Investigação da participação da amídala basolateral e suas aferências corticais no comportamento de reinstalação da busca de etanol induzida pelo contexto em ratos
Data
2022
Autores
Tavares, Gabrielle Emily Boaventura 
Orientadores
Cruz, Fábio Cardoso
Tipo
Dissertação de mestrado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
O etanol é a substância de abuso lícita mais consumida em diversos países. Seu uso de forma crônica, pode desencadear o transtorno por uso de substâncias. Um dos maiores desafios para o tratamento destes transtornos são as altas taxas de recaída mesmo após longos períodos de abstinência. Estudos clínicos e pré-clínicos indicam que a exposição a estímulos ambientais associados ao uso desta droga de abuso pode desencadear uma recaída. Entretanto, os mecanismos neurais envolvidos nesse
fenômeno, ainda não estão totalmente elucidados. Neste contexto, a amídala basolateral é apontada na literatura como uma região importante nos comportamentos de aprendizado associativo e respostas emocionais ao uso de drogas. Além disso, as projeções corticais recebidas por esta região têm se mostrado importantes na regulação de comportamentos aprendidos. Portanto, o objetivo do presente estudo foi analisar o papel da amídala basolateral e de suas aferências provindas do córtex orbitofrontal, pré-límbico e infralímbico, na reinstalação da busca de etanol induzida pelo contexto. Para tanto, ratos Long-Evans, foram treinados a autoadministrarem etanol, em caixas operantes, contextualmente específicas (Contexto A). Em seguida, as respostas nas barras ativas foram extintas (respostas não reforçadas) em caixas com contextos diferentes (Contexto B). Por fim, a capacidade do contexto de induzir a reinstalação da busca por etanol, foi avaliada expondo os animais aos contextos A ou B nas mesmas condições de extinção. Utilizando esse protocolo, avaliamos: a) a participação da amídala basolateral no comportamento de reinstalação de busca de etanol induzida pelo contexto, através da técnica imunofluorescência para a proteína Fos (marcador de ativação neuronal) e com a inibição farmacológica desta região; b) o envolvimento das projeções do córtex orbitofrontal, pré-límbico e infralímbico, para a amídala basolateral na reinstalação da busca de etanol induzida pelo contexto,
através da técnica de dupla marcação por imunofluorescência para marcadores neurais retrógrados e anticorpo contra a proteína Fos e c) as alterações moleculares em sinaptoneurossomas da amídala na reinstalação da busca de etanol induzida pelo contexto, através da técnica de western blotting para proteínas alvo. Nossos resultados não indicaram diferenças no número de células positivas para Fos na
amídala basolateral entre os animais expostos aos diferentes contextos, bem como a inibição farmacológica da amídala basolateral não atenuou a reinstalação da busca de etanol induzida pelo contexto. Além disso, observamos que a reinstalação da busca de etanol induzida pelo contexto foi correlacionada ao aumento da ativação das projeções do córtex pré-límbico para a amídala basolateral, mas não das projeções vindas do córtex infralímbico e orbitofrontal. Por fim, não encontramos diferenças no nível de expressão das proteínas avaliadas em sinaptoneurossomas da amídala de animais expostos aos contextos A, B e Home-cage. Nossos resultados sugerem que a amídala basolateral está envolvida na reinstalação da busca de etanol induzida pelo contexto, e que a via do córtex pré-límbico parece mediar esse envolvimento.
