The aim of this study was to examine mitogen-activated protein kinase (ERK1/2) activation in the human neonatal colonic enteric nervous system. For this, we investigated by immunocytochemistry the cellular localization of phosphorylated ERK1/2 (P-ERK) in a series of normal human colon samples removed from newborns and in patients with intestinal obstruction such as Hirschsprung's disease (HSCR), stenosis and atresia. We checked the presence of P-ERK in the three distinct histological layers of normal colon. Phosphorylated ERK was detected in the colonic mucosa, in the enteric nervous system and in endothelial cells. In the mucosa from normal colon, P-ERK was detected at the upper part of the crypt, while P-ERK activation in epithelial cells is altered in HSCR, stenosis and atresia. In the normal colon, strong P-ERK staining was detected in myenteric and submucosal enteric plexuses. Using confocal microscopy analyses, we observed that P-ERK staining was localized in enteric glial cells and not in enteric neurons. Strong P-ERK staining was also observed in plexuses from stenosis and atresia whereas in HSCR, hypertrophic nerve fibres were not stained.