Immunohistochemical and Biochemical Evidence for the Presence of Serotonin in Amphibian Adrenal Chromaffin Cells

The presence of serotonin (5-HT) in chromaffin cells of the frog adrenal (inter-renal) gland has been demonstrated both by immunocytochemical and biochemical techniques. Using antisera against 5-HT and tyrosine hydroxylase (TH) on consecutive sections, we found by means of the indirect immunofluorescence technique that a majority of chromaffin cells were also immunopositive for 5-HT. When antibodies to 5-HT and phenylethanolamine-N-methyltransferase (PNMT) were applied on consecutive sections, 5-HT-like immunoreactivity was observed in almost all epinephrine-producing cells which represented about 90% of the total chromaffin cells. No 5-HT-containing fibres could be detected. At the ultrastructural level, using a pre-embedding procedure associated with gold-silver intensification of the immunoperoxidase reaction, 5-HT-immunoreactivity was visualized in secretory vesicles essentially located in the periphery of epinephrine cells. Combination of high performance liquid chromatography and electrochemical detection showed the presence of both 5-HT and its metabolite 5-hydroxyindolacetic acid (5-HIAA) in frog adrenal extracts. Transection of the splanchnic nerve enhanced 5-HT immunoreactivity and augmented the amount of 5-HT in adrenal extracts. Taken together, these results indicate that epinephrine-producing cells of the frog adrenal contain significant amounts of serotonin. The observation of the storage of 5-HT in secretory vesicles of epinephrine cells suggests that serotonin may be released with catecholamines under stress conditions.