The contractility of vascular smooth muscle cells (VSMCs) in resistance arteries is the major contributor of vascular tone and blood pressure. In VSMCs, increase of intracellular Ca²⁺ level directly generates the force to contract arteries. A major source of intracellular Ca²⁺ is entry through voltage dependent Ca²⁺ channels (VDCCs) on plasma membrane. While mitochondria are now recognised as key regulators of intracellular Ca²⁺ homeostasis by modulating internal Ca²⁺ store in several cell types including smooth muscle cells and endothelial cells, their role in modulating Ca²⁺ signalling generated by VDCC in intact VSMCs is still under studied. Given the potential interaction between mitochondria and VDCCs in VSMCs, we hypothesize that mitochondria can directly regulate Ca²⁺ release from sarcoplasmic reticulum in VSMCs, hence regulate Ca²⁺ signalling and vascular tone generated mediated by VDCCs.

The interplay between mitochondria and VDCCs was investigated by imaging and analysing intracellular Ca²⁺ signals in smooth muscle cells in intact arteries from rat mesentery (n = 5). Summarized data were analyzed and presented as mean ± SD of n biological replicates. When the data extracted from same preparations under different treatment, data were analyzed by using paired t-test.

Depolarization of the plasma membrane potential, by high potassium (30 mM) physiological saline solution, triggered Ca²⁺ entry through VDCCs and a sustained increase in intracellular Ca2+ on which repetitive Ca²⁺ oscillations occurred. All Ca2+ signals were abolished by removal of external Ca²⁺ and by dihydropyridine inhibitors of VDCCs. Significantly, the repetitive Ca²⁺ oscillations, but not the sustained Ca²⁺ signals, were blocked by the IP₃ receptor inhibitor 2-APB and SERCA inhibitor cyclopiazonic acid. Neither the repetitive Ca²⁺ oscillations or the sustained Ca²⁺ signals were altered by the ryanodine receptor inhibitors ryanodine and dantrolene (paired t-test). These results suggest that Ca²⁺ entry via VDCC triggers Ca²⁺-induced Ca²⁺ release via IP₃ receptors in intact mesenteric arteries.

Depolarization of the mitochondrial membrane potential (ψm, indicated by a membrane sensitive dye, TMRE), by the uncoupler CCCP (1 µM), but not ATP deprivation with the ATP synthesis blocker oligomycin (1 µM), inhibited VDCCs evoked IP₃ mediated Ca²⁺ oscillations but not the sustained Ca²⁺ signals. Furthermore, in intact arteries, depolarization of ψm directly suppressed inositol triphosphate receptors (IP₃Rs) mediated Ca²⁺ release from the internal Ca²⁺ store evoked by photolysis of caged IP₃. These results suggest that mitochondria regulate Ca²⁺ release via IP₃Rs triggered by voltage dependent Ca²⁺ entry. In return, Ca²⁺ entry via VDCCs did not alter ψm. In addition, depolarization of ψm supressed sustained vasocontraction that was mediated by voltage dependent Ca²⁺ entry.

Together, these results suggest that mitochondria regulate Ca²⁺-induced Ca²⁺ release at IP₃ receptors triggered by Ca²⁺ entry via VDCCs but do not directly regulate VDCCs activity.