Outward currents in smooth muscle cells isolated from sheep mesenteric lymphatics

Abstract

1. The patch-clamp technique was used to measure membrane currents in isolated smooth muscle cells dispersed from sheep mesenteric lymphatics. Depolarizing steps positive to -30 mV evoked rapid inward currents followed by noisy outward currents. 2. Nifedipine (1 microM) markedly reduced the outward current, while Bay K 8644 (1 microM) enhanced it. Up to 90% of the outward current was also blocked by iberiotoxin (Kd = 36 nM). 3. Large conductance (304 +/- 15 pS, 7 cells), Ca(2+)- and voltage-sensitive channels were observed during single-channel recordings on inside-out patches using symmetrical 140 mM K+ solutions (at 37 degrees C). The voltage required for half-maximal activation of the channels (V1/2) shifted in the hyperpolarizing direction by 146 mV per 10-fold increase in [Ca2+]i. 4. In whole-cell experiments a voltage-dependent outward current remained when the Ca(2+)-activated current was blocked with penitrem A (100 nM). This current activated at potentials positive to -20 mV and demonstrated the phenomenon of voltage-dependent inactivation (V1/2 = -41 +/- 2 mV, slope factor = 18 +/- 2 mV, 5 cells). 6. Tetraethylammonium (TEA; 30 mM) reduced the voltage-dependent current by 75% (Kd = 3.3 mM, 5 cells) while a maximal concentration of 4-aminopyridine (4-AP; 10 mM) blocked only 40% of the current. TEA alone had as much effect as TEA and 4-AP together, suggesting that there are at least two components to the voltage-sensitive K+ current. 7. These results suggest that lymphatic smooth muscle cells generate a Ca(2+)-activated current, largely mediated by large conductance Ca(2+)-activated K+ channels, and several components of voltage-dependent outward current which resemble 'delayed rectifier' currents in other smooth muscle preparations.