The Diversity of Cortical Interneurons
A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy
The cortex is involved in diverse higher cognitive processes including decision making, motor planning, sensory discrimination, and memory consolidation. The cortical interneurons are key elements of the cortical system. These interneurons stabilize networks, but at the same time they also add non-linear effects to the excitatory system to make the cortical network more dynamic. To achieve this, cortical interneurons form a very heterogeneous group, making it hard to classify them without markers. We took a BACtrap approach performing translating ribosome affinity purifications on transgenic mice with Bacterial artificial chromosome, for systematic discovery of markers for different cell types. First, we generated BACtrap lines for known markers of mixed interneuron populations. After IHC (immnohistochemistry) characterization of each line, we picked 4 lines for Dlx1, Nek7, Htr3a and Cort genes for further studies. We collected mRNAs from targeted neurons in each line and performed gene profiling. Based on IHC and gene profiling studies, we found that each of the 4 lines labeled different but overlapping interneuron populations in the cortex. Second, we performed a comparative microarray analysis to find genes that showed differential enrichment in each of the 4 populations and we found ~20 genes as candidate marker genes. To examine their potential role as marker genes, we generated BAC transgenic mice for these candidate genes and also examined their DRP (Density Recovery Profile) on ISH images from the ABA (Allen Brain Atlas). We found that a number of candidate genes showed regular spacing of cell bodies, suggesting that those genes might label a functionally homogenous group. Third, we characterized new Cre lines for candidate marker genes, Rbp4 and Oxtr, to investigate their cell types and functional roles by using Cre/loxP system. Both Rbp4 and Oxtr Cre are heterogeneous in terms of their neurochemical profiles, but DRP analysis on Oxtr Cre neurons suggested their potential to be a functionally homogenous group. Cre dependent AAV injection also revealed a tiling property of Oxtr Cre neurons in the somatosensory cortex. Connectivity of three different Cre lines (Rbp4, Oxtr, Htr3a) was also examined using retrograde monosynaptic rabies virus tracers. Although three lines expressed Cre proteins in different interneuron populations, the presynaptic inputs were almost identical except for a few differences. However, each line had a different preference in inputs and we found line specific inputs from the hippocampus and the dopaminergic nuclei. In short, we carried out systematic marker searches and the generation of transgenic mice. Our findings suggest the existence of better markers for interneuron cell types, and we also showed that a group that is heterogeneous at the cellular level could work as a functionally homologous group. New interneuronal Cre lines showed a few differences in presynaptic inputs and created new opportunities for us to understand the functional differences of distinct cell types.