Purinergic receptor regulation of signal transduction in NCB-20 cells.
In the present paper, P1 and P2 purinergic receptors and their control of signal transduction pathways were investigated in NCB-20 cells. ATP elicited an increase in [Ca2+]i. The purinergic receptor subtype involved was identified by comparing the actions of a range of nucleotides. UTP was the most potent agonist in elevating [Ca2+]i, with an EC50 value of 6.2 +/- 0.5 microM. UTP, ATP (EC50, 17.3 +/- 1.5 microM), adenosine-5'-O-(3-thio)triphosphate (23 +/- 3 microM), and ITP (55 +/- 4 microM) exerted similar maximal effects. Other nucleotides tested, including beta, gamma-methylene-ATP and 2-methylthio-ATP, which are considered prototypic agonists for P2x and P2y receptors, respectively, were ineffective; in general, modifications in the ribose-triphosphate chain and substitution on the 2-position of the purines reduced the efficacy of nucleotides. This pharmacological characterization indicated that a putative P2u receptor mediates the [Ca2+]i elevation elicited by nucleotides in NCB-20 cells. The increase in [Ca2+]i originates from intracellular Ca2+ stores; blockade of Ca2+ entry does not affect the rise in [Ca2+]i. In contrast, pretreatment with the Ca(2+)-ATPase inhibitor thapsigargin or with bradykinin, a hormone that releases Ca2+ from inositol trisphosphate-sensitive stores, does preclude the increase in [Ca2+]i induced by ATP. ATP and UTP also transiently inhibit cAMP accumulation in the intact cell, presumably via a Ca(2+)-mediated mechanism. The finding of a P2u receptor in NCB-20 cells adds to a growing perception that P2 receptors are widely distributed. Besides the P2u receptor, NCB-20 cells express adenosine A2 receptors, coupled to stimulation of cAMP accumulation. The presence of both P1 and P2 purinergic receptors permits a sequential modulation of distinct second messenger levels associated with a common stimulus, ATP.
Garritsen
A
A
Department of Pharmacology, University of Colorado Health Sciences Center, Denver 80262.
Zhang
Y
Y
Cooper
D M
DM
D000227:Adenine Nucleotides / Q000494:pharmacology; D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002413:Cations, Divalent; D002855:Chromatography, Thin Layer; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D006822:Hybrid Cells; D007293:Inosine Triphosphate / Q000494:pharmacology; D051379:Mice; D011983:Receptors, Purinergic / Q000187:drug effects / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D014544:Uridine Triphosphate / Q000494:pharmacology
D000227:Adenine Nucleotides; D002413:Cations, Divalent; D011983:Receptors, Purinergic; D013729:Terpenes; D007293:Inosine Triphosphate; D004533:Egtazic Acid; D019284:Thapsigargin; D000242:Cyclic AMP; D001920:Bradykinin; D002118:Calcium; D014544:Uridine Triphosphate