Influence of Mitochondrial Inhibition on Global and Local [Ca ²⁺ ] _i in Rat Tail Artery

Abstract

Abstract-Inhibition of oxidative metabolism is often found to decrease contractility of systemic vascular smooth muscle, but not to reduce global [Ca 2ϩ ] i . In the present study, we probe the hypothesis that it is associated with an altered pattern of intracellular Ca 2ϩ oscillations (waves) influencing force development. In the rat tail artery, mitochondrial inhibitors (rotenone, antimycin A, and cyanide) reduced ␣ 1 -adrenoceptor-stimulated force by 50% to 80%, but did not reduce global [Ca 2ϩ ] i . Less relaxation (about 30%) was observed after inhibition of myosin phosphatase activity with calyculin A, suggesting that part of the metabolic sensitivity involves the regulation of myosin 20-kDa light chain phosphorylation, although no decrease in phosphorylation was found in freeze-clamped tissue. Confocal imaging revealed that the mitochondrial inhibitors increased the frequency but reduced the amplitude of asynchronous cellular Ca 2ϩ waves elicited by ␣ 1 stimulation. The altered wave pattern, in association with increased basal [Ca 2ϩ ] i , accounted for the unchanged global [Ca 2ϩ ] i . Inhibition of glycolytic ATP production by arsenate caused similar effects on Ca 2ϩ waves and global [Ca 2ϩ ] i , developing gradually in parallel with decreased contractility. Inhibition of wave activity by the InsP 3 receptor antagonist 2-APB correlated closely with relaxation. Furthermore, abolition of waves with thapsigargin in the presence of verapamil reduced force by about 50%, despite unaltered global [Ca 2ϩ ] i , suggesting that contraction may at least partly depend on Ca 2ϩ wave activity. This study therefore indicates that mitochondrial inhibition influences Ca 2ϩ wave activity, possibly due to a close spatial relationship of mitochondria and the sarcoplasmic reticulum and that this contributes to metabolic vascular relaxation. Key Words: arterial smooth muscle Ⅲ metabolic inhibition Ⅲ myosin phosphorylation Ⅲ calcium waves Ⅲ confocal imaging T he distribution of blood flow in tissue depends on the sensitivity of arterial tone to the local oxygen tension, but the nature of the mediating signal is yet unclear. Proposed mechanisms include reduced Ca 2ϩ inflow across the cell membrane, 1-4 decreased myosin phosphorylation, 5 or inhibition of cross-bridge cycling. 6 In some cases of vascular hypoxia or mitochondrial inhibition, the intracellular free Ca 2ϩ concentration ([Ca 2ϩ ] i ), measured as an average from multiple cells, is unchanged or even increased at high stimulus levels while force is reduced. [7][8][9][10] Thus mechanisms based on decreased global [Ca 2ϩ ] i are insufficient to explain hypoxic relaxation. It is puzzling also that in many studies myosin light chain phosphorylation does not decrease, 6,9 even though exceptions are seen. 5 Finally, insufficient MgATP for cross-bridge interaction, although clearly a possibility with severe metabolic inhibition, does not seem to be a general explanation for hypoxic relaxation. 10 Within the vessel wall, individual cells may exhibit asynchron...

Vasomotor responses in MnSOD-deficient mice

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Mitochondrial regulation of cytosolic Ca2+ signals in smooth muscle

“…Mitochondria were shown to modulate control IP 3 -evoked Ca 2+ release by operating within an IP 3 R cluster, and when mitochondrial Ca 2+ uptake was prevented, a localised Ca 2+ release from an IP 3 R (a Ca 2+ puff) was inhibited (Fig. 3) [17,22,57,76,78]. The results were consistent with Ca 2+ uptake by mitochondria limiting a negative feedback effect of Ca 2+ on the IP 3 R cluster.…”

“…Several early studies have shown that Ca 2+ increases evoked by IP 3 -generating agonists were almost fully inhibited when mitochondrial Ca 2+ uptake was prevented [17,22,57,76,78], i.e. mitochondria appeared to determine whether or not a Ca 2+ signal occurred at all.…”

Mitochondrial involvement in carbachol-induced intracellular Ca2+ mobilization and contraction in rat gastric smooth muscle

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