The development of excitatory neurons in the chick cloaca

Abstract

The enteric nervous system of the chick embryo hindgut is derived from the vagal and sacral neural tube. Nerve cells from these regions produce various neuronal phenotypes, including inhibitory and excitatory nerve cells, providing intrinsic innervation to the chick embryo cloaca. We hypothesised that the vagal and sacral neural tubes provide the cloaca, with phenotypically similar nerve cells. The aim of our study was to investigate the origin of excitatory neurotransmission in the developing cloaca. Chicken embryos were incubated until the 10-12 somite stage (ss). To study the vagal neural tube contribution to the cloaca, this region was microsurgically ablated in ovo and replaced with the corresponding region from age-matched quail embryos. To study the sacral neural tube contribution to the cloaca, the vagal neural tube was ablated at the 10-12 ss, but not replaced with quail neural tube, thus, only the sacral neural crest cells remained. All embryos were harvested at E12 and E14, embedded in paraffin wax and serially sectioned. Immunohistochemistry was carried out on all embryos using human natural killer-1, quail non-chick perinuclear, choline acetyltransferase (ChAT) and substance P (SubP) antibodies. Choline acetyltransferase- and SubP-positive neurons were seen to originate in both the vagal and the sacral neural tube. The vagal neural tube provided the majority of the nerve cells to the chick embryo cloaca and expressed both ChAT and SubP in both the myenteric and submucosal plexus. The sacral neural tube contributed a lesser amount of nerve cells to the chick embryo cloaca, but was also seen to produce both ChAT- and SubP-positive nerve cells in both ganglionated plexus. This study shows, for the first time, that the excitatory ChAT- and SubP-expressing neurons in the developing cloaca originate in both the vagal and the sacral neural tube. These results highlight the origin of phenotypically similar nerve cells in both regions of the neural tube, providing new insights into the developmental origin of the intrinsic innervation of the persistent cloaca.