Expression of Calcium Channel Subunit Variants in Small Mesenteric Arteries of WKY and SHR.

ABSTRACT: Enhanced function of dihydropyridine-sensitive Ca2+ channels (CaV) in hypertensive arterial myocytes (HAM) is well accepted. Increased protein expression of pore forming ?1-subunits contributes to this effect, but cannot explain all of the differences in CaV properties in HAM. We hypothesized that differences in expression of CaV subunits and/or their splice variants also contribute.RNA, protein, and myocytes were isolated from small mesenteric arteries (SMA) of 20-week-old male WKY and SHR and analyzed by polymerase chain reaction (PCR), sequencing, immunoblotting, and patch clamp methods.Cav1.2 ?1, ?2c, and ?2?1d were the dominant subunits expressed in both WKY and SHR with a smaller amount of ?3a. Real-time PCR indicated that the mRNA abundance of ?3a and ?2?1 but not total Cav1.2 ?1 or ?2c were significantly larger in SHR. Analysis of alternative splicing of Cav1.2 ?1 showed no differences in abundance of mutually exclusive exons1b, 8, 21 and 32 or alternative exons33 and 45. However, inclusion of exon9* was higher and a 73 nucleotide (nt) deletion in exon15 (exon15?73) was lower in SHR. Immunoblot analysis showed higher protein levels of Cav1.2 ?1 (1.61±0.05), ?3 (1.80±0.32), and ?2?1 (1.80±0.24) but not ?2 in SHR.The lower abundance of exon15?73 transcripts in SHR results in a larger fraction of total Cav1.2 mRNA coding for full-length CaV protein, and the higher abundance of exon9* transcripts and CaV?3a protein likely contribute to differences in gating and kinetics of CaV currents in SHR. Functional studies of Ca2+ currents in native SMA myocytes and HEK cells transiently transfected with CaV subunits support these conclusions.

SUBMITTER: Cox RH 

PROVIDER: S-EPMC4675841 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature