Estratégias de investigação in vitro e ex vivo dos efeitos do canabidiol sobre aspectos morfológicos e funcionais de células neurais
Data
2021-03-08
Autores
Quintella, Miguel Lins 
Orientadores
Monteiro, Beatriz de Oliveira
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
Introdução: Evidências recentes indicam que o canabidiol (CBD) possui ação neuroprotetora e anti-inflamatória, podendo atuar também como anticonvulsivante, antioxidante e ansiolítico. No entanto, alguns estudos têm demonstrado efeitos tóxicos e pró-apoptóticos do CBD, e o modo pelo qual o CBD atua sobre o sistema nervoso central ainda é desconhecido. Portanto, para compreender esses efeitos contraditórios uma possível estratégia é investigar a ação do CBD in vitro.
Objetivo: Assim, o presente estudo teve como proposta analisar in vitro, em células neurais e em fatias (slices) de cérebro, os efeitos do CBD em aspectos morfológicos da doença de Alzheimer (AD), e funcionais da dependência química.
Métodos: A tese foi organizada em dois capítulos. No capítulo 1 investigamos as alterações evidenciadas na formação das esferas, migração, crescimento de neuritos e morte neuronal de células neuroprogenitoras (NPC) e de células de neuroblastoma (N2a), na presença do CBD. O peptídeo β-amiloide (Aβ) foi adicionado à cultura como modelo de uma situação patológica simulando aspectos do processo inflamatório da AD e da toxicidade de Aβ. No capítulo 2, investigamos a influência do CBD na liberação de dopamina em fatias do estriado de camundongos tratados com CBD, avaliada por voltametria cíclica, mimetizando um aspecto da dependência química. Assim, foram utilizadas técnicas in vitro de culturas de neuroesferas e de células N2a, e ferramenta eletroquímica de voltametria cíclica de registro em slice, para avaliar aspectos morfológicos e funcionais das possíveis alterações causadas pelo tratamento com CBD.
Resultados: Verificamos que o CBD exerce efeito no crescimento e formação de esferas, além de promover morte celular e reduzir a migração e o brotamento de neuritos. Além disso, células tratadas com Aβ também apresentaram redução da viabilidade celular, do crescimento da neuroesfera e de neuritos. As análises por voltametria mostraram não haver diferenças nos níveis de dopamina liberada no estriado de animais tratados com CBD. Em culturas de NPC e N2a foi possível identificar alterações morfológicas promovidas pelo tratamento com CBD. Porém, as análises por voltametria cíclica indicaram que o tratamento prévio com CBD não alterou a liberação de dopamina no estriado.
Conclusão: CBD modificou o padrão de viabilidade, proliferação, migração e projeções de neuritos, que poderiam potencializar as alterações presentes na AD. Este estudo oferece insights in vitro sobre os efeitos do CBD.
Introduction: Recent evidence indicates that cannabidiol (CBD) has neuroprotective and anti-inflammatory effects, and can also act as an anticonvulsant, antioxidant, and anxiolytic substance. However, some studies have shown toxic and pro-apoptotic effects of CBD, and the way by which CBD acts on the central nervous system is still unknown. Therefore, to understand these contradictory effects, a possible strategy is to investigate the action of CBD in vitro. Objectives: The present study aims to analyze in vitro and ex vivo, in neural cells and brain slices, the effects of CBD on morphological aspects of Alzheimer's disease (AD), and functional aspects of chemical dependency. Methods: The thesis was organized into two chapters. In chapter 1 we investigated changes evidenced in the formation of spheres, migration, neurite growth and neuronal death of neural progenitor cells (NPC) and neuroblastoma cells (N2a) in the presence of CBD. The amyloid-β peptide (Aβ) was added to the culture as a model of a pathological situation simulating aspects of the inflammatory process of AD and Aβ toxicity. In chapter 2, we investigated the influence of CBD on the dopamine release in slices of the striatum of mice treated with CBD, assessed by cyclic voltammetry, mimicking one aspect of chemical dependence. Thus, in vitro techniques of neurosphere and N2a cell culture, as well as electrochemical cyclic voltammetry of slice recording, were utilized to evaluate morphological and functional aspects of possible changes caused by treatment with CBD. Results: We found that CBD decreases growth and formation of neurospheres, promotes cell death, reduces cell migration and neurite sprouting. Cells treated with Aβ also shows reduced cell viability, neurosphere growth and neurite sprouting. Voltammetry analysis show no difference in levels of dopamine release in the striatum of animals treated with CBD. Thus, in NPC and N2a cultures it was possible to identify morphological alterations promoted by treatment with CBD. However, cyclic voltammetry analysis indicated that previous treatment with CBD did not alter the release of dopamine in the striatum. Conclusion: CBD modified the pattern of viability, proliferation, migration, and projections of neurites, which can potentiate the changes present in the AD. This study provides in vitro insights about the effects of CBD.
Introduction: Recent evidence indicates that cannabidiol (CBD) has neuroprotective and anti-inflammatory effects, and can also act as an anticonvulsant, antioxidant, and anxiolytic substance. However, some studies have shown toxic and pro-apoptotic effects of CBD, and the way by which CBD acts on the central nervous system is still unknown. Therefore, to understand these contradictory effects, a possible strategy is to investigate the action of CBD in vitro. Objectives: The present study aims to analyze in vitro and ex vivo, in neural cells and brain slices, the effects of CBD on morphological aspects of Alzheimer's disease (AD), and functional aspects of chemical dependency. Methods: The thesis was organized into two chapters. In chapter 1 we investigated changes evidenced in the formation of spheres, migration, neurite growth and neuronal death of neural progenitor cells (NPC) and neuroblastoma cells (N2a) in the presence of CBD. The amyloid-β peptide (Aβ) was added to the culture as a model of a pathological situation simulating aspects of the inflammatory process of AD and Aβ toxicity. In chapter 2, we investigated the influence of CBD on the dopamine release in slices of the striatum of mice treated with CBD, assessed by cyclic voltammetry, mimicking one aspect of chemical dependence. Thus, in vitro techniques of neurosphere and N2a cell culture, as well as electrochemical cyclic voltammetry of slice recording, were utilized to evaluate morphological and functional aspects of possible changes caused by treatment with CBD. Results: We found that CBD decreases growth and formation of neurospheres, promotes cell death, reduces cell migration and neurite sprouting. Cells treated with Aβ also shows reduced cell viability, neurosphere growth and neurite sprouting. Voltammetry analysis show no difference in levels of dopamine release in the striatum of animals treated with CBD. Thus, in NPC and N2a cultures it was possible to identify morphological alterations promoted by treatment with CBD. However, cyclic voltammetry analysis indicated that previous treatment with CBD did not alter the release of dopamine in the striatum. Conclusion: CBD modified the pattern of viability, proliferation, migration, and projections of neurites, which can potentiate the changes present in the AD. This study provides in vitro insights about the effects of CBD.