ION PERMEATION AND CONDUCTION IN A HUMAN RECOMBINANT 5-HT3 RECEPTOR SUBUNIT (H5-HT3A)

1. A human recombinant homo-oligomeric 5-HT3 receptor (h5-HT3A) expres sed in a human embryonic kidney cell line (HEK 293) was characterized using the whole-cell recording configuration of the patch clamp techni que. 2. 5-KT evoked transient inward currents (EC50 = 3.4 mu M; Hill c oefficient = 1.8) that were blocked by the 5-HT3 receptor antagonist o ndansetron (IC50 = 103 pM) and by the nonselective agents metocloprami de (IC50 = 69 nM), cocaine (IC50 = 459 nM) and (+)-tubocurarine (IC50 = 2.8 mu M). 3. 5-HT-induced currents rectified inwardly and reversed in sign (E5-HT) at a potential of -2.2 mV. N-Methyl-D-glucamine was fi nitely permeant. Permeability ratios P-Na/P-Cs and P-NMDG/P-Cs were 0. 90 and 0.083, respectively. 4. Permeability towards divalent cations w as assessed from measurements of E5-HT in media where Ca2+ and Mg2+ re placed Na+.P-Ca/P-Cs and P-Mg/P-Cs were calculated to be 1.00 and 0.61 , respectively. 5. Single channel chord conductance (gamma) estimated from fluctuation analysis of macroscopic currents increased with membr ane hyperpolarization from 243 fS at -40 mV to 742 fS at -100 mV. 6. R educing [Ca2+](o) from 2 to 0.1 mM caused an increase in the whole-cel l current evoked by 5-HT. A concomitant reduction in [Mg2+](o) produce d further potentiation. Fluctuation analysis indicates that a voltage- independent augmentation of gamma contributes to this phenomenon. 7. T he data indicate that homo-oligomeric receptors composed of h5-HT3A su bunits form inwardly rectifying cation-selective ion channels of low c onductance that are permeable to Ca2+ and Mg2+.