GABA-mediated inhibition is crucial for the function and plasticity of neural circuits in neocortex, but the cellular diversity of the GABAergic system has been resistant to conventional anatomical, physiological, and genomic approaches. For example, maturation of inhibitory circuitry in primary visual cortex (V1) promotes the development of ocular dominance (OD) plasticity and its critical period, but the cellular and molecular mechanisms underlying the developmental and experience-dependent maturation of GABA interneurons remain poorly understood. Analysis of gene expression profiles should provide fundamental insights into this issue, but the heterogeneity of GABA interneurons has precluded such studies. Here we propose a genetic strategy to implement a novel method for cell type-based analysis of gene expression in GABAergic system and in complex brain tissues in general. Using Cre/loxP-regulated gene expression strategy, we will generate knockin mice expressing an epitope (FLAG)-tagged polyA binding protein (PABP) in different classes of interneurons. Actively translated mRNAs from interneurons in V1 will be harvested by co-immunoprecipitation against the FLAG peptide and subjected to microarray analysis. Three major classes of interneurons will be analyzed during the critical period of OD plasticity. Since the critical period can be either accelerated or delayed by BDNF (brain-derived neurotrophic factor) overexpression or dark-rearing (DR), respectively, the same cell types will be analyzed in BDNF transgenic and dark-reared mice.
We aim to identify genes in specific cell types whose expression 1) correlate with the progression of critical period, 2) are regulated by BDNF and DR in a manner that corresponds to alterations in critical period. These studies will provide a comprehensive picture of transcriptomes in defined GABAergic cell types during the critical period, unravel the molecular mechanisms which direct experience-dependent maturation of inhibitory interneurons, and guide future experiments to examine specific cellular processes. Because cell types are functional units of neural circuits, this approach will 1) substantially increase the sensitivity for detecting alterations of gene expression in complex brain tissues, 2) allow meaningful interpretation of gene expression data in the context of neural circuit development and function, 3) greatly enhance the power of gene expression analysis in system neuroscience and in mouse model of brain disorders.
Here we proposed to establish a cell type-based genetic strategy to study gene expression profiles in mouse brain. This method will greatly enhance the sensitivity and explanatory power of genomic studies of mouse model of brain disorders, such as Rett Syndrome, schizophrenia, and autism.