Title

Layer-specific local and callosal projections in rat neocortex

Date of Completion

January 2007

Keywords

Biology, Neuroscience

Degree

Ph.D.

Abstract

Neuronal lamination is a characteristic cytoarchitectonic feature of the neocortex and there exists a complex circuitry of connections among neurons found within and between cortical lamina. Interhemispheric cortical connections also exist, made by neurons found in several lamina with axons forming the corpus callosum. An ongoing challenge in understanding normal cortical function, as well as cortical dysfunction due to disease or injury, is the revelation of molecules that regulate lamination in the developing cortex as well as description of the rules of connectivity among neocortical neurons. ^ The present report details our evaluation of intraventricular injection and in utero electroporation as a tool for labeling newly-generated rat cortical neurons. Electroporation of fluorescent proteins targeting early-born neurons reliably resulted in the labeling of neurons found in deep neocortical lamina. The cellular compartments which could be visualized included dendrites and spines, somata, axons and collaterals, and often axon terminals. Electroporation of fluorescent proteins targeting late-born neurons reliably resulted in the labeling of neurons found in upper neocortical lamina. Interestingly, glia including astrocytes, oligodendrocytes, and putative microglia were also labeled. Experiments performed in mice revealed similar results. Using this technique, as well as sequential electroporation which targeted both early and late-born neurons, it was possible to describe the pattern of intracortical and callosal connections made by neurons found in different lamina. ^ Combining electroporation and RNAi targeting Doublecortin (Dcx), it was possible to determine how this molecule regulates neocortical lamination and the development of cortical connections. RNAi of Dcx resulted in disruption in neuronal migration with neurons found in inappropriate lamina as well as trapped in the white matter forming a subcortical band heterotopia. Interestingly, similar experiments in mice also resulted in migration disruption, however, heterotopia were not observed. Intracortical and callosal connections were examined in brains with migration deficits due to Dcx RNAi, which revealed only minor changes in the pattern of callosal connections. ^ Results from the present experiments highlight the use of electroporation and RNAi as tools for visualization and molecular perturbation of neurons in the developing neocortex. As the list of molecules that participate in neocortical development grows, so too will the potential to use these techniques to elucidate the regulation of neocortical lamination and the establishment intracortical and callosal connections. ^