Bannister John P, Adebiyi Adebowale, Zhao Guiling, Narayanan Damodaran, Thomas Candice M, Feng Jessie Y, Jaggar Jonathan H
Department of Physiology, University of Tennessee Health Science Center, Memphis, TN 38163, USA.
Circ Res. 2009 Nov 6;105(10):948-55. doi: 10.1161/CIRCRESAHA.109.203620. Epub 2009 Oct 1.
Voltage-dependent L-type (Ca(V)1.2) Ca(2+) channels are a heteromeric complex formed from pore-forming alpha(1) and auxiliary alpha(2)delta and beta subunits. Ca(V)1.2 channels are the principal Ca(2+) influx pathway in arterial myocytes and regulate multiple physiological functions, including contraction. The macromolecular composition of arterial myocyte Ca(V)1.2 channels remains poorly understood, with no studies having examined the molecular identity or physiological functions of alpha(2)delta subunits.
We investigated the functional significance of alpha(2)delta subunits in myocytes of resistance-size (100 to 200 mum diameter) cerebral arteries.
alpha(2)delta-1 was the only alpha(2)delta isoform expressed in cerebral artery myocytes. Pregabalin, an alpha(2)delta-1/-2 ligand, and an alpha(2)delta-1 antibody, inhibited Ca(V)1.2 currents in isolated myocytes. Acute pregabalin application reversibly dilated pressurized arteries. Using a novel application of surface biotinylation, data indicated that >95% of Ca(V)1.2 alpha(1) and alpha(2)delta-1 subunits were present in the arterial myocyte plasma membrane. Alpha(2)delta-1 knockdown using short hairpin RNA reduced plasma membrane-localized Ca(V)1.2 alpha(1) subunits, caused a corresponding elevation in cytosolic Ca(V)1.2 alpha(1) subunits, decreased intracellular Ca(2+) concentration, inhibited pressure-induced vasoconstriction ("myogenic tone"), and attenuated pregabalin-induced vasodilation. Prolonged (24-hour) pregabalin exposure did not alter total alpha(2)delta-1 or Ca(V)1.2 alpha(1) proteins but decreased plasma membrane expression of each subunit, which reduced myogenic tone.
alpha(2)delta-1 is essential for plasma membrane expression of arterial myocyte Ca(V)1.2 alpha(1) subunits. alpha(2)delta-1 targeting can block Ca(V)1.2 channels directly and inhibit surface expression of Ca(V)1.2 alpha(1) subunits, leading to vasodilation. These data identify alpha(2)delta-1 as a novel molecular target in arterial myocytes, the manipulation of which regulates contractility.
电压依赖性L型（Ca(V)1.2）钙通道是由形成孔道的α1亚基以及辅助性α2δ和β亚基组成的异源复合物。Ca(V)1.2通道是动脉肌细胞中主要的钙内流途径，并调节多种生理功能，包括收缩。动脉肌细胞Ca(V)1.2通道的大分子组成仍知之甚少，尚无研究检测α2δ亚基的分子特性或生理功能。
我们研究了α2δ亚基在阻力型（直径100至200μm）脑动脉肌细胞中的功能意义。
α2δ-1是脑动脉肌细胞中唯一表达的α2δ亚型。普瑞巴林，一种α2δ-1/-2配体，以及一种α2δ-1抗体，可抑制分离肌细胞中的Ca(V)1.2电流。急性应用普瑞巴林可使加压动脉可逆性扩张。使用表面生物素化的新方法，数据表明超过95%的Ca(V)1.2α1和α2δ-1亚基存在于动脉肌细胞质膜中。使用短发夹RNA敲低α2δ-1可减少质膜定位的Ca(V)1.2α1亚基，导致胞质Ca(V)1.2α1亚基相应升高，降低细胞内钙浓度，抑制压力诱导的血管收缩（“肌源性张力”），并减弱普瑞巴林诱导的血管舒张。长时间（24小时）暴露于普瑞巴林不会改变总α2δ-1或Ca(V)1.2α1蛋白，但会降低每个亚基的质膜表达，从而降低肌源性张力。
α2δ-1对于动脉肌细胞Ca(V)1.2α1亚基的质膜表达至关重要。靶向α2δ-1可直接阻断Ca(V)1.2通道并抑制Ca(V)1.2α1亚基的表面表达，从而导致血管舒张。这些数据确定α2δ-1是动脉肌细胞中的一个新分子靶点，对其进行调控可调节收缩性。
