Mechanisms regulating interneuron diversity and maturation

Eunice Kennedy Shriver National Institute Of Child Health & Human Development

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Abstract

HOW THE ENVIRONMENT SCULPTS INTERNEURON DIVERSITY AND MATURATION Interneurons undergo an extensive tangential migration period before reaching their terminal brain region, upon which they interact with the local environment to differentiate and mature. The composition of interneuron subtypes varies significantly between different brain regions. Numerous experiments indicate that general interneuron classes (e.g., parvalbumin- (PV) or somatostatin-expressing (SST)) are determined as cells become postmitotic during embryogenesis, but when other features that define a mature interneuron subtype (neurochemical markers, cell type and subcellular location of synaptic partners, electrophysiology properties, etc.) are established remains unknown. One hypothesis is that interneurons undergo an initial differentiation into cardinal classes during embryogenesis, and maturation into definitive subgroups requires active interaction with their mature environment. An alternate hypothesis is that immature interneurons are already genetically hard-wired into definitive subgroups during embryogenesis and the environment more passively sculpts the maturation of these cells. To test these competing hypotheses, we are harvesting early postnatal interneuron precursors (P0-P2) in specific brain regions and transplanting them into wildtype hosts either homotopically (cortex-to-cortex) or heterotopically (cortex-to-hippocampus or cortex-to-striatum). This technique allows us to determine if transplanted interneurons adopt properties of the host environment (indicating a strong role for the environment in regulating interneuron diversity) or if they retain subtype features more consistent with the donor region. Our initial experiments indicate that the environment largely determines the composition of interneuron subtypes in a brain region regardless of donor region. However, some interneuron subtypes seem to be more genetically predefined and resistant to environmental influences. In the future, we hope to combine this transplantation approach with other analytic techniques to more fully characterize how the environment sculpts interneuron diversity. DEVELOPING A NOVEL APPROACH TO IDENTIFY GENETIC CASCADES UNDERLYING INITIAL INTERNEURON FATE DECISIONS The ability to longitudinally track gene expression within defined populations is essential for understanding how changes in expression mediate both development and plasticity. Previous screens that were designed to identify genes and transcription factors specific to SST- or PV-fated interneurons were largely unsuccessful because several issues significantly hinder these types of studies. First, these interneurons originate from the medial ganglionic eminence (MGE), which is a heterogeneous population of progenitors that gives rise to both interneurons and a variety of GABAergic projection neurons, making it difficult to segregate interneuron progenitors from other cell types. Additionally, many markers that define mature interneuron subtypes are not expressed embryonically, and thus these class-defining markers are not helpful for studying MGE progenitors. In an ideal scenario, we would like to identify actively transcribed genes in MGE progenitors undergoing fate decisions while retaining the capacity to identify whether these cells become PV- or Sst-expressing interneurons in the postnatal brain. To this end, we are developing a spatially and temporally inducible form of DNA adenine methylase identification (DamID) that will allow us to label the transcriptome of MGE progenitors. Labeled cells can be harvested at maturity when we have the tools to distinguish specific interneuron cell types. Then the methylated genomic DNA will be analyzed, allowing us to retrospectively look back in time to identify candidate fate determining genes expressed in specific interneuron populations. Our hope is that this strategy could be widely applicable so that an investigator could characterize the temporal gene expression pattern of their favorite cell type.