KMUP-1 activates BKCa channels in basilar artery myocytes via cyclic nucleotide-dependent protein kinases

This study investigated whether KMUP-1, a synthetic xanthine-based derivative, augments the delayed-rectifier potassium (K(DR))- or large-conductance Ca2+-activated potassium (BKCa) channel activity in rat basilar arteries through protein kinase-dependent and -independent mechanisms. Cerebral smooth muscle cells were enzymatically dissociated from rat basilar arteries. Conventional whole cell, perforated and inside-out patch-clamp electrophysiology was used to monitor K+- and Ca2+ channel activities. KMUP-1 (1 microM) had no effect on the K(DR) current but dramatically enhanced BKCa channel activity. This increased BKCa current activity was abolished by charybdotoxin (100 nM) and iberiotoxin (100 nM). Like KMUP-1, the membrane-permeable analogs of cGMP (8-Br-cGMP) and cAMP (8-Br-cAMP) enhanced the BKCa current. BKCa current activation by KMUP-1 was markedly inhibited by a soluble guanylate cyclase inhibitor (ODQ 10 microM), an adenylate cyclase inhibitor (SQ 22536 10 microM), competitive antagonists of cGMP and cAMP (Rp-cGMP, 100 microM and Rp-cAMP, 100 microM), and cGMP- and cAMP-dependent protein kinase inhibitors (KT5823, 300 nM and KT5720, 300 nM). Voltage-dependent L-type Ca2+ current was significantly suppressed by KMUP-1 (1 microM), and nearly abolished by a calcium channel blocker (nifedipine, 1 microM). In conclusion, KMUP-1 stimulates BKCa currents by enhancing the activity of cGMP-dependent protein kinase, and in part this is due to increasing cAMP-dependent protein kinase. Physiologically, this activation would result in the closure of voltage-dependent calcium channels and the relaxation of cerebral arteries.