Alternative splicing of [Ca.sub.v]1.2 channel exons in smooth muscle cells of resistance-size arteries generates currents with unique electrophysiological properties

Voltage-dependent calcium ([Ca.sup.2+], [Ca.sub.v]1.2) channels are the primary [Ca.sup.2+] entry pathway in smooth muscle cells of resistance-size (myogenic) arteries, but their molecular identity remains unclear. Here we identified and quantified [Ca.sub.v]1.2 [[alpha].sub.1]-subunit splice variation in myocytes of rat resistance-size (100-200 [micro]m diameter) cerebral arteries. Full-length clones containing either exon 1b or the recently identified exon 1c exhibited additional primary splice variation at exons 9*, 21/22, 31/32, and [+ or -] 33. Real-time PCR confirmed the findings from full-length clones and indicated that the major [Ca.sub.v]1.2 variant contained exons 1c, 8, 21, and 32+33, with ~57% containing 9*. Exon 9* was more prevalent in clones containing 1c (72%) than in those containing 1b (33%), suggesting exon-selective combinatorial splicing. To examine the functional significance of this splicing profile, membrane currents produced by each of the four exon 1b/c/ [+ or -] 9* variants were characterized following transfection in HEK293 cells. Exon 1c and 9* caused similar hyperpolarizing shifts in both current-voltage relationships and voltage-dependent activation of currents. Furthermore, exon 9* induced a hyperpolarizing shift only in the voltage-dependent activation of channels containing exon 1b, but not in those containing exon 1c. In contrast, exon 1b, 1c, or +9* did not alter voltage-dependent inactivation. In summary, we have identified the [Ca.sub.v]1.2 [[alpha].sub.1]-subunit splice variant population that is expressed in myocytes of resistance-size arteries and the unique electrophysiological properties of recombinant channels formed by exon 1 and 9* variation. The predominance of exon 1c and 9* in smooth muscle cell [Ca.sub.v]1.2 channels causes a hyperpolarizing shift in the voltage sensitivity of currents toward the physiological arterial voltage range. voltage-dependent calcium channel; myogenic artery; cloning; ribonucleic acid splicing