beta-Adrenoceptor stimulation activates large-conductance Ca2+-activated K+ channels in smooth muscle cells from basilar artery of guinea pig.
We studied the effect of isoproterenol on the Ca2+-activated K+ (BK) channel in smooth muscle cells isolated from the basilar artery of the guinea pig. Cells were studied in a whole-cell configuration to allow the clamping of intracellular Ca2+ concentration, [Ca2+]i. Macroscopic BK channel currents were recorded during depolarizing test pulses from a holding potential (VH) of 0 mV, which was used to inactivate the outward rectifier. The outward macroscopic current available from a VH of 0 mV was highly sensitive to block by external tetraethylammonium Cl (TEA) and charybdotoxin, and was greatly augmented by increasing [Ca2+]i from 0.01 to 1.0 microM. With [Ca2+]i between 0.1 and 1.0 microM, 0.4 microM isoproterenol increased this current by 58.6 +/- 17.1%, whereas with [Ca2+]i at 0.01 microM a sixfold smaller increase was observed. With [Ca2+]i > or = 0.1 microM, 100 microM dibutyryl -adenosine 3':5: cyclic monophosphate (cAMP) and 1 microM forskolin increased this current by 58.5 +/- 24.1% and 59.7 +/- 10.3%, respectively. The increase with isoproterenol was blocked by 4.0 microM propranolol extracellularly, and by 10 U/ml protein kinase inhibitor intracellularly. Single-channel openings during depolarizing test pulses from a VH of 0 mV recorded in the whole-cell configuration under the same conditions (outside-out-whole-cell recording) indicated a slope conductance of 260 pS. In conventional outside-out patches, this 260-pS channel was highly sensitive to block by external TEA, and in inside-out patches, its probability of opening was greatly augmented by increasing [Ca2+]i from 0.01 to 1.0 microM. Outside-out-whole-cell recordings with [Ca2+]i > or = 0.1 microM indicated that 100 microM dibutyryl-cAMP increased the probability of opening of the 260-pS channel by 152 +/- 115%. In inside-out patches, the catalytic subunit of protein kinase A increased the probability of opening, and this effect also depended on [Ca2+]i , with a 35-fold larger effect observed with 0.1-0.5 microM Ca2+ compared to 0.01 microM Ca2+. We conclude that the BK channel in cerebrovascular smooth muscle cells can be activated by beta-adrenoceptor stimulation, that the effect depends strongly on [Ca2+]i, and that the effect is mediated by cAMP-dependent protein kinase A with no important contribution from a direct G-protein or phosphorylation-independent mechanism. Our data indicate that the BK channel may participate in beta-adrenoceptor-mediated relaxation of cerebral vessels, although the importance of this pathway in obtaining vasorelaxation remains to be determined.
Song
Y
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Neuroscience Program, University of Texas Medical Branch, Galveston, TX 77550, USA.
Simard
J M
JM
D000818:Animals; D001488:Basilar Artery / Q000166:cytology / Q000502:physiology*; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D000242:Cyclic AMP / Q000494:pharmacology / Q000502:physiology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000494:pharmacology / Q000502:physiology; D004553:Electric Conductivity; D006168:Guinea Pigs; D007545:Isoproterenol / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000502:physiology*; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology*; D011943:Receptors, Adrenergic, beta / Q000502:physiology*
D015221:Potassium Channels; D011943:Receptors, Adrenergic, beta; D003994:Bucladesine; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D007545:Isoproterenol; D002118:Calcium