RESUMO
The techniques evolving from the rapidly developing field of small RNAs promise accessible approaches to dissecting cellular and molecular mechanisms of higher brain function. Here, a current overview of the technology is presented, along with an outline of how these approaches might help neuroscientists to more rapidly uncover the cellular and molecular bases of behavior.
Assuntos
Encéfalo/fisiologia , Neurônios/fisiologia , Interferência de RNA/fisiologia , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , Animais , Marcação de Genes/métodos , Marcação de Genes/tendências , Vetores Genéticos/genética , Humanos , Modelos Animais , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , RNA Mensageiro/genéticaRESUMO
In vivo cellular microenvironments are not well-mimicked in present in vitro cell culture systems. Microtechnology, and microfluidics in particular, provides the tools to create in vivo-like cellular microenvironments in vitro. Features of in vitro cellular microenvironments are discussed and compared to macroscale cell culture environments; the concept of an effective culture volume (ECV) is introduced to facilitate the comparison. Current research using microtechnology to investigate in vitro cellular microenvironments is presented and areas where more research is needed in characterizing the in vitro microenvironment are outlined.
Assuntos
Técnicas de Cultura de Células/métodos , Microfluídica/métodos , Adsorção , Animais , Materiais Biocompatíveis/química , Reatores Biológicos , Adesão Celular , Técnicas de Cultura de Células/instrumentação , Células Cultivadas , Difusão , Desenho de Equipamento , Microfluídica/instrumentação , Estresse MecânicoRESUMO
Persistent activity in the brain is involved in working memory and motor planning. The ability of the brain to hold information 'online' long after an initiating stimulus is a hallmark of brain areas such as the prefrontal cortex. Recurrent network loops such as the thalamocortical loop and reciprocal loops in the cortex are potential substrates that can support such activity. However, native brain circuitry makes it difficult to study mechanisms underlying such persistent activity. Here we propose a platform to study synaptic mechanisms of such persistent activity by constraining neuronal networks to a recurrent loop like geometry. Using a polymer stamping technique, adhesive proteins are transferred onto glass substrates in a precise ring shape. Primary rat hippocampal cultures were capable of forming ring-shaped networks containing 40-60 neurons. Calcium imaging of these networks show evoked persistent activity in an all-or-none manner. Blocking inhibition with bicuculline methaiodide (BMI) leads to an increase in the duration of persistent activity. These persistent phases were abolished by blockade of asynchronous neurotransmitter release by ethylene glycol tetraacetic acid (EGTA-AM).