Anatomical organization and origins of VGLUT3-positive axon terminals in the lateral septum.

The lateral septum (LS) integrates afferents from multiple brain regions, including the raphe nuclei. The organization of these inputs contributes to the regionalization of LS functions, for example spatial coding in dorsal LS and emotional regulation in ventral LS. Raphe-LS projections include glutamatergic axons expressing the vesicular glutamate transporter type 3 (VGLUT3), which often form pericellular baskets around LS neurons. This study provides an anatomical characterization of the organization and origins of VGLUT3-positive (VGLUT3+) raphe inputs to the LS. We mapped VGLUT3+ axon terminal density across the rostro-caudal extent of the LS and quantified colocalization with serotonin (5-HT) using immunohistochemistry. Our results showed that VGLUT3 density was highest in the ventral LS, whereas VGLUT3/5-HT colocalization was strongest in the dorsal LS. Retrograde viral vector-mediated tracing identified predominant inputs from the median raphe and B9 neuron group. Interestingly, the ventral hippocampus, a functionally related region which is known to also receive raphe VGLUT3 inputs, showed collaterals with the LS. Additional VGLUT3+ inputs to the LS arose from the interpeduncular nucleus, bed nucleus of the stria terminalis, nucleus incertus and pontine central gray. Anterograde tracing revealed that inputs from these brain regions target distinct and largely non-overlapping domains in the LS. Our findings highlight multiple sources of VGLUT3+ inputs to the LS, beyond the raphe nuclei, and suggest that distinct VGLUT3 circuits could contribute to LS functional specialization.