Projections of nitric oxide synthase and vasoactive intestinal polypeptide-reactive submucosal neurons in the human colon

Background: The submucosal plexus is important in the control of secretomotor and motor function of the intestine. Our aim was to describe the projections of submucosal neurons to the mucosa within the submucosal plexus and to the circular muscle of human colon and to determine whether submucosal neurons that projected to different layers were located at different levels of the submucosa. Methods: A retrogradely transported fluorescent dye was applied to the mucosa, submucosa or circular muscle layer of human colon which was then maintained in organotypic culture for 5 days. The submucosa was then dissected into two preparations, one containing the inner layer of the submucosal plexus and the other containing both the intermediate and outer layers. The dissected preparations were labelled with antibodies to nitric oxide synthase (NOS) or vasoactive intestinal peptide (VIP). Results: Submucosal neurons projected to the mucosa, submucosa and circular muscle layers for mean distances of 3.7, 3.0 and 4.3 mm, respectively. Ninety-seven per cent of submucosal neurons labelled from the circular muscle were located in the outer or the intermediate layers, while 51% of those projecting to the mucosa were in inner layer and 49% in the intermediate/outer layers of the submucosal plexus. Eleven per cent of submucosal neurons projecting to the circular muscle were immunoreactive for NOS and 12% were immunoreactive for VIP. Forty-five per cent of those projecting within the submucosa were immunoreactive for VIP and 38% of those projecting to the mucosa were immunoreactive for VIP. Conclusions: Submucosal neurons in the human colon innervate the mucosa, circular muscle and submucosa and different functional classes of neurons are located in different layers of the submucosal plexus.