An improved immunocytochemical method for subcellular localization of serotonin in rat enterochromaffin cells.
Serotonin-like immunoreactivity (5-HT-LI) has been localized at the ultrastructural level in enterochromaffin (EC) cells of rat gastrointestinal tract. Ultra-thin sections of tissues embedded in epoxy resin were incubated with 5-HT antisera and antibody binding sites were visualized with protein A-gold. Three different antisera were compared and were shown to require different fixation regimens for optimal preservation of 5-HT-LI. For one antiserum, tissues fixed in glutaraldehyde and osmium tetroxide could be used to demonstrate 5-HT-LI in EC cells. Immunocytochemical localization of 5-HT can thus be performed with good ultrastructural preservation of tissues. Quantitative evaluation of the intracellular distribution of 5-HT-LI was performed on EC cells from antrum, duodenum, and proximal colon, fixed in glutaraldehyde only. In all three locations, the majority of the gold particles (90%) in EC cells were localized over the dense core of the secretory granules, while a minor fraction (10%) were localized in parts of the cytoplasm devoid of granules. In EC cells fixed in glutaraldehyde and post-fixed in osmium tetroxide, 5-HT-LI was reduced by about 85%, although intracellular distribution was essentially the same as in cells fixed in glutaraldehyde alone. The results indicate that 5-HT in EC cells is stored mainly in secretory granules, with a small fraction of 5-HT being localized outside the granules.
D000818:Animals; D002838:Chromaffin System / Q000648:ultrastructure*; D003106:Colon / Q000648:ultrastructure; D003593:Cytoplasm / Q000032:analysis; D003594:Cytoplasmic Granules / Q000032:analysis; D004064:Digestive System / Q000648:ultrastructure*; D004386:Duodenum / Q000648:ultrastructure; D004759:Enterochromaffin Cells / Q000648:ultrastructure*; D006046:Gold; D006651:Histocytochemistry; D007106:Immune Sera; D007159:Immunologic Tests; D008297:Male; D008854:Microscopy, Electron; D011706:Pyloric Antrum / Q000648:ultrastructure; D051381:Rats; D011919:Rats, Inbred Strains; D012701:Serotonin / Q000032:analysis* / Q000276:immunology; D013205:Staphylococcal Protein A
D007106:Immune Sera; D013205:Staphylococcal Protein A; D012701:Serotonin; D006046:Gold