Ca2+ permeability of human heteromeric nAChRs expressed by transfection in human cells

The Ca²⁺ permeability of the human heteromeric α3β4, α4β2 and α4β4 neuronal nicotinic acetylcholine receptors (nAChRs) was estimated by measuring the fractional Ca²⁺ current (P_f) flowing through the ligand-activated receptor-channels. Simultaneous recordings of transmembrane currents and fluorescence transients, using the whole-cell patch-clamp technique combined with fura-2 fluorescence microscopy, were performed in transiently transfected human cells. The human α4β2 nAChR showed a P_f value of 2.6%, while the human α3β4 nAChR showed a similar P_f value of 2.7%. Conversely, α4β4 nAChR exhibited a P_f value (1.5%) significantly smaller than those of both α4β2 and α3β4 nAChRs. In test experiments performed in HEK 293 cells stably expressing rat GluR1 AMPA receptor subunit, we repeated the determination of P_f, whose value (3.2%) has previously been reported by others using the same fluorescent dye; and we found a very similar P_f value (3.5%). In further test experiments, we found that P_f values of chick α3β4 (4.4%) and α4β4 (2.1 %) matched those previously reported by us using confocal fluorescence microscopy. Thus, our findings are consistent with those elsewhere reported even using different experimental procedures, giving a strong support to the following sequence of Ca²⁺ permeability: h-α3β4 > h-α4β2 > h-αAβ4.