Azelnidipine treatment reduces the expression of Ca(v)1.2 protein

Aims: Azelnidipine, a third-generation dihydropyridine calcium channel blocker (DHP CCB), has a characteristic hypotensive effect that persists even after it has disappeared from the plasma, which is thought to be due to its high hydrophobicity. However, because azelnidipine is unique, it might have other unknown effects on L-type Ca(v)1.2 channels that result in the long-lasting decrease of blood pressure. The aim of this study was to investigate the potential quantitative modification of Ca(v)1.2 by azelnidipine. Main methods: HEK293 cells were used to express Ca(v)1.2 channels. Immunocytochemical analysis was performed to detect changes in the surface expression of the pore-forming subunit of the Ca(v)1.2 channel, Ca(v)1.2 alpha(1c). Western blotting analysis was performed to evaluate changes in expression levels of total Ca(v)1.2 alpha(1c) and Ca-v beta(2c). Key findings: The surface expression of Ca(v)1.2 alpha(1c) was markedly reduced by treatment with azelnidipine, but not with other DHP CCBs (amlodipine and nicardipine). Results obtained with a dynamin inhibitor and an early endosome marker suggested that the reduction of surface Ca(v)1.2 alpha(1c) was not likely caused by internalization. Azelnidipine reduced the total amount of Ca(v)1.2 alpha(1c) protein in HEK293 cells and rat pulmonary artery smooth muscle cells. The reduction of Ca(v)1.2 alpha(1c) was rescued by inhibiting proteasome activity. In contrast, azelnidipine did not affect the amount of auxiliary Ca-v beta(2c) subunits that function as a chaperone of Ca(v)1.2. Significance: This study is the first to demonstrate that azelnidipine reduces the expression of Ca(v)1.2 alpha(1c), which might partly explain its long-lasting hypotensive effect.