Diane Sherman

Development of serotonergic neurons in the chick duodenum

The ontogenetic sequence of neuron formation has been examined in the duodenum of the developing chick. Enteric serotonergic neurons were selected as an example of an intrinsic neuron, and the adrenergic innervation was studied as an example of an extrinsic supply. Light and electron microscopic radioautography following incubation with tritiated serotonin (3H-5-HT) was used to identify serotonergic elements, and histofluorescence was used to detect adrenergic neurites. At 7 days of incubation (Day E7) the primitive myenteric plexus had formed and the gut is known to contain cholinergic elements [Smith, J.,et al. (1977).Cell Different.6, 199–216]. Nevertheless, in the duodenum there was no uptake of3H-5-HT outside the mucosa. However, on Days E8–E9 many primitive-appearing cells, containing abundant free ribosomes, located in the outer gut mesenchyme surrounding the developing islands of myenteric plexus, took up3H-5-HT. In addition, there was also labeling of axons and growth cones within the myenteric plexus. As development proceeded and the gut matured, the number of labeled cells outside the enteric plexuses declined (rapidly between Days E9 and E13) and the amount of labeling of axons within the plexuses correspondingly increased. Labeling was found in the submucosal plexus on Day E11; substantial numbers of mature-appearing varicosities were labeled by Day E18; the labeled cells in the mesenchyme outside the myenteric plexus disappeared after Day E18. Cells labeled by3H-5-HT were found apparently entering the myenteric plexus from the surrounding mesenchyme. This suggested the possibility that the labeled cells in the mesenchyme were precursors of serotonergic neurons that enter the myenteric plexus and then extend an axon. After this occurs the uptake of3H-5-HT may be lost from the cell bodies and restricted to axons. Adrenergic neurites were first detected in the gut on Day E12. Thus, a sequential pattern of development of the enteric innervation of a single intestinal region was found. Cholinergic neurons are followed by serotonergic neurons, and both precede the ingrowth of the adrenergic innervation. Sequential development of neurons may be important to the expression of the multiplicity of enteric neuronal phenotypes.

Development of the monoaminergic innervation of the avian gut: transient and permanent expression of phenotypic markers

Specific cellular accumulation of [3H]5-hydroxytryptamine ([3H]5-HT) occurs during development of the avian gut. This accumulation is transient in extraganglionic mesenchymal cells (TES cells) but is a permanent characteristic of enteric serotonergic neurons (ESN). Species-specific differences were found in the location of TES cells and ESN. In chicks TES cells surrounded myenteric ganglia and ESN were restricted to the myenteric plexus. In quails TES cells surrounded submucosal ganglia and [3H]5-HT-labeled submucosal as well as myenteric neurons. [3H]Norepinephrine accumulated only in noradrenergic terminals and not in TES cells or ESN. The origins of TES cells and ESN were studied in chimeras, in which neuraxis from appropriate or inappropriate axial levels was grafted from quail to chick. Both types of chimeric bowel contained TES cells and ESN. Most TES cells in chimeras were chick in origin and distributed as in chicks (around myenteric ganglia); however, some TES cells and all ESN were quail cells. To test whether crest cells are required for development of TES cells and ESN, aneuronal chick hindgut was explanted and grown alone, or with quail neuraxis, as chorioallantoic membrane (CAM) grafts. TES cells appeared in CAM grafts whether or not crest cells were present; however ESN only appeared in explants when quail neuraxis was included. In addition, an ectopic [3H]5-HT-labeled chromaffin-like cell, also of quail origin, was found in enteric plexuses in these combined explants of crest and gut. Most TES cells, therefore, are neither derived from nor dependent on the presence of crest cells in the gut wall. Since even an inappropriate axial level of crest was found to produce ESN when it was experimentally induced to colonize the bowel the enteric microenvironment probably plays a critical role in serotonergic neural development. The species-specific location of TES cells and ESN is consistent with the hypothesis that TES cells constitute an important component of this microenvironment.