Expression of purinergic P2X2 receptor-channels and their role in calcium signaling in pituitary cells

Abstract

Pituitary cells express purinergic receptor-channels (P2XR), the activation of which by ATP is associated with the facilitation of Ca2+ influx. Pharmacological, RT-PCR, and nucleotide sequence analyses confirm the presence of a wild type P2X2aR and a spliced isoform P2X2bR, which lacks a portion of carboxyl terminal amino acids. Wild type and spliced isoform receptors have a similar EC50 for ATP and time-course for activation, but the spliced isoform exhibits rapid and complete desensitization, whereas the wild type channel desensitizes slowly and incompletely. Deletion and insertion studies have revealed that a 6 residue sequence located in carboxyl tail (Arg371-Pro376) is required for sustained Ca2+ influx through wild type receptors. When co-expressed, the wild type and spliced channels form functional heteropolymeric channels. The patterns of Ca2+ signaling in the majority of pituitary cells expressing ATP-gated receptor-channels are highly comparable to those observed in cells co-transfected with P2X2aR and P2X2bR. ATP-induced [Ca2+]i response in pituitary cells is partially inhibited by nifedipine, a blocker of voltage-gated L-type Ca2+ channels, suggesting that P2X2R not only drive Ca2+ into the cell, but also activate voltage-gated Ca2+ entry. Our results indicate that ATP represents a paracrine and (or) autocrine factor in the regulation of Ca2+ signaling, and that its actions are mediated in part by heteropolymeric P2X2R. Key words: ATP-gated channels, gonadotrophs, lactotrophs, somatotrophs.