Title: Molecular and cellular mechanisms of axon collateral development in the murine neocortex

     In the cerebral cortex, spatial regulation of axon targeting and collateral elaboration is critical for achieving ordered connectivity among distinct cell types. Layer 2/3 (L2/3) cortical projection neurons (CPNs) in primary somatosensory (S1) cortex display laminar-specific axon branching, extending collaterals into layers 5 and 2/3, but not 4. CPN axons are also involved in long-range targeting of homotypic contralateral cortical areas. We reasoned that genes specifically enriched in L2/3 neurons during early postnatal development are likely determinants of L2/3-specific axon projection patterns. To screen for their function in regulating L2/3 CPN axon collateral formation and long-range targeting, we employed inducible, cell-autonomous manipulation of gene-function in vivo combined with robust sparse labeling of individual neurons.

      We find that ablation of the transcription factor Mef2c in S1 L2/3 CPNs leads to defects in the innervation of long-range targets in the contralateral hemisphere. Though Mef2c is known to regulate cortical neuron cell-fate determination and survival, to our knowledge this is a first demonstration of a post-mitotic role in axon targeting. We are currently employing conditional Mef2c mutants to separate L2/3-autonomous functions in axon targeting from non-autonomous roles in cortical patterning. Further, we are interrogating Mef2c function in regulating contralateral axon targeting through its potential downstream targets, including components of ephrin/EphR signaling.

    We are also investigating axon collateral development in layer 6 corticothalamic neurons (L6CThNs), which elaborate intracortical collaterals in a pattern complementary to L2/3 neurons. After addressing technical challenges to achieve sparse labeling of L6CThNs during postnatal development, we are employing brain clearing and high-resolution lightsheet imaging to reconstruct their local axon collaterals during the first two weeks of postnatal development. Comparing molecular mechanisms that distinguish L6CThN development from L2/3 CPNs will further our understanding of intracellular signaling events and extrinsic spatial cues that produce distinct morphological outcomes in different cortical cell types.