Localization of zinc-enriched neurons in the mouse peripheral sympathetic system

Growing evidence supports the notion that zinc ions located in the synaptic vesicles of zinc-enriched neurons (ZEN) play important physiological roles and are involved in certain pathological changes in the central nervous system. Here we present data revealing the distribution of zinc ions and the co-localization of zinc transporter 3 (ZnT3) and tyrosine hydroxylase (TH) in crush-operated sciatic nerves and lumbar sympathetic ganglia of mice, using zinc selenide autometallography (ZnSe(AMG)) and ZnT3 immunofluorescence combined with confocal scanning microscopy, respectively. Six hours after the crush operation, ZnSe(AMG) grains and ZnT3 immunoreactivity were predominantly present in a subpopulation of thin unmyelinated sciatic nerve axons. In order to identify the type(s) of ZEN axons involved, double labeling with ZnT3 and (1) TH, (2) vesicular acetylcholine transporter (VAChT), (3) calcitonin gene-related peptide (CGRP), and (4) neuropeptide Y (NPY) was performed. Confocal microscopic observations showed that ZnT3 was located in a subpopulation of sciatic axons in distended parts proximal and distal to the crush site. Most, if not all, ZnT3-positive axons contained TH immunofluorescence, a few showed co-localization of ZnT3 and VAChT with very weak immunostaining, while no congruence was observed between ZnT3 and CGRP or NPY. Studies of the lumbar sympathetic ganglia showed that not more than 5% of the neurons were ZnT3-positive and that almost all of these were TH-positive. Furthermore, approximately 5% of total lumbar sympathetic ganglionic cells were ZnSe(AMG) positive, 48 h after a local injection of sodium selenide into the sciatic nerve. The present data support the notion that a subgroup of mouse sympathetic postganglionic neurons are ZEN neurons.