title: Effects of organ culture on arterial gene expression and hypoxic relaxation: role of the ryanodine receptor.
abstract: Organ culture specifically inhibits vasorelaxation to acute hypoxia and preferentially decreases specific voltage-dependent K(+) channel expression over other K(+) and Ca(2+) channel subtypes. To isolate further potential oxygen-sensing mechanisms correlated with altered gene expression, we performed differential display analysis on RNA isolated from control and cultured coronary arterial rings. We hypothesize that organ culture results in altered gene expression important for vascular smooth muscle contractility important to the mechanism of hypoxia-induced relaxation. Our results indicate a milieu of changes suggesting both up- and downregulation of several genes. The altered expression pattern of two positive clones was verified by Northern analysis. Subsequent screening of a porcine cDNA library indicated homology to the ryanodine receptor (RyR). RT-PCR using specific primers to the three subtypes of RyR shows an upregulation of RyR2 and RyR3 after organ culture. Additionally, the caffeine- and/or ryanodine-sensitive intracellular Ca(2+) store was significantly more responsive to caffeine activation after organ culture. Our data indicate that organ culture increases expression of specific RyR subtypes and inhibits hypoxic vasorelaxation. Importantly, ryanodine blunted hypoxic relaxation in control coronary arteries, suggesting that upregulated RyR might play a novel role in altered intracellular Ca(2+) handling during hypoxia.
authors.affiliation: Department of Molecular and Cellular Physiology, University of Cincinnati, College of Medicine, Cincinnati, Ohio 45267-0576, USA. george.thorne@amedd.army.mil
authors.forename: George D
authors.initials: GD
authors.lastname: Thorne
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Paul
mesh_terms: D000818:Animals; D001158:Arteries / Q000503:physiopathology*; D001483:Base Sequence / Q000235:genetics; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D018076:DNA, Complementary; D015870:Gene Expression; D020869:Gene Expression Profiling; D015723:Gene Library; D000860:Hypoxia / Q000503:physiopathology*; D007425:Intracellular Membranes / Q000378:metabolism; D008969:Molecular Sequence Data; D009924:Organ Culture Techniques; D020033:Protein Isoforms / Q000378:metabolism; D012016:Reference Values; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012433:Ryanodine / Q000494:pharmacology; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry / Q000502:physiology*; D013552:Swine; D014664:Vasodilation
chemical_list: D018076:DNA, Complementary; D020033:Protein Isoforms; D019837:Ryanodine Receptor Calcium Release Channel; D012433:Ryanodine; D002110:Caffeine; D002118:Calcium