Importância dos interneurônios GABAérgicos Parvalbumina-Positivos do córtex insular para comportamentos relacionados à impulsividade e de procura de etanol
Data
2023-04-10
Autores
Silva, Fernando Bezerra Romualdo da 
Orientadores
Cruz, Fábio Cardoso
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
O álcool é a substância de abuso mais consumida no mundo. Seu consumo está relacionado a índices elevados de morbidade e mortalidade. Estudos com adolescentes correlacionam o uso dessa substância comportamentos impulsivos. A impulsividade é definida como ações mal avaliadas que implicam, geralmente, em respostas prejudiciais ao indivíduo. A ínsula parece desempenhar papel importante no controle de comportamentos impulsivos. Estudos demonstram que essa modulação pode ser exercida por interneurônios GABAérgicos Parvalbuminérgicos (IPV+). O objetivo do presente estudo foi investigar o papel dos IPV+, do córtex insular, nos comportamentos relacionados à impulsividade e consumo de etanol em camundongos machos C57BL/6J adolescentes (Dia Pós Natal – DPN 28-47). No primeiro experimento, os animais foram submetidos aos seguintes testes comportamentais: Labirinto em Zero Elevado (LZE), Esquiva Inibitória (ESIN), Inibição Pré-Pulso (PPI) e Omissão em Comando Operante (OCO), durante a adolescência e posteriormente na idade adulta (DPN64-83), para escolha do teste mais sensível para avaliação da expressão de comportamentos relacionados à impulsividade. Os dados provenientes dessa avaliação foram submetidos à Análise de Componentes Principais (PCA), que nos indicou que os comportamentos relacionados à impulsividade foram mais expressos durante a adolescência, não se mantiveram na idade adulta e que a OCO foi o melhor teste para essa avaliação. No segundo experimento, grupos distintos de animais adolescentes e adultos foram avaliados por meio do protocolo de OCO e pelo teste de Livre escolha entre duas garrafas para análise da impulsividade, consumo e preferência de etanol (10%). Observamos que camundongos adolescentes, apresentaram maior impulsividade, consumo e preferência por etanol, mesmo quando essa substância foi adulterada por quinino. E, ainda, através da PCA, foi demonstrado correlação entre comportamentos relacionados a impulsividade e consumo de álcool, em camundongos adolescentes. Após os testes comportamentais parte dos animais foram perfundidos para análise imunoistoquímica da proteína Fos e outra parte foi submetido a eutanásia para a análise das quantidades de proteínas Sinaptopodina e GAD67 na insula. Não observamos diferenças entre os grupos para essas análises. No terceiro experimento, os animais foram submetidos ao protocolo de OCO e em seguida perfundidos para realização da técnica de imunofluorescência para as proteínas Fos, NeuN e Parvalbumina, no córtex insular. Nossos resultados demonstram que camundongos adolescentes, apresentaram menor ativação de IPV+ e maior ativação neuronal da ínsula, além de escores mais elevados de comportamentos relacionados à impulsividade. No quarto experimento, utilizamos a técnica de quimiogenética para modular a atividade dos IPV+, em camundongos C57/BL6 transgênicos (PV-Cre), durante o teste de OCO. Observamos causalidade dos IPV+ nos comportamentos relacionados à impulsividade. No último experimento, utilizamos o mesmo procedimento anterior para analisar a participação dos IPV+ na OCO e consumo operante de etanol. Foi observado que a ativação desses neurônios diminuiu o comportamento impulsivo e a busca de etanol. Em conclusão, nossos resultados sugerem que a diminuição da atividade de IPV+, do córtex insular, parece estar associada aos comportamentos relacionados à impulsividade e procura por etanol.