Ethanol is the legal drug most used in many countries. Chronic use can trigger substance use disorder. One major challenge in treating these disorders is the high relapse rates, even after periods of abstinence. Clinical and preclinical studies indicate that exposure to environmental cues associated with drug use could trigger a relapse. However, the neural mechanisms involved in this phenomenon are not fully elucidated. In this context, basolateral amygdala has been implicated in the literature as a critical region in associative learning behaviors and emotional responses to drug use. Furthermore, the cortical projections received by this region have been shown to play a role in modulating learned behaviors. Therefore, this study aimed to analyze the role of basolateral amygdala and its cortical inputs from the orbitofrontal, prelimbic, and infralimbic cortex in context-induced reinstatement of ethanol-seeking. Therefore, Long-Evans rats were trained to self-administer ethanol in contextually specific operating chambers (Context A). Then, the responses in the active lever were extinct (non-reinforced responses) in contextually different chambers (Context B). Finally, the role of context in the reinstatement of ethanol-seeking was evaluated by exposing the animals to contexts A or B under the same extinction conditions. Using this protocol, we evaluated: a) participation of basolateral amygdala in context-induced reinstatement of ethanol-seeking behavior, by the immunofluorescence technique for the Fos protein (neuronal activation marker) and with pharmacological inhibition; b) the involvement of projections from the orbitofrontal, prelimbic, and infralimbic cortex, to basolateral amygdala in context-induced reinstatement of ethanol-seeking, by the immunofluorescence double labeling technique for retrograde neural markers and antibodies for Fos protein and c) molecular changes in amygdala synaptoneurosomes in context-induced reinstatement of ethanol-seeking, using western blotting technique for target proteins. Our results did not indicate differences in the number of Fos-positive cells in basolateral amygdala between animals exposed to different contexts. Also, pharmacological inhibition of the basolateral amygdala did not attenuate contextinduced reinstatement of ethanol-seeking. Furthermore, we observed that context-induced reinstatement of ethanol-seeking was correlated with increased activation of projections from prelimbic cortex to basolateral amygdala, but not to projections from infralimbic and orbitofrontal cortex. Finally, we did not find differences in expressions levels of evaluated proteins in amygdala synaptoneurosomes of animals exposed to contexts A, B, and home-cage. Our results suggest that basolateral amygdala has a role in context-induced reinstatement of ethanol-seeking behavior and the prelimbic cortex pathway appears to mediate this involvement.
Ethanol is the legal drug most used in many countries. Chronic use can trigger substance use disorder. One major challenge in treating these disorders is the high relapse rates, even after periods of abstinence. Clinical and preclinical studies indicate that exposure to environmental cues associated with drug use could trigger a relapse. However, the neural mechanisms involved in this phenomenon are not fully elucidated. In this context, basolateral amygdala has been implicated in the literature as a critical region in associative learning behaviors and emotional responses to drug use. Furthermore, the cortical projections received by this region have been shown to play a role in modulating learned behaviors. Therefore, this study aimed to analyze the role of basolateral amygdala and its cortical inputs from the orbitofrontal, prelimbic, and infralimbic cortex in context-induced reinstatement of ethanol-seeking. Therefore, Long-Evans rats were trained to self-administer ethanol in contextually specific operating chambers (Context A). Then, the responses in the active lever were extinct (non-reinforced responses) in contextually different chambers (Context B). Finally, the role of context in the reinstatement of ethanol-seeking was evaluated by exposing the animals to contexts A or B under the same extinction conditions. Using this protocol, we evaluated: a) participation of basolateral amygdala in context-induced reinstatement of ethanol-seeking behavior, by the immunofluorescence technique for the Fos protein (neuronal activation marker) and with pharmacological inhibition; b) the involvement of projections from the orbitofrontal, prelimbic, and infralimbic cortex, to basolateral amygdala in context-induced reinstatement of ethanol-seeking, by the immunofluorescence double labeling technique for retrograde neural markers and antibodies for Fos protein and c) molecular changes in amygdala synaptoneurosomes in context-induced reinstatement of ethanol-seeking, using western blotting technique for target proteins. Our results did not indicate differences in the number of Fos-positive cells in basolateral amygdala between animals exposed to different contexts. Also, pharmacological inhibition of the basolateral amygdala did not attenuate contextinduced reinstatement of ethanol-seeking. Furthermore, we observed that context-induced reinstatement of ethanol-seeking was correlated with increased activation of projections from prelimbic cortex to basolateral amygdala, but not to projections from infralimbic and orbitofrontal cortex. Finally, we did not find differences in expressions levels of evaluated proteins in amygdala synaptoneurosomes of animals exposed to contexts A, B, and home-cage. Our results suggest that basolateral amygdala has a role in context-induced reinstatement of ethanol-seeking behavior and the prelimbic cortex pathway appears to mediate this involvement.