Chemical coding of zinc-enriched neurons in the intramural ganglia of the porcine jejunum

Zinc ions in the synaptic vesicles of zinc-enriched neurons (ZEN) seem to have an important role in normal physiological and pathophysiological processes in target organ innervation. The factor directly responsible for the transport of zinc ions into synaptic vesicles is zinc transporter 3 (ZnT3), a member of the divalent cation zinc transporters and an excellent marker of ZEN neurons. As data concerning the existence of ZEN neurons in the small intestine is lacking, this study was designed to disclose the presence and neurochemical coding of such neurons in the porcine jejunum. Cryostat sections (10 mμ thick) of porcine jejunum were processed for routine double- and triple-immunofluorescence labeling for ZnT3 in various combinations with immunolabeling for other neurochemicals including pan-neuronal marker (PGP9.5), substance P (SP), somatostatin (SOM), vasoactive intestinal peptide (VIP), nitric oxide synthase (NOS), leu-enkephalin (LENK), vesicular acetylcholine transporter (VAChT), neuropeptide Y (NPY), galanin (GAL), and calcitonin-gene related peptide (CGRP). Immunohistochemistry revealed that approximately 39%, 49%, and 45% of all PGP9.5- positive neurons in the jejunal myenteric (MP), outer submucous (OSP), and inner submucous (ISP) plexuses, respectively, were simultaneously ZnT3(+). The majority of ZnT3(+) neurons in all plexuses were also VAChT-positive. Both VAChT-positive and VAChT-negative ZnT3(+) neurons co-expressed a variety of active substances with diverse patterns of co-localization depending on the plexus studied. In the MP, the largest populations among both VAChT-positive and VAChT-negative ZnT3(+) neurons were NOS-positive cells. In the OSP and ISP, substantial subpopulations of ZnT3(+) neurons were VAChT-positive cells co-expressing SOM and GAL, respectively. The broad-spectrum of active substances that co-localize with the ZnT3(+) neurons in the porcine jejunum suggests that ZnT3 takes part in the regulation of various processes in the gut, both in normal physiological and during pathophysiological processes.