Role of aspartate 298 in mouse 5-HT3A receptor gating and modulation by extracellular Ca2+

摘要: The TM2–TM3 extracellular loop is critical for activation of the Cys-loop family ligand-gated ion channels. contribution aspartate 298 (D298), an amino acid that links transmembrane domain 2 (TM2) to loop, in mouse 5-hydroxytryptamine3A (5-HT3A) receptor function was probed with site-directed mutagenesis present study. This negatively charged residue replaced alanine neutralize charge, a glutamate conserve or arginine reverse charge. Human embryonic kidney 293 (HEK 293) cells transfected wild-type and mutant receptors were studied by combining whole-cell patch-clamp recording fast agonist application. D→A D→R mutations resulted reduced 5-HT potency, accelerated kinetics desensitization deactivation. In addition, efficacy partial agonists mutation. D→E mutation produced properties similar those receptor. potential role this modulation calcium ([Ca2+]o) investigated. Increasing [Ca2+]o inhibited 5-HT-activated currents altered manner D298E receptors, alteration eliminated mutations. Our data suggest charge at D298 participates transitions between functional states 5-HT3A receptor, provide evidence side-chain contributes channel gating crucial Ca2+ modulation.