In vitro continuous amperometric monitoring of 5-hydroxytryptamine release from enterochromaffin cells of the guinea pig ileum

Abstract

A diamond microelectrode was used to sensitively, reproducibly and stably record overflow of 5-hydroxytryptamine (5-HT, serotonin) from enterochromaffin cells (EC) of the intenstinal mucosal layer. 5-HT is an important neurotransmitter and paracrine signalling molecule in the gastrointestinal tract. The diamond microelectrode was formed by overcoating a sharpened 76 µm diameter Pt wire with a thin layer of conducting diamond. After insulation with polypropylene, the conically-shaped microelectrode had a diameter of about 10 µm at the tip and 80 µm at the cylindrical portion. The exposed length was 100–200 µm. Continuous amperometry with the microelectrode poised at a detection potential of 700 mV vs. Ag|AgCl was used to measure 5-HT overflow as an oxidation current. 5-HT overflow was elicited by both mechanical and electrical stimulation. Some minor electrode fouling, a common problem with the oxidative detection of 5-HT, was seen for diamond but the response stabilized enabling recording in vitro. Both 5-HT and the paracrine hormone, melatonin, were detected in the extracellular solution. The 5-HT oxidation current increased in the presence of the serotonin transporter (SERT) inhibitor, fluoxetine (1 µM), providing evidence that the oxidation current was associated with 5-HT.