Divergent roles of glycolysis and the mitochondrial electron transport chain in hypoxic pulmonary vasoconstriction of the rat: identity of the hypoxic sensor

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Divergent roles of glycolysis and the mitochondrial electron transport chain in hypoxic pulmonary vasoconstriction of the rat: identity of the hypoxic sensor

Richard M. Leach, Heidi M. Hill, Vladimir A. Snetkov, Thomas P. Robertson * and Jeremy P. T. Ward

Department of Respiratory Medicine and Allergy, GKT School of Medicine, Centre for Cardiovascular Biology and Medicine, King's College London, Guy's Hospital Campus, London SE1 9RT, UK and * Department of Physiology and Pharmacology, Veterinary School, University of Georgia, Athens, GA, USA

The mechanisms responsible for sensing hypoxia and initiating hypoxic pulmonary vasoconstriction (HPV) are unclear. We therefore examined the roles of the mitochondrial electron transport chain (ETC) and glycolysis in HPV of rat small intrapulmonary arteries (IPAs).
HPV demonstrated a transient constriction (phase 1) superimposed on a sustained constriction (phase 2). Inhibition of complex I of the ETC with rotenone (100 nM) or complex III with myxothiazol (100 nM) did not cause vasoconstriction in normoxia, but abolished both phases of HPV. Rotenone inhibited the hypoxia-induced rise in intracellular Ca²⁺ ([Ca²⁺]_i). Succinate (5 mM), a substrate for complex II, reversed the effects of rotenone but not myxothiazol on HPV, but did not affect the rise in NAD(P)H fluorescence induced by hypoxia or rotenone. Inhibition of cytochrome oxidase with cyanide (100 µM) potentiated phase 2 constriction.
Phase 2 of HPV, but not phase 1, was highly correlated with glucose concentration, being potentiated by 15 mM but abolished in its absence, or following inhibition of glycolysis by iodoacetate or 2-deoxyglucose. Glucose concentration did not affect the rise in [Ca²⁺]_i during HPV.
Depolarisation-induced constriction was unaffected by hypoxia except in the absence of glucose, when it was depressed by ~50 %. Depolarisation-induced constriction was depressed by rotenone during hypoxia by 23 ± 4 %; cyanide was without effect.
Hypoxia increased 2-deoxy-[³H]glucose uptake in endothelium-denuded IPAs by 235 ± 32 %, and in mesenteric arteries by 218 ± 38 %.
We conclude that complex III of the mitochondrial ETC acts as the hypoxic sensor in HPV, and initiates the rise in smooth muscle [Ca²⁺]_i by a mechanism unrelated to changes in cytosolic redox state per se, but more probably by increased production of superoxide. Additionally, glucose and glycolysis are essential for development of the sustained phase 2 of HPV, and support an endothelium-dependent Ca²⁺-sensitisation pathway rather than the rise in [Ca²⁺]_i.

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				J. P. T. Ward Point:Counterpoint: Hypoxic pulmonary vasoconstriction is/is not mediated by increased production of reactive oxygen species J Appl Physiol, September 1, 2006; 101(3): 993 - 995. [Full Text] [PDF]

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				P. J. Kemp Detecting acute changes in oxygen: will the real sensor please stand up? Exp Physiol, September 1, 2006; 91(5): 829 - 834. [Abstract] [Full Text] [PDF]

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				T. P. Robertson, P. I. Aaronson, and J. P. T. Ward Ca2+ sensitization during sustained hypoxic pulmonary vasoconstriction is endothelium dependent Am J Physiol Lung Cell Mol Physiol, June 1, 2003; 284(6): L1121 - L1126. [Abstract] [Full Text] [PDF]

				R. Paddenberg, B. Ishaq, A. Goldenberg, P. Faulhammer, F. Rose, N. Weissmann, R. C. Braun-Dullaeus, and W. Kummer Essential role of complex II of the respiratory chain in hypoxia-induced ROS generation in the pulmonary vasculature Am J Physiol Lung Cell Mol Physiol, May 1, 2003; 284(5): L710 - L719. [Abstract] [Full Text] [PDF]

				Y.-X. Wang, Y.-M. Zheng, I. Abdullaev, and M. I. Kotlikoff Metabolic inhibition with cyanide induces calcium release in pulmonary artery myocytes and Xenopus oocytes Am J Physiol Cell Physiol, February 1, 2003; 284(2): C378 - C388. [Abstract] [Full Text] [PDF]

				C. Schroedl, D. S. McClintock, G. R. S. Budinger, and N. S. Chandel Hypoxic but not anoxic stabilization of HIF-1alpha requires mitochondrial reactive oxygen species Am J Physiol Lung Cell Mol Physiol, November 1, 2002; 283(5): L922 - L931. [Abstract] [Full Text] [PDF]

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				G. B. Waypa, J. D. Marks, M. M. Mack, C. Boriboun, P. T. Mungai, and P. T. Schumacker Mitochondrial Reactive Oxygen Species Trigger Calcium Increases During Hypoxia in Pulmonary Arterial Myocytes Circ. Res., October 18, 2002; 91(8): 719 - 726. [Abstract] [Full Text] [PDF]

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