CR) by presenting just the stimulus alone1,2 . Once fear is acquired, it can be diminished using extinction training, whereby the conditioned stimulus is repeatedly presented without the aversive outcome until fear is no longer expressed3. This inhibitory learning creates a new, safe representation for the CS, which competes for expression with the original fear memory4. Although extinction is effective at inhibiting fear, it is not permanent. Fear can spontaneously recover with the passage of time. Exposure to stress or returning to the context of initial learning can also cause fear to resurface3,4.
Our protocol addresses the transient nature of extinction by targeting the reconsolidation window to modify emotional memory in a more permanent manner. Ample evidence suggests that reactivating a consolidated memory returns it to a labile state, during which the memory is again susceptible to interference5-9. This window of opportunity appears to open shortly after reactivation and close approximately 6hrs later5,11,16, although this may vary depending on the strength and age of the memory15. By allowing new information to incorporate into the original memory trace, this memory may be updated as it reconsolidates10,11. Studies involving non-human animals have successfully blocked the expression of fear memory by introducing pharmacological manipulations within the reconsolidation window, however, most agents used are either toxic to humans or show equivocal effects when used in human studies12-14. Our protocol addresses these challenges by offering an effective, yet non-invasive, behavioral manipulation that is safe for humans.
By prompting fear memory retrieval prior to extinction, we essentially trigger the reconsolidation process, allowing new safety information (i.e., extinction) to be incorporated while the fear memory is still susceptible to interference. A recent study employing this behavioral manipulation in rats has successfully blocked fear memory using these temporal parameters11. Additional studies in humans have demonstrated that introducing new information after the retrieval of previously consolidated motor16, episodic17, or declarative18 memories leads to interference with the original memory trace14. We outline below a novel protocol used to block fear recovery in humans.

Conditioned fear can be diminished through an inhibitory process called extinction, but can resurface under conditions such as the passage of time or exposure to stress. Our protocol presents a novel way of preventing fear recovery by introducing extinction during the reconsolidation window (the re-storage phase of a reactivated memory).

Formação extinção durante a janela de reconsolidação impede a recuperação do Medo

Fear is maladaptive when it persists long after circumstances have become safe. It is therefore crucial to develop an approach that persistently prevents ...

Optical Recording of Suprathreshold Neural Activity with Single-cell and Single-spike Resolution

Signaling of information in the vertebrate central nervous system is often carried by populations of neurons rather than individual neurons. Also propagation of suprathreshold spiking activity involves populations of neurons. Empirical studies addressing cortical function directly thus require recordings from populations of neurons with high resolution. Here we describe an optical method and a deconvolution algorithm to record neural activity from up to 100 neurons with single-cell and single-spike resolution. This method relies on detection of the transient increases in intracellular somatic calcium concentration associated with suprathreshold electrical spikes (action potentials) in cortical neurons. High temporal resolution of the optical recordings is achieved by a fast random-access scanning technique using acousto-optical deflectors (AODs)1. Two-photon excitation of the calcium-sensitive dye results in high spatial resolution in opaque brain tissue2. Reconstruction of spikes from the fluorescence calcium recordings is achieved by a maximum-likelihood method. Simultaneous electrophysiological and optical recordings indicate that our method reliably detects spikes (&gt;97% spike detection efficiency), has a low rate of false positive spike detection (&lt; 0.003 spikes/sec), and a high temporal precision (about 3 msec) 3. This optical method of spike detection can be used to record neural activity in vitro and in anesthetized animals in vivo3,4.

Understanding the function of the vertebrate central nervous system requires recordings from many neurons because cortical function arises on the level of populations of neurons. Here we describe an optical method to record suprathreshold neural activity with single-cell and single-spike resolution, dithered random-access scanning. This method records somatic fluorescence calcium signals from up to 100 neurons with high temporal resolution. A maximum-likelihood algorithm deconvolves the underlying suprathreshold neural activity from the somatic fluorescence calcium signals. This method reliably detects spikes with high detection efficiency and a low rate of false positives and can be used to study neural populations in vitro and in vivo.

A gravação ótica da atividade neural supraliminares com resolução de uma única célula e um único pico

Signaling of information in the vertebrate central nervous system is often carried by populations of neurons rather than individual neurons. Also ...

