Inhibition of in vitro enteric neuronal development by endothelin-3: mediation by endothelin B receptors

The terminal colon is aganglionic in mice lacking endothelin-3 or its recep tor, endothelin B. To analyze the effects of endothelin-3/endothelin B on t he differentiation of enteric neurons, E11-13 mouse gut was dissociated, an d positive and negative immunoselection with antibodies to p75(NTR) were us ed to isolate neural crest- and non-crest-derived cells. mRNA encoding endo thelin B was present in both the crest-and non-crest-derived cells, but tha t encoding preproendothelin-3 was detected only in the noncrest-derived pop ulation. The crest- and non-crest-derived cells were exposed in vitro to en dothelin-3, IRL 1620 (an endothelin B agonist), and/or BQ 788 (an endotheli n B antagonist). Neurons and glia developed only in cultures of crest-deriv ed cells, and did so even when endothelin-3 was absent and BQ 788 was prese nt. Endothelin-3 inhibited neuronal development, an effect that was mimicke d by IRL 1620 and blocked by BQ 788, Endothelin-3 failed to stimulate the i ncorporation of [H-3]thymidine or bromodeoxyuridine, Smooth muscle developm ent in noncrest-derived cell cultures was promoted by endothelin-3 and inhi bited by BQ 788, In contrast, transcription of laminin ctl, a smooth muscle -derived promoter of neuronal development, was inhibited by endothelin-3, b ut promoted by BQ 788. Neurons did not develop in explants of the terminal bowel of E12 Is/ls (endothelin-3-deficient) mice, but could be induced to d o so by endothelin-3 if a source of neural precursors was present. We sugge st that endothelin-3/endothelin B normally prevents the premature different iation of crest-derived precursors migrating to and within the fetal bowel, enabling the precursor population to persist long enough to finish coloniz ing the bowel.