Allosteric regulation of BK channel gating by Ca2+ and Mg2+ through a nonselective, low affinity divalent cation site

The ability of membrane voltage to activate high conductance, calcium-activ ated (BK-type) K+ channels is enhanced by cytosolic calcium (Ca2+). Activat ion is sensitive to a range of [Ca2+] that spans over four orders of magnit ude. Here, we examine the activation of BK channels resulting from expressi on of cloned mouse Slog a subunits at [Ca2+] and [Mg2+] up to 100 mM. The H alf-activation voltage (V-0.5) is steeply dependent on [Ca2+] in the microm olar range, but shows a tendency towards saturation over the range of 60-30 0 muM Ca2+. As [Ca2+] is increased to millimolar levels, the V-0.5 is stron gly shifted again to more negative potentials. When channels are activated by 300 muM Ca2+, further addition of either mM Ca2+ or mM Mg2+ produces sim ilar negative shifts in steady-state activation. Millimolar Mg2+ also produ ces shifts of similar magnitude in the complete absence of Ca2+. The abilit y of millimolar concentrations of divalent cations to shift activation is p rimarily correlated with a slowing of BK current deactivation. At voltages where millimolar elevations in [Ca2+] increase activation rates, addition o f 10 mM Mg2+ to 0 Ca2+ produces little effect on activation tinge course, w hile markedly slowing deactivation. This suggests that Mg2+ does not partic ipate in Ca2+-dependent steps that influence current activation rate. We co nclude that millimolar Mg2+ and Ca2+ concentrations interact with low affin ity, relatively nonselective divalent cation binding sites that are distinc t from higher affinity, Ca2+-selective binding sites that increase current activation rates. A symmetrical model with four independent higher affinity Ca2+ binding steps, four voltage sensors, and four independent lower affin ity Ca2+/Mg2+ binding steps describes well the behavior of G-V curves over a range of Ca2+ and Mg2+. The ability of a broad range of [Ca2+] to produce shifts in activation of Slo1 conductance can, therefore, be accounted for by multiple types of divalent cation binding sites.