Peculiarities of Selectivity of Three Subtypes of Low-Threshold T-Type Calcium Channels

Abstract

Despite the progress in studies of the properties and functions of low-threshold calcium channels (LTCCs) [1], the mechanisms of their selectivity and permeability remain unstudied in detail. We performed a comparative analysis of the selectivity of three cloned pore-forming LTCC subunits (α1G, α1H, and α1I) functionally expressed in Xenopus oocytes with respect to bivalent alkaline-earth metal cations (Ва 2+ , Са 2+ , and Sr 2+ ). The relative conductivities (G) of these channels were determined according to the amplitudes of macroscopic currents (I) and potentials of zero currents (E). The currents were recorded after preliminary intracellular injection of a fast calcium buffer, BАРТА, in order to suppress the endogenous calciumdependent chloride conductivity. Channels formed by α1G subunits demonstrated the following ratios of the amplitudes of macroscopic currents and potentials of zero current: I Ca :I Ba :I Sr = 1.00:0.75:1.12 and E Ca ≈ E Ba ≈ E Sr . For channels that were formed by α1H and α1I subunits, these ratios were as follows: I Ca :I Ba :I Sr = 1.00:1.20:1.17, E Ca ≈ E Ba ≈ E Sr and I Ca :I Ba :I Sr = 1.00:1.48:1.45, E Ca ≈ E Ba ≈ E Sr , respectively. The different macroscopic conductivities and similar potentials of zero current typical of α1G and α1І channels indicate that, probably, various bivalent cations can in a differential manner influence the stochastic parameters of functioning of these channels. At the same time, channels formed by α1Н subunits are characterized by more positive potentials of zero current for Са 2+ . It seems possible that the selectivity of the above channels is determined by mechanisms that mediate the selectivity of most high-threshold calcium channels (more affine binding of Са 2+ inside the pore).

Sodium/calcium selectivity of cloned calcium T-type channels

“…This is why they are to a lesser extent selective with respect to Са 2+ than HTCCs. As compared with the HTCCs, cloned LTCCs are also characterized by nearly equal permeabilities for Са 2+ , Sr 2+ , and Ва 2+ [15]. In amino acid sequences of Р loops of the Са v 3 channels, there are two residues of glutamate and two aspartate residues (D, also charged negatively) instead of four glutamate residues that form a selective filter of the L-type HTCCs, [7].…”

“…Procedures of preparation of matrix RNA (mRNA) of cloned LTCCs and injections of this mRNA into Xenopus oocytes, as well as techniques of isolation and preparation of Xenopus oocytes, were described earlier in detail [15,17].…”

“…Currents through Са v 3.3 channels showed the slowest inactivation [12], and each α1 subunit possessed a definite sequence of integral conductances with respect to divalent cations (Са 2+ , Ва 2+ , and Sr 2+ ). Nevertheless, this peculiarity was determined not only by different permeabilities for these cations [15] but, perhaps, also by their different actions on the probability of the open state of these channels [16]. The latter above-mentioned peculiarity is not typical of the HTCCs, and, therefore, it can be mediated by somewhat dissimilar mechanisms of selectivity realized in the LTCCs.…”

“…The latter above-mentioned peculiarity is not typical of the HTCCs, and, therefore, it can be mediated by somewhat dissimilar mechanisms of selectivity realized in the LTCCs. This is why we continued examination of the selectivity and permeability of the cloned LTCCs [15]; special attention was paid to discrimination of mono-and divalent cations by these channels (on the example of two most physiologically important ions, Na + and Са 2+ ).…”