Co-analysis of Brain Structure and Function using fMRI and Diffusion-weighted Imaging

The study of complex computational systems is facilitated by network maps, such as circuit diagrams. Such mapping is particularly informative when studying the brain, as the functional role that a brain area fulfills may be largely defined by its connections to other brain areas. In this report, we describe a novel, non-invasive approach for relating brain structure and function using magnetic resonance imaging (MRI). This approach, a combination of structural imaging of long-range fiber connections and functional imaging data, is illustrated in two distinct cognitive domains, visual attention and face perception. Structural imaging is performed with diffusion-weighted imaging (DWI) and fiber tractography, which track the diffusion of water molecules along white-matter fiber tracts in the brain (Figure 1). By visualizing these fiber tracts, we are able to investigate the long-range connective architecture of the brain. The results compare favorably with one of the most widely-used techniques in DWI, diffusion tensor imaging (DTI). DTI is unable to resolve complex configurations of fiber tracts, limiting its utility for constructing detailed, anatomically-informed models of brain function. In contrast, our analyses reproduce known neuroanatomy with precision and accuracy. This advantage is partly due to data acquisition procedures: while many DTI protocols measure diffusion in a small number of directions (e.g., 6 or 12), we employ a diffusion spectrum imaging (DSI)1, 2 protocol which assesses diffusion in 257 directions and at a range of magnetic gradient strengths. Moreover, DSI data allow us to use more sophisticated methods for reconstructing acquired data. In two experiments (visual attention and face perception), tractography reveals that co-active areas of the human brain are anatomically connected, supporting extant hypotheses that they form functional networks. DWI allows us to create a "circuit diagram" and reproduce it on an individual-subject basis, for the purpose of monitoring task-relevant brain activity in networks of interest.

We describe a novel approach for simultaneous analysis of brain function and structure using magnetic resonance imaging (MRI). We assess brain structure with high-resolution diffusion-weighted imaging and white-matter fiber tractography. Unlike standard structural MRI, these techniques allow us to directly relate anatomical connectivity to functional properties of brain networks.

Co-análise de estrutura e função cerebral usando ressonância magnética funcional e imagem ponderada em difusão

The study of complex computational systems is facilitated by network maps, such as circuit diagrams. Such mapping is particularly informative when ...

Whole Cell Patch Clamp for Investigating the Mechanisms of Infrared Neural Stimulation

It has been demonstrated in recent years that pulsed, infrared laser light can be used to elicit electrical responses in neural tissue, independent of any further modification of the target tissue. Infrared neural stimulation has been reported in a variety of peripheral and sensory neural tissue in vivo, with particular interest shown in stimulation of neurons in the auditory nerve. However, while INS has been shown to work in these settings, the mechanism (or mechanisms) by which infrared light causes neural excitation is currently not well understood. The protocol presented here describes a whole cell patch clamp method designed to facilitate the investigation of infrared neural stimulation in cultured primary auditory neurons. By thoroughly characterizing the response of these cells to infrared laser illumination in vitro under controlled conditions, it may be possible to gain an improved understanding of the fundamental physical and biochemical processes underlying infrared neural stimulation.

Infrared nerve stimulation has been proposed as an alternative to electrical stimulation in a range of nerve types, including those associated with the auditory system. This protocol describes a patch clamp method for studying the mechanism of infrared nerve stimulation in a culture of primary auditory neurons.

Todo patch clamp celular para investigar os mecanismos de estimulação neural Infrared

It has been  demonstrated in recent years that pulsed, infrared laser light can be used to  elicit electrical responses in neural tissue, independent of ...

