RETRACTED: [CA2+](I) INHIBITION OF K+ CHANNELS IN CANINE RENAL-ARTERY - NOVEL MECHANISM FOR AGONIST-INDUCED MEMBRANE DEPOLARIZATION (Retracted Article. See vol. 94, pg E15, 2004)

The patch-clamp technique was used to examine the inhibition of delayed rectifier K+ channels by agents that release intracellular Ca2+. During voltage-clamp experiments on isolated myocytes with 4-aminopyridine (4-AP, 10 mmol/L) and niflumic acid (100 mu mol/L) present to inhibit delayed rectifier K+ current (I-K(dr)) and Ca2+-activated Cl- current (I-Cl(Ca)), angiotensin II (Ang II) and caffeine increased Ca2+- activated K+ current (I-K(Ca)) between -25 and 80 mV (n=5). Conversely, with charybdotoxin (ChTX, 100 nmol/L) and niflumic acid (100 mu mol/L) present to inhibit I-K(Ca) and I-Cl(Ca), Ang II and caffeine only caused inhibition of I-K(dr). Block was achieved within 15 seconds of drug application and was reversible upon washout (n=5). The effects of Ang II on I-K(Ca) and I-K(dr) were inhibited by the specific Ang II receptor antagonist losartan (1 mmol/L, n=3). Intracellular BAPTA (10 mmol/L) also abolished the effects of Ang II and caffeine on both I-K(Ca) and I-K(dr). In current-clamp experiments, the application of ChTX (100 nmol/L) and niflumic acid (100 mu mol/L) caused little change in resting membrane potential; however, subsequent application of caffeine (10 mmol/L) caused a 26+/-2.9 mV depolarization from -54+/-3.1 to -28+/-1.7 mV (n=6). 4-AP (10 mmol/L) blocked the caffeine-induced depolarization. When isolated cells were loaded with the Ca2+ indicator indo 1 (100 mu mol/L), Ang II, caffeine, and 4-AP increased [Ca2+](i) and depolarized the cells. Both Ang II and caffeine caused an increase in [Ca2+](i) that preceded membrane depolarization, whereas 4-AP depolarized the cell first and then caused an increase in [Ca2+](i) (n=4). In inside-out patches, with 200 nmol/L ChTX in the patch pipette to block large-conductance Ca2+-activated K+ channels, a 45+/-7-picosiemen 4-AP-sensitive K+ channel was identified that was sensitive to cytoplasmic Ca2+ (n=6). Increasing intracellular Ca2+ decreased channel opening probability [NxP(open), where N is the number of functional channels in a patch and P(open) is the opening probability] at all membrane potentials examined. At 0 mV, increasing Ca2+ from <5 to 200 and 600 nmol/L free Ca2+ decreased NxP(open) by 52+/-3% and 73+/-7%, respectively (n=6). The decrease in opening probability of the delayed rectifier K+ channel resulted from a concentration- and voltage-dependent decrease in mean open time. The decrease in mean open time reflected significant decreases and increases in open and closed time constants, respectively. These results suggest that agonist-induced changes in intracellular Ca2+ can alter vascular smooth muscle membrane potential through regulation of delayed rectifier K+ channels.