Animal studies suggest a mediator role for neuroendocrine peptides and amines in regulating cell proliferation in the gastrointestinal epithelium. Our aim was to examine the effect of serotonin and selected gastrointestinal peptides on DNA synthesis in a rat and human small intestinal cell line in vitro. IEC-6 and FHs-74 cells were incubated with epidermal growth factor (EGF), insulin-like growth factor II, glucagon, substance P, neurokinin A, calcitonin gene-related peptide (GRP, CCGRP), neurotensin and serotonin. The cells were labelled with [methyl-3H] thymidine and processed for autoradiography. DNA synthesis was evaluated by the labelling index. Epidermal growth factor, insulin-like growth factor II, glucagon, and substance P increased the labelling index in a dose-related manner (P < 0.003). In contrast, a significant dose-dependent reduction of the labelling index was observed after administration of serotonin and neurokinin A (P < 0.0001). Neurotensin and CGRP did not affect the labelling index. EGF, insulin-like growth factor II, glucagon, substance P, serotonin and neurokinin A may be important physiological regulators of proliferation, of gastrointestinal cells.