Cellular target of voltage and calcium-dependent K+ channel blockers involved in EDHF-mediated responses in rat superior mesenteric artery

Abstract

1. We have investigated the cellular target of K+ channel blockers responsible for the inhibition of the EDHF-mediated relaxation in the rat mesenteric artery by studying their effects on tension, smooth muscle cell (SMC) membrane potential and endothelial cell Ca2+ signal ([Ca2+]endo). 2. In arteries contracted with prostaglandin F2α (2.5-10 μM), relaxation evoked by ACh (0.01-3 μM) was abolished by a combination of charybdotoxin (ChTX, 0.1 μM) plus apamin (Apa, 0.1 μM) and was inhibited by 68 ± 6% (n=6) by 4-aminopyridine (4-AP, 5 mM). 3. ACh (0.001-3 μM) increased [Ca2+]endo and hyperpolarized SMCs with the same potency, the pD2 values were equal to 7.2±0.08 (n=4) and 7.2±0.07 (n=9), respectively. SMCs hyperpolarization to ACh (1 μM) was abolished by high K+ solution or by ChTX/Apa. It was decreased by 66±5% (n=6) by 4-AP. 4. The increase in [Ca2+]endo evoked by ACh (1 μM) was insensitive to ChTX/Apa but was depressed by 58±16% (n=6) and 27±4% (n=7) by raising external K+ concentration and by 4-AP, respectively. 5. The effect of 4-AP on [Ca2+]endo was not affected by increasing external K+ concentration. In Ca-free/EGTA solution, the transient increase in [Ca2+]endo evoked by ACh (1 μM) was abolished by thapsigargin (1 μM) and was decreased by 75±7% (n=5) by 4-AP. 6. These results show that inhibition of EDHF-evoked responses by 4-AP may be attributed to a decrease in the Ca2+ release activated by ACh in endothelial cells. The abolition of SMCs hyperpolarization to ACh by ChTX/Apa is not related to an interaction with the [Ca2+]endo.