Role of endothelial cell hyperpolarization in EDHF‐mediated responses in the guinea‐pig carotid artery

Experiments were performed to identify the potassium channels involved in the acetylcholine‐induced endothelium‐dependent hyperpolarization of the guinea‐pig internal carotid artery. Smooth muscle and endothelial cell membrane potentials were recorded in isolated arteries with intracellular microelectrodes. Potassium currents were recorded in freshly‐dissociated smooth muscle cells using patch clamp techniques. In single myocytes, iberiotoxin (0.1 μM)‐, charybdotoxin (0.1 μM)‐, apamin (0.5 μM)‐ and 4‐aminopyridine (5 mM)‐sensitive potassium currents were identified indicating the presence of large‐ and small‐conductance calcium‐sensitive potassium channels (BKCa and SKCa) as well as voltage‐dependent potassium channels (KV). Charybdotoxin and iberiotoxin inhibited the same population of BKCa but a conductance specifically sensitive to the combination of charybdotoxin plus apamin could not be detected. 4‐aminopyridine (0.1–25 mM) induced a concentration‐dependent inhibition of KV without affecting the iberiotoxin‐ or the apamin‐sensitive currents. In isolated arteries, both the endothelium‐dependent hyperpolarization of smooth muscle and the hyperpolarization of endothelial cells induced by acetylcholine or by substance P were inhibited by 5 mM 4‐aminopyridine. These results indicate that in the vascular smooth muscle cells of the guinea‐pig carotid artery, a conductance specifically sensitive to the combination of charybdotoxin plus apamin could not be detected, comforting the hypothesis that the combination of these two toxins should act on the endothelial cells. Furthermore, the inhibition by 4‐aminopyridine of both smooth muscle and endothelial hyperpolarizations, suggests that in order to observe an endothelium‐dependent hyperpolarization of the vascular smooth muscle cells, the activation of endothelial potassium channels is likely to be required. British Journal of Pharmacology (2000) 129, 1103–1112; doi:10.1038/sj.bjp.0703175