Anatomical and neurochemical features of the extrinsic and intrinsic innervation of the striated muscle in the porcine esophagus: evidence for regional and species differences

Studies of the intrinsic and extrinsic innervation patterns of esophageal motor endplates (MEPs) are mainly confined to small rodents. Therefore, an immunocytochemical, denervation and tracing study was conducted on the pig, an experimental model in which the distribution of the striated esophageal muscle portion more closely resembles the human situation. The purpose of this study was to analyze the origin and neurochemical content of the nerve fibers participating in the myoneural synapse. Fifteen 6-week-old domestic pigs were studied by immunohistochemistry combined with a-bungarotoxin labeling to define the co-innervation patterns of nitrergic and peptidergic nerve terminals in MEPs. Some animals were subjected to unilateral infra- or supranodose vagotomy to determine the origin of the nerve terminals in MEPs. Special attention was paid to the interregional differences in terms of co-innervation rates, and these findings were compared with literature data on small mammals. Double stainings revealed that most of the nNOS-immunoreactive (ir) terminals in MEN co-stained for VIP, GAL and NPY, but not for PACAP and L-ENK. PACAP- and L-ENK-ir terminals were coarser than nNOS-ir terminals, and largely co-localized VAChT. High percentages of MEN at the cervical level were contacted by PACAP- (similar to94%) and L-ENK-ir (similar to78%) terminals, but the proportion of both decreased in the rostrocaudal direction. Vagotomy significantly reduced their presence in MEPs at the thoracic and abdominal levels, while nNOS-ir terminals observed in approximately 30% of the MEN were unaffected by vagotomy. Immunostainings on brainstem cryosections after retrograde tracing from the cervical esophagus showed that a large number of FB-positive cells in the nucleus ambiguus were PACAP-ir (similar to72%). C-kit-positive interstitial cells of Cajal were seen adjacent to the striated muscle fibers, apparently without direct relationship to MEPs. Similar to mouse esophagus, intrinsic nitrergic fibers were found to run close to, or even spiral around, these interstitial cells, an association that might point to a role as specialized spindle proprioceptors. In conclusion, the cholinergic terminals - part of which coexpress PACAP and/or L-ENK - that innervate MEN in the porcine esophagus have a vagal origin, whereas the nNOS/VIP/GAL/NPY-ir fibers co-innervating these MEN are intrinsic in nature. The regional differences observed along the esophageal length pertain to the neurochemical content of the vagal motor innervation of the MEPs.