Neurotransmitter phenotype and axonal projection patterns of VIP-expressing neurons in the inferior colliculus

Neurons in the inferior colliculus (IC), the midbrain hub of the central auditory pathway, send ascending and descending projections to other auditory brain regions, as well as projections to other sensory and non-sensory brain regions. However, the axonal projection patterns of individual classes of IC neurons remain largely unknown. Vasoactive intestinal polypeptide (VIP) is a neuropeptide expressed by subsets of neurons in many brain regions. We recently identified a class of IC stellate neurons that we called VIP neurons because they are labeled by tdTomato (tdT) expression in VIP-IRES-Cre x Ai14 mice. Here, using fluorescence in situ hybridization, we found that tdT+ neurons in VIP-IRES-Cre x Ai14 mice express Vglut2, a marker of glutamatergic neurons, and VIP, suggesting that VIP neurons use both glutamatergic and VIPergic signaling to influence their postsynaptic targets. Next, using viral transfections with a Cre-dependent eGFP construct, we labeled the axonal projections of VIP neurons. As a group, VIP neurons project intrinsically, within the ipsilateral and contralateral IC, and extrinsically to all the major targets of the IC. Within the auditory system, VIP neurons sent axons and formed axonal boutons in higher centers, including the medial geniculate nucleus and the nucleus of the brachium of the IC. Less dense projections terminated in lower centers, including the nuclei of the lateral lemniscus, superior olivary complex, and dorsal cochlear nucleus. VIP neurons also project to several non-auditory brain regions, including the superior colliculus, periaqueductal gray, and cuneiform nucleus. The diversity of VIP projections compared to the homogeneity of VIP neuron intrinsic properties suggests that VIP neurons play a conserved role at the microcircuit level, likely involving neuromodulation through glutamatergic and VIPergic signaling, but support diverse functions at the systems level through their participation in different projection pathways.