Optogenetic Perturbation of Neural Activity with Laser Illumination in Semi-intact Drosophila Larvae in Motion

Drosophila larval locomotion is a splendid model system in developmental and physiological neuroscience, by virtue of the genetic accessibility of the underlying neuronal components in the circuits1-6. Application of optogenetics7,8 in the larval neural circuit allows us to manipulate neuronal activity in spatially and temporally patterned ways9-13. Typically, specimens are broadly illuminated with a mercury lamp or LED, so specificity of the target neurons is controlled by binary gene expression systems such as the Gal4-UAS system14,15. In this work, to improve the spatial resolution to "sub-genetic resolution", we locally illuminated a subset of neurons in the ventral nerve cord using lasers implemented in a conventional confocal microscope. While monitoring the motion of the body wall of the semi-intact larvae, we interactively activated or inhibited neural activity with channelrhodopsin16,17 or halorhodopsin18-20, respectively. By spatially and temporally restricted illumination of the neural tissue, we can manipulate the activity of specific neurons in the circuit at a specific phase of behavior. This method is useful for studying the relationship between the activities of a local neural assembly in the ventral nerve cord and the spatiotemporal pattern of motor output.

Optogenetic Perturbation of Neural Activity with Laser Illumination in Semi-intact Drosophila Larvae in Motion

Here we describe a protocol for optogenetic manipulation of motoneuronal activity while monitoring changes in motor output (muscle contraction) in semi-intact Drosophila larvae using lasers within a conventional confocal microscope. This technique enables researchers to achieve local perturbation of neural activity in a few neuromeres to elucidate the dynamics of motor circuits.

Optogenetic Perturbação da atividade neural com Iluminação Laser no Semi-intacta<em&gt; Drosophila</em&gt; As larvas in Motion

Drosophila larval locomotion is a splendid model system in developmental and physiological neuroscience, by virtue of the genetic accessibility of the ...

Deep Brain Stimulation with Simultaneous fMRI in Rodents

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for using blood oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI) to image rodents with simultaneous DBS. DBS fMRI presents a number of technical challenges, including accuracy of electrode implantation, MR artifacts created by the electrode, choice of anesthesia and paralytic to minimize any neuronal effects while simultaneously eliminating animal motion, and maintenance of physiological parameters, deviation from which can confound the BOLD signal.&#160;Our laboratory has developed a set of procedures that are capable of overcoming most of these possible issues. For electrical stimulation, a homemade tungsten bipolar microelectrode is used, inserted stereotactically at the stimulation site in the anesthetized subject. In preparation for imaging, rodents are fixed on a plastic headpiece and transferred to the magnet bore. For sedation and paralysis during scanning, a cocktail of dexmedetomidine and pancuronium is continuously infused, along with a minimal dose of isoflurane; this preparation minimizes the BOLD ceiling effect of volatile anesthetics. In this example experiment, stimulation of the subthalamic nucleus (STN) produces BOLD responses which are observed primarily in ipsilateral cortical regions, centered in motor cortex. Simultaneous DBS and fMRI allows the unambiguous modulation of neural circuits dependent on stimulation location and stimulation parameters, and permits observation of neuronal modulations free of regional bias. This technique may be used to explore the downstream effects of modulating neural circuitry at nearly any brain region, with implications for both experimental and clinical DBS.

This protocol describes a standard method for simultaneous functional magnetic resonance imaging and deep brain stimulation in the rodent. The combined use of these experimental tools allows for the exploration of global downstream activity in response to electrical stimulation at virtually any brain target.

Estimulação Cerebral Profunda com fMRI simultâneo em Roedores

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for ...

A Protocol for the Administration of Real-Time fMRI Neurofeedback Training

Neurologic disorders are characterized by abnormal cellular-, molecular-, and circuit-level functions in the brain. New methods to induce and control neuroplastic processes and correct abnormal function, or even shift functions from damaged tissue to physiologically healthy brain regions, hold the potential to dramatically improve overall health. Of the current neuroplastic interventions in development, neurofeedback training (NFT) from functional Magnetic Resonance Imaging (fMRI) has the advantages of being completely non-invasive, non-pharmacologic, and spatially localized to target brain regions, as well as having no known side effects. Furthermore, NFT techniques, initially developed using fMRI, can often be translated to exercises that can be performed outside of the scanner without the aid of medical professionals or sophisticated medical equipment. In fMRI NFT, the fMRI signal is measured from specific regions of the brain, processed, and presented to the participant in real-time. Through training, self-directed mental processing techniques, that regulate this signal and its underlying neurophysiologic correlates, are developed. FMRI NFT has been used to train volitional control over a wide range of brain regions with implications for several different cognitive, behavioral, and motor systems. Additionally, fMRI NFT has shown promise in a broad range of applications such as the treatment of neurologic disorders and the augmentation of baseline human performance. In this article, we present an fMRI NFT protocol developed at our institution for modulation of both healthy and abnormal brain function, as well as examples of using the method to target both cognitive and auditory regions of the brain.

