Developmental Changes in the Protein Expression Levels and Localization of the Vesicular Zinc Transporter (ZnT3) in the Mouse Central Auditory System

Vesicular (synaptic) zinc is a neuromodulator that fine-tunes synaptic transmission and sensory processing across many brain areas, including the brainstem, hippocampus, amygdala, and neocortex. Throughout the central auditory system, synaptic zinc plays a crucial role in modulating neurotransmission as well as baseline and adaptive sound processing. However, the developmental changes in the protein expression levels and localization of the vesicular zinc transporter (ZnT3), which loads synaptic zinc into presynaptic vesicles, remain unknown—due in part to the lack of robust ZnT3 antibodies. To address this question, we used the recently developed and validated ZnT3-HA transgenic mouse line, in which the ZnT3 protein contains a C-terminal HA epitope tag. We performed immunohistochemical staining and confocal microscopy in the central auditory system across development to localize and quantify changes in ZnT3 expression and explore potential colocalization of ZnT3 with the vesicular glutamate and GABA transporters VGLUT1 and VGAT, respectively. We found that ZnT3 expression increased significantly between P7 and P14 in both the dorsal cochlear nucleus (DCN) and the auditory cortex (AC), reaching a stable overall expression level and layer distribution by P21. Both in the DCN and AC, ZnT3 was mainly colocalized with VGLUT1. We did not find any significant levels of ZnT3 expression in either the IC or the auditory thalamus. Together, these results highlight major developmental changes in zinc signaling that may affect synaptic transmission and plasticity, as well as normal and pathological sound processing.