Four components to the outward potassium currents of rat ventricular myocytes have been identified on the basis of their voltage- and time-dependent kinetics and pharmacology (Himmel et al. 1999; James et al. 2002). We have previously shown that the steady-state background K+ current, Iss, is inhibited by the G-protein-coupled receptor agonists endothelin-1 and phenylephrine (PE; James et al. 2001a,b). The molecular basis to Iss is unclear but may involve the acid-sensitive background K+ channel, TASK-1 (James et al. 2001a). The aim of the present study was to investigate in detail the properties of the PE-sensitive current.
Male Wistar rats were humanely killed and ventricular myocytes isolated by enzymatic and mechanical dispersion. Whole-cell patch-clamp recordings of membrane currents were made at 35°C (pH 7.35) using a K+-rich pipette solution and standard external Tyrode solution containing nifedipine (3 µM) and atenolol (1 µM). Outward currents were elicited by depolarisation to +40 mV (holding potential = -70 mV) in the absence and presence of 10 µM PE. The time course of inactivation of the currents was fitted by a double exponential equation and Iss defined as the time-independent component. PE-sensitive difference currents were obtained by subtraction of Iss recorded in the absence and presence of PE. Data are expressed as means ± S.E.M. and compared using Student’s unpaired t test. P < 0.05 was accepted as significant.
Under control conditions, the PE-sensitive current was 301 ± 36 pA (n = 40). Replacement of pipette K+ with either caesium or lithium largely abolished the PE-sensitive current (12.4 ± 8.5 pA; n = 6 and -20.2 ± 4.2 pA; n = 3, respectively; P < 0.001), demonstrating the K+-selective nature of the PE-sensitive current. The PE-sensitive current was also greatly attenuated by reduction of external pH to 6.1 (38.5 ± 25.4 pA; n = 7, P < 0.001). Moreover, the PE-sensitive current was inhibited by the TASK-1 K+ channel blocker, anandamide (10 µM, 175 ± 29 pA; n = 7, P < 0.02). On the other hand, the TASK-3 K+ channel blocker, ruthenium red (20 µM), did not significantly inhibit the PE-sensitive current (237 ± 38 pA; n = 5). Neither was the PE-sensitive current (370 ± 72 pA, n = 11) affected by incorporation of the voltage-gated K+ channel blocker, tetraethylammonium (TEA, 20 mM) in the pipette solution.
Taken together, these data suggest that PE inhibits an acid-sensitive K+ channel with the properties of TASK-1 in rat ventricular myocytes.
Financial support from the British Heart Foundation (PG/98091) is gratefully acknowledged.