Alcohol is the most widely consumed substance of abuse in the world. Its consumption is related to high rates of morbidity and mortality. Studies with adolescents correlate the use of this substance with impulsive behavior. Impulsivity is defined as poorly evaluated actions that usually imply responses that are harmful to the individual. The insula seems to play an important role in controlling impulsive behavior. Studies show that this modulation can be exerted by GABAergic Parvalbumin interneurons (IPV+). The aim of the present study was to investigate the role of IPV+, from the insular cortex, in impulsivity-related behaviors and ethanol consumption in adolescent male C57BL/6J mice (Post Natal Day - PND 28-47). In the first experiment, the animals were submitted to the following behavioral tests: Elevated Zero Maze (LZE), Inhibitory Avoidance (ESIN), Pre-Pulse Inhibition (PPI) and Omission in Operant Conditioning (OOC), during adolescence and later in adulthood (DPN64-83), to choose the most sensitive test to evaluate the expression of impulsivity-related behaviors. The data from this evaluation were submitted to Principal Component Analysis (PCA), which indicated that impulsivity-related behaviors were more expressed during adolescence and did not persist into adulthood, and that the OOC was the best test for this evaluation. In the second experiment, different groups of adolescent and adult animals were evaluated using the OOC protocol and the Two-Bottle Choice test to analyze impulsivity, consumption and preference of ethanol (10%). We observed that adolescent mice presented higher impulsivity, consumption and preference for ethanol, even when this substance was adulterated with quinine. Furthermore, through PCA, a correlation between impulsivity and alcohol consumption was demonstrated in adolescent mice. After the behavioral tests, part of the animals were perfused for immunohistochemical analysis of Fos protein and another part was euthanized to analyze the amounts of Synaptopodin and GAD67 proteins in the insula. We did not observe differences between the groups for these analyses. In the third experiment, the animals were submitted to the OOC protocol and then perfused to perform the immunofluorescence technique for Fos, NeuN, and Parvalbumin proteins in the insular cortex. Our results demonstrate that adolescent mice, showed less activation of IPV+ and more neuronal activation in the insula, as well as higher scores of impulsivity-related behaviors. In the fourth experiment, we used the chemogenetic technique to modulate the activity of IPV+, in C57/BL6 transgenic mice (PV-Cre), during the OOC test. We observed IPV+ causality on impulsivity-related behaviors. In the last experiment, we used the same previous procedure to analyze the participation of BTI+ in OOC and operant ethanol consumption. It was observed that activation of these neurons decreased impulsive behavior and ethanol seeking. In conclusion, our results suggest that decreased IPV+ activity, from the insular cortex, seems to be associated with impulsivity and ethanol-seeking related behaviors.
Alcohol is the most widely consumed substance of abuse in the world. Its consumption is related to high rates of morbidity and mortality. Studies with adolescents correlate the use of this substance with impulsive behavior. Impulsivity is defined as poorly evaluated actions that usually imply responses that are harmful to the individual. The insula seems to play an important role in controlling impulsive behavior. Studies show that this modulation can be exerted by GABAergic Parvalbumin interneurons (IPV+). The aim of the present study was to investigate the role of IPV+, from the insular cortex, in impulsivity-related behaviors and ethanol consumption in adolescent male C57BL/6J mice (Post Natal Day - PND 28-47). In the first experiment, the animals were submitted to the following behavioral tests: Elevated Zero Maze (LZE), Inhibitory Avoidance (ESIN), Pre-Pulse Inhibition (PPI) and Omission in Operant Conditioning (OOC), during adolescence and later in adulthood (DPN64-83), to choose the most sensitive test to evaluate the expression of impulsivity-related behaviors. The data from this evaluation were submitted to Principal Component Analysis (PCA), which indicated that impulsivity-related behaviors were more expressed during adolescence and did not persist into adulthood, and that the OOC was the best test for this evaluation. In the second experiment, different groups of adolescent and adult animals were evaluated using the OOC protocol and the Two-Bottle Choice test to analyze impulsivity, consumption and preference of ethanol (10%). We observed that adolescent mice presented higher impulsivity, consumption and preference for ethanol, even when this substance was adulterated with quinine. Furthermore, through PCA, a correlation between impulsivity and alcohol consumption was demonstrated in adolescent mice. After the behavioral tests, part of the animals were perfused for immunohistochemical analysis of Fos protein and another part was euthanized to analyze the amounts of Synaptopodin and GAD67 proteins in the insula. We did not observe differences between the groups for these analyses. In the third experiment, the animals were submitted to the OOC protocol and then perfused to perform the immunofluorescence technique for Fos, NeuN, and Parvalbumin proteins in the insular cortex. Our results demonstrate that adolescent mice, showed less activation of IPV+ and more neuronal activation in the insula, as well as higher scores of impulsivity-related behaviors. In the fourth experiment, we used the chemogenetic technique to modulate the activity of IPV+, in C57/BL6 transgenic mice (PV-Cre), during the OOC test. We observed IPV+ causality on impulsivity-related behaviors. In the last experiment, we used the same previous procedure to analyze the participation of BTI+ in OOC and operant ethanol consumption. It was observed that activation of these neurons decreased impulsive behavior and ethanol seeking. In conclusion, our results suggest that decreased IPV+ activity, from the insular cortex, seems to be associated with impulsivity and ethanol-seeking related behaviors.
Descrição
Citação
SILVA, Fernando Bezerra Romualdo da. Importância dos interneurônios GABAérgicos Parvalbumina-Positivos do córtex insular para comportamentos relacionados à impulsividade e de procura de etanol. 2023. 171 f. Tese (Doutorado em Farmacologia) - Escola Paulista de Medicina, Universidade Federal de São Paulo (UNIFESP). São Paulo, 2023.