Distinct hyperpolarizing and relaxant roles for gap junctions and endothelium-derived H2O2 in NO-independent relaxations of rabbit arteries.
We have compared the contributions of gap junctional communication and chemical signaling via H2O2 to NO-independent relaxations evoked by the Ca2+ ionophore A23187 and acetylcholine (ACh) in rabbit ilio-femoral arteries. Immunostaining confirmed the presence of connexins (Cxs) 37 and 40 in the endothelium and Cxs 40 and 43 in smooth muscle. Maximal endothelium-dependent subintimal smooth muscle hyperpolarizations evoked by A23187 and ACh were equivalent (approximately 20 mV) and almost abolished by an inhibitory peptide combination targeted against Cxs 37, 40, and 43. However, maximal NO-independent relaxations evoked by A23187 were unaffected by such peptides, whereas those evoked by ACh were depressed by approximately 70%. By contrast, the enzyme catalase, which destroys H2O2, attenuated A23187-induced relaxations over a broad range of concentrations, but only minimally depressed the maximum response to ACh. Catalase did not affect A23187- or ACh-evoked hyperpolarizations. After loading with an H2O2-sensitive probe, A23187 caused a marked increase in endothelial fluorescence that correlated temporally with relaxation, whereas only a weak delayed increase was observed with ACh. In arteries without endothelium, the H2O2-generating system xanthine/xanthine oxidase induced a catalase-sensitive relaxation that mimicked the gap junction-independent response to A23187 as it was maximally equivalent to approximately 80% of induced tone, but associated with a smooth muscle hyperpolarization <5 mV. We conclude that myoendothelial gap junctions underpin smooth muscle hyperpolarizations evoked by A23187 and ACh, but that A23187-induced relaxation is dominated by extracellular release of H2O2. Endothelium-derived H2O2 may thus be regarded as a relaxing factor, but not a hyperpolarizing factor, in rabbit arteries.
Chaytor
Andrew T
AT
Department of Diagnostic Radiology, Wales Heart Research Institute, University of Wales College of Medicine, Heath Park, Cardiff CF14 4XN, United Kingdom.
Edwards
David H
DH
Bakker
Linda M
LM
Griffith
Tudor M
TM
D000109:Acetylcholine / Q000378:metabolism; D000818:Animals; D001158:Arteries / Q000378:metabolism*; D001681:Biological Assay; D000001:Calcimycin / Q000494:pharmacology; D018031:Connexin 43 / Q000378:metabolism; D017630:Connexins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004727:Endothelium / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism; D017629:Gap Junctions / Q000502:physiology*; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism*; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials; D018613:Microscopy, Confocal; D009126:Muscle Relaxation; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D011817:Rabbits; D014969:Xanthine Oxidase / Q000378:metabolism; D000097003:Gap Junction alpha-5 Protein; D000097006:Gap Junction alpha-4 Protein
D000109:Acetylcholine; D000001:Calcimycin; D018031:Connexin 43; D017630:Connexins; D006861:Hydrogen Peroxide; D007476:Ionophores; D009569:Nitric Oxide; D010455:Peptides; D014969:Xanthine Oxidase; D000097003:Gap Junction alpha-5 Protein; D000097006:Gap Junction alpha-4 Protein