Effects of organ culture on arterial gene expression and hypoxic relaxation: role of the ryanodine receptor

Organ culturespecifically inhibits vasorelaxation to acute hypoxia andpreferentially decreases specific voltage-dependent K⁺channel expression over other K⁺ and Ca²⁺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 culturedcoronary arterial rings. We hypothesize that organ culture results inaltered gene expression important for vascular smooth musclecontractility important to the mechanism of hypoxia-induced relaxation.Our results indicate a milieu of changes suggesting both up- anddownregulation of several genes. The altered expression pattern of twopositive clones was verified by Northern analysis. Subsequent screeningof a porcine cDNA library indicated homology to the ryanodine receptor(RyR). RT-PCR using specific primers to the three subtypes of RyR showsan upregulation of RyR2 and RyR3 after organ culture. Additionally, thecaffeine- and/or ryanodine-sensitive intracellular Ca²⁺store was significantly more responsive to caffeine activation afterorgan culture. Our data indicate that organ culture increases expression of specific RyR subtypes and inhibits hypoxicvasorelaxation. Importantly, ryanodine blunted hypoxic relaxation incontrol coronary arteries, suggesting that upregulated RyR might play anovel role in altered intracellular Ca²⁺ handling during hypoxia.