BLOCK OF LARGE-CONDUCTANCE CA2-ACTIVATED K+ CHANNELS IN RABBIT VASCULAR MYOCYTES BY INTERNAL MG2+ AND NA+()

1. We studied the biophysical properties of single large conductance ( >200 pS in symmetrical K+ pipette and bath solutions) Ca2+-activated K + (BKCa) channels of rabbit portal vein and coronary arterial smooth m uscle cells using the cell-attached and inside-out variants of the pat ch-clamp technique (at 22 degrees C). 2. The unitary conductance of BK Ca channels recorded in cell-attached patches with K+ concentrations i n the range 5.4-140 mM was significantly lower than that predicted on the basis of the conductance measured in inside-out patches with symme trical K+ pipette and bath solutions (140 mM) and the constant field e quation. In cell-attached patches from cells bathed in depolarizing me dium (140 mM) with 5.4 mM K+ in the pipette solution, BKCa channels we re difficult to detect in the physiological range of membrane potentia ls (approximately -50 mV). Unitary currents were smaller at all voltag es in the range -50 to 0 mV and the i-V relationship exhibited strong inward rectification at potentials >0 mV. These channels were unequivo cally identified as BKCa channels due to their sensitivity to caffeine (10 mM) and iberiotoxin (20 nM), and their non-stationary kinetic pro perties. 3. Exposure of the cytoplasmic side of excised patches to [Mg 2+] in the range 0-15 mM produced two effects on BKCa channel activity : the slope conductance and open probability were reduced and enhanced , respectively in a concentration-dependent manner by this cation. The Mg2+-induced reduction in conductance exhibited weak voltage dependen ce. 4. Application of 20 mM Na+ to the internal face of BKCa channels recorded in the inside-out configuration produced a flickery block at potentials greater than or equal to+20 mV resulting in reduced unitary current amplitudes and strong inward rectification of the i-V relatio nship. Exposure of inside-out patches to a combination of 20 mM Na+ an d 2 mM Mg2+ further reduced unitary current amplitude to a level simil ar to the algebraic sum of the effect of each cation in isolation. 5. We conclude that Ca2+-dependent K+ channels of vascular smooth muscle cells display a lower unitary conductance when recorded under physiolo gical conditions than that previously estimated on the basis of their behaviour in excised membrane patches. Our data indicate that the decr eased permeation through BKCa channels may be partly attributed to blo ck by intracellular Mg2+ and Na+, which appear to interact with distin ct binding sites along the inner side of the pore.