Inactivation properties of human recombinant class E calcium channels

The electrophysiological and pharmacological properties of alpha(1E)-conrai ning Ca2+ channels were investigated by using the patch-clamp technique in the whole cell configuration, in HEK 293 cells stably expressing the human alpha(1E) together with alpha(2b), and beta(1b) accessory subunits. These c hannels had current-voltage (I-V) characteristics resembling those of high- voltage-activated (HVA) Ca2+ channels (threshold at -30 mV and peak amplitu de at +10 mV in 5 mM Ca2+). The currents activated and deactivated with a f ast rate, in a time- and voltage-dependent manner. No difference was found in their relative permeability to Ca2+ and Ba2+. Inorganic Ca2+ channel blo ckers: (Cd2+, Ni2+) blocked completely and potently the alpha(1E),/alpha(2b )delta/beta(1b) beta(1b) mediated currents (IC50 = 4 and 24.6 mu M, respect ively). alpha(1E)- mediated currents inactivated rapidly and mainly in a no n-Ca2+ dependent manner, as evidenced by the fact that I) decreasing extrac ellular Ca2+ from 10 to 2 mM and 2) changing the intracellular concentratio n of the Ca2+ chelator 1.2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid (BAPTA), did not affect the inactivation characteristics; 3) there wa s no clear-cut bell-shaped relationship between test potential and inactiva tion, as would be expected from a Ca2+-dependent event. Although Ba2+ subst itution did not affect the inactivation of alpha(IE) channels, Na+ substitu tion revealed a small but significant reduction in the extent and rate of i nactivation, suggesting that besides the presence of dominant voltage-depen dent inactivation, alpha(IE) channels are also affected by a divalent catio n-dependent inactivation process. We have analyzed the Ca2+ currents produc ed by a range of imposed action potential-like voltage protocols (APVPs). T he amplitude and area of the current were dependent on the duration of the waveform employed and were relatively similar to those described for HVA Ca lcium channels. However, the peak latency resembled that obtained for low-v oltage-activated (LVA) calcium channels. Short bursts of APVPs applied at 1 00 Hz produced a depression of the Ca2+ current amplitude, suggesting an ac cumulation of inactivation likely to be calcium dependent. The human alpha( IE) gene seems to participate to a Ca2+ channel type with biophysical and p harmacological properties partly resembling those of LVA and those of HVA c hannels, with inactivation characteristics more complex than previously bel ieved.