Development and birthdates of vasoactive intestinal peptide immunoreactive neurons in the chick proventriculus.

To gain insight into the mechanisms regulating expression of transmitter phenotypes in the enteric nervous system, we have studied the development and birthdate of vasoactive intestinal peptide immunoreactive (VIP-IR) myenteric neurons in the chicken proventriculus (secretory portion of the avian stomach) by a combination of immunocytochemistry and radioautography. The appearance and numbers of VIP-IR neurons in whole mounts of the myenteric plexus from chick embryos and chickens were examined. We found that VIP-IR neurons first appeared at embryonic day (E) 5.5-6.5 in the distal part of the proventriculus. At E7.5, VIP-IR neurons were found singly, in pairs, or in small groups, which together with unlabeled cells formed primitive myenteric ganglia. VIP-IR fibers were found within the developing fiber tracts which connected the ganglia. The number of VIP-IR neurons was found to be maximum in the E15.5 embryo and to decline to 68% of maximum in the 4 week old chicken. Birthdate studies were performed by application of either single pulses or cumulative doses of [3H]-thymidine to embryos between E3 and E14. Whole mounts of the myenteric plexus from the proventriculus of these embryos were immunostained for VIP at E10 or E17. The whole mounts were subsequently sectioned and processed for radioautography. We found that VIP-IR myenteric neurons were born between E3 and E10 with a peak at E7. Most cells underwent terminal division between E5 and E9. These data will be useful in determining the time and conditions when cells make decisions about transmitter phenotypes.