The ability to induce and/or control neural plasticity may be critical in future treatments for neurologic disorders and the recovery from brain injury. In this paper, we present a protocol on the use of neurofeedback training with functional magnetic resonance imaging to modulate human brain function.

Um protocolo para a administração do real-time fMRI treinamento de Neurofeedback

Neurologic disorders are characterized by abnormal cellular-, molecular-, and circuit-level functions in the brain. New methods to induce and control ...

Time-dependent Increase in the Network Response to the Stimulation of Neuronal Cell Cultures on Micro-electrode Arrays

Micro-electrode arrays (MEAs) can be used to investigate drug toxicity, design paradigms for next-generation personalized medicine, and study network dynamics in neuronal cultures. In contrast with more traditional methods, such as patch-clamping, which can only record activity from a single cell, MEAs can record simultaneously from multiple sites in a network, without requiring the arduous task of placing each electrode individually. Moreover, numerous control and stimulation configurations can be easily applied within the same experimental setup, allowing for a broad range of dynamics to be explored. One of the key dynamics of interest in these in vitro studies has been the extent to which cultured networks display properties indicative of learning. Mouse neuronal cells cultured on MEAs display an increase in response following training induced by electrical stimulation. This protocol demonstrates how to culture neuronal cells on MEAs; successfully record from over 95% of the plated dishes; establish a protocol to train the networks to respond to patterns of stimulation; and sort, plot, and interpret the results from such experiments. The use of a proprietary system for stimulating and recording neuronal cultures is demonstrated. Software packages are also used to sort neuronal units. A custom-designed graphical user interface is used to visualize post-stimulus time histograms, inter-burst intervals, and burst duration, as well as to compare the cellular response to stimulation before and after a training protocol. Finally, representative results and future directions of this research effort are discussed.

Mouse neuronal cells cultured on multi-electrode arrays display an increase in response following electrical stimulation. This protocol demonstrates how to culture neurons, how to record activity, and how to establish a protocol to train the networks to respond to patterns of stimulation.

Aumento Time-dependente na resposta da rede à estimulação de culturas de pilha neuronais em matrizes do micro-elétrodo

Micro-electrode arrays (MEAs) can be used to investigate drug toxicity, design paradigms for next-generation personalized medicine, and study network ...

Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls body functions and internal organ rhythms when animals are exposed to emotional challenges and changes in their living environments. In general experiments, signals from different organs, such as the brain and the heart, are recorded by independent recording systems that require multiple recording devices and different procedures for processing the data files. This study describes a new method that can simultaneously monitor electrical biosignals, including tens of local field potentials in multiple brain regions, electrocardiograms that represent the cardiac rhythm, electromyograms that represent awake/sleep-related muscle contraction, and breathing signals, in a freely moving rat. The recording configuration of this method is based on a conventional micro-drive array for cortical local field potential recordings in which tens of electrodes are accommodated, and the signals obtained from these electrodes are integrated into a single electrical board mounted on the animal's head. Here, this recording system was improved so that signals from the peripheral organs are also transferred to an electrical interface board. In a single surgery, electrodes are first separately implanted into the appropriate body parts and the target brain areas. The open ends of all of these electrodes are then soldered to individual channels of the electrical board above the animal's head so that all of the signals can be integrated into the single electrical board. Connecting this board to a recording device allows for the collection of all of the signals into a single device, which reduces experimental costs and simplifies data processing, because all data can be handled in the same data file. This technique will aid the understanding of the neurophysiological correlates of the associations between central and peripheral organs.

This study introduces a method for the simultaneous recording of local field potentials in the brain, electrocardiograms, electromyograms, and breathing signals of a freely moving rat. This technique, which reduces experimental costs and simplifies data analysis, will contribute to the understanding of the interactions between the brain and peripheral organs.

Gravações simultâneas de potenciais de campo corticais Local, eletrocardiograma, eletromiograma e respirar o ritmo de um rato andando livremente

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls ...