GLUN3A DELETION DISRUPTS REGION- AND LAMINAR-SPECIFIC TARGETING OF INTERHEMISPHERIC CALLOSAL AXONS

Callosal projections are long-range axonal projections that connect the two cortical hemispheres via the corpus callosum, allowing bilateral integration of sensory information. Callosal axons originate mainly from layer (L) 2/3 pyramidal neurons in primary sensory areas and project to homotypic regions of the contralateral hemisphere. Accurate information transfer relies on a highly stereotyped region and layer-specific axonal pattern, targeting distinct dendritic domains of contralateral L2/3 neurons. In mouse somatosensory cortex such precise innervation is established during the second and third postnatal weeks but the molecular determinants are largely unknown. Using in utero electroporation of fluorescent reporters to label L2/3 neurons, we show that loss-of-function of Grin3a (gene encoding the non-conventional NMDA receptor subunit GluN3A) disrupts region and layer-specific callosal axon targeting without altering early axonal navigation steps. Rather than concentrating in the border between primary/secondary somatosensory cortex (S1/S2), callosal axons in GluN3A knockouts form an ectopic second column laterally in S2. Within the S1/S2 border, axonal arbors fail to innervate their normal destinations in L1 and are shifted towards inner regions of L2/3. Conditional loss of function experiments, together with altered apical dendritic branching in Grin3a knockouts, suggest that the dendritic architecture of postsynaptic L2/3 neurons guides callosal axon patterns during critical periods of circuit maturation and implicate GluN3A NMDA receptor subunits in determining the timing and specificity of this process. Finally, we identify GluN3A modulation of Kcna1 and Crmp4 expression as downstream targets, linking their tempo of expression with the spatial precision of dendritic arbor development.