P2Y receptor subtypes evoke different Ca²⁺ signals in cultured aortic smooth muscle cells

Abstract

Adenine and uridine nucleotides evoke Ca²⁺ signals via four subtypes of P2Y receptor in cultured aortic smooth muscle cells, but the mechanisms underlying the different patterns of these Ca²⁺ signals are unresolved. Cytosolic Ca²⁺ signals were recorded from single cells and populations of cultured rat aortic smooth muscle cells, loaded with a fluorescent Ca²⁺ indicator and stimulated with agonists that allow subtype-selective activation of P2Y1, P2Y2, P2Y4, or P2Y6 receptors. Activation of P2Y1, P2Y2, and P2Y6 receptors caused homologous desensitisation, while activation of P2Y2 receptors also caused heterologous desensitisation of the other subtypes. The Ca²⁺ signals evoked by each P2Y receptor subtype required activation of phospholipase C and release of Ca²⁺ from intracellular stores via inositol 1,4,5-trisphosphate (IP₃) receptors, but they were unaffected by inhibition of ryanodine or nicotinic acid adenine dinucleotide phosphate (NAADP) receptors. Sustained Ca²⁺ signals were independent of the Na⁺/Ca²⁺ exchanger and were probably mediated by store-operated Ca²⁺ entry. Analyses of single cells established that most cells express P2Y2 receptors and at least two other P2Y receptor subtypes. We conclude that four P2Y receptor subtypes evoke Ca²⁺ signals in cultured aortic smooth muscle cells using the same intracellular (IP₃ receptors) and Ca²⁺ entry pathways (store-operated Ca²⁺ entry). Different rates of homologous desensitisation and different levels of receptor expression account for the different patterns of Ca²⁺ signal evoked by each P2Y receptor subtype.