Potassium channels play a key role in the regulation of excitation-contraction coupling in vascular SMCs and Kv channels, being ubiquitous in the vasculature, are important contributors to the regulation of vascular tone. Intracellular mechanisms controlling function of Kv channels in arteries are poorly understood. We have previously found that the replacement of 5 mM MgATP with 5 mM MgCl₂ in the pipette solution caused a significant negative shift in the Kv channel current (I_Kv) availability in RASMCs (Tammaro et al. 2001). Gelband et al. (1993) showed previously that increase in intracellular divalent cation concentration significantly suppressed I_Kv in isolated SMCs. Since ATP is a potent chelator of Mg²⁺, a possibility of the direct effect of intracellular Mg²⁺ on I_Kv was further investigated in aortic SMCs isolated from humanely killed male Wistar rats.

Kv channel currents were isolated using paxilline (1 µM) and glibenclamide (10 µM) in the external solution to block Ca²⁺-activated and ATP-sensitive K⁺ currents, respectively. Pipette solution contained (mM): 110 KCl, 10 NaCl, 10 Hepes, 10 EGTA and 0.5 CaCl₂; pH 7.2. Intracellular [Mg²⁺] was altered by adding MgCl₂. Increasing the pipette concentration of MgCl₂ from 0.5 mM (used as control) to 5 mM did not significantly affect either I_Kv steady-state activation (calculated from I-V relationships) or the whole-cell maximum conductance (0.077 ± 0.009 pS pF^-1, n = 12 and 0.074 ± 0.007 pS pF^-1, n = 15; mean ± S.E.M.). However, I_Kv availability, measured with 10 s conditioning pre-pulses, was significantly shifted by ~9 mV (P < 0.013, unpaired t test) with V_0.5 = -45.4 ± 1.7 mV (n = 6) and V_0.5 = -53.7 ± 1.9 mV (n = 11) for 0.5 and 5 mM MgCl₂, respectively. Increase in the pipette concentration of MgCl₂ to 10 mM produced further shifts in I_Kv availability (V_0.5 = 61.7 ± 5.7 mV, n = 6). Although steady-state activation was not affected (P > 0.05), whole-cell maximum conductance was decreased by 47 % to 0.04 ± 0.004 (n = 6, P < 0.002) by 10 mM MgCl₂. Perfusion of RASMCs with pipette solution containing 0.5 mM MgCl₂ with pH adjusted to 6.2 did not significantly affect either voltage dependency I_Kv activation and inactivation or the maximum conductance.

A selective effect of intracellular Mg²⁺ on I_Kv inactivation and not activation dependencies and the lack of the effect of increased intracellular concentration of protons on I_Kv does not support direct interaction between Mg²⁺ and fixed surface charges near the internal channel mouth, and could indicate a novel mechanism of the modulation of Kv channels by physiological concentrations of intracellular Mg²⁺ in VSMCs.

This work was supported by the BHF.