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(Circulation Research. 2009;105:948.)
© 2009 American Heart Association, Inc.

Cellular Biology

Smooth Muscle Cell ₂-1 Subunits Are Essential for Vasoregulation by Ca_V1.2 Channels

John P. Bannister, Adebowale Adebiyi, Guiling Zhao, Damodaran Narayanan, Candice M. Thomas, Jessie Y. Feng, Jonathan H. Jaggar

From the Department of Physiology, University of Tennessee Health Science Center, Memphis.

Correspondence to Jonathan H. Jaggar, University of Tennessee Health Science Center, Department of Physiology, 894 Union Ave, Nash Building, Memphis, TN 38163. E-mail jjaggar{at}physio1.utmem.edu

Rationale: Voltage-dependent L-type (Ca_V1.2) Ca²⁺ channels are a heteromeric complex formed from pore-forming ₁ and auxiliary ₂ and β subunits. Ca_V1.2 channels are the principal Ca²⁺ influx pathway in arterial myocytes and regulate multiple physiological functions, including contraction. The macromolecular composition of arterial myocyte Ca_V1.2 channels remains poorly understood, with no studies having examined the molecular identity or physiological functions of ₂ subunits.

Objective: We investigated the functional significance of ₂ subunits in myocytes of resistance-size (100 to 200 µm diameter) cerebral arteries.

Methods and Results: ₂-1 was the only ₂ isoform expressed in cerebral artery myocytes. Pregabalin, an ₂-1/-2 ligand, and an ₂-1 antibody, inhibited Ca_V1.2 currents in isolated myocytes. Acute pregabalin application reversibly dilated pressurized arteries. Using a novel application of surface biotinylation, data indicated that >95% of Ca_V1.2 ₁ and ₂-1 subunits were present in the arterial myocyte plasma membrane. ₂-1 knockdown using short hairpin RNA reduced plasma membrane-localized Ca_V1.2 ₁ subunits, caused a corresponding elevation in cytosolic Ca_V1.2 ₁ subunits, decreased intracellular Ca²⁺ concentration, inhibited pressure-induced vasoconstriction ("myogenic tone"), and attenuated pregabalin-induced vasodilation. Prolonged (24-hour) pregabalin exposure did not alter total ₂-1 or Ca_V1.2 ₁ proteins but decreased plasma membrane expression of each subunit, which reduced myogenic tone.

Conclusions: ₂-1 is essential for plasma membrane expression of arterial myocyte Ca_V1.2 ₁ subunits. ₂-1 targeting can block Ca_V1.2 channels directly and inhibit surface expression of Ca_V1.2 ₁ subunits, leading to vasodilation. These data identify ₂-1 as a novel molecular target in arterial myocytes, the manipulation of which regulates contractility.

Key Words: L-type Ca²⁺ channels • arterial contractility • auxiliary subunit