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pubmed:abstractText |
To demonstrate a Ca(2+)-independent component of hypoxic vasorelaxation and to investigate its mechanism, we utilized permeabilized porcine coronary arteries, in which [Ca(2+)] could be clamped. Arteries permeabilized with beta-escin developed maximum force in response to free Ca(2+) (6.6 microm), concomitant with a parallel increase in myosin regulatory light chain phosphorylation (MRLC-P(i)), from 0.183 +/- 0.023 to 0.353 +/- 0.019 MRLC-P(i) (total light chain)(-1). Hypoxia resulted in a significant decrease in both force (-31.9 +/- 4.1% prior developed force) and MRLC-P(i) (from 0.353 to 0.280 +/- 0.023), despite constant [Ca(2+)] buffered by EGTA (4 mm). Forces developed in response to Ca(2+) (6.6 microm), Ca(2+) (0.2 microm) + GTPgammaS (1 mM), or in the absence of Ca(2+) after treatment with ATPgammaS (1 mM), were of similar magnitude. Hypoxia also relaxed GTPgammaS contractures but importantly, arteries could not be relaxed after treatment with ATPgammaS. Permeabilization with Triton X-100 for 60 min also abolished hypoxic relaxation. The blocking of hypoxic relaxation after ATPgammaS suggests that this Ca(2+)-independent mechanism(s) may operate through alteration of MRLC-P(i) or of phosphorylation of the myosin binding subunit of myosin light chain phosphatase. Treatment with the Rho kinase inhibitor Y27632 (1 microm) relaxed GTPgammaS and Ca(2+) contractures; but the latter required a higher concentration (10 microm) for consistent relaxation. Relaxations to N(2) and/or Y27632 averaged 35% and were not additive or dependent on order. Our data suggest that the GTP-mediated, Rho kinase-coupled pathway merits further investigation as a potential site of this novel, Ca(2+)-independent O(2)-sensing mechanism. Importantly, these results unambiguously show that hypoxia-induced vasorelaxation can occur in permeabilized arteries where the Ca(2+) is clamped at a constant value.
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pubmed:affiliation |
Department of Molecular and Cellular Physiology, University of Cincinnati College of Medicine, 231 Albert Sabin Way, Cincinnati, OH 45267-0576, USA.
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