P. Pacaud et al., CHARACTERIZATION OF THE P-2Y-PURINOCEPTOR INVOLVED IN THE ATP-INDUCEDRISE IN CYTOSOLIC CA2+ CONCENTRATION IN RAT ILEAL MYOCYTES, British Journal of Pharmacology, 118(8), 1996, pp. 2213-2219
1 The P-2-purinoceptor subtype and the intracellular signalling mechan
ism(s) involved in the rise in the free cytosolic Ca2+ concentration (
[Ca2+](i)) induced by ATP and analogues were analyzed in myocytes isol
ated from the longitudinal muscle layer of rat ileum by means of molec
ular and physiological techniques. 2 The P-2-purinoceptor expressed by
ileal smooth muscle cells shared 100% amino acid identity with the ra
t P-2Y1-receptor. 3 Short applications of the purinoceptor agonists in
duced a transient rise in [Ca2+](i) in an all-or-nothing manner. The r
ank order of potency of the analogues of ATP and ADP, determined by me
asuring the percentage of responding cells was 2-methylthioATP=2-chlor
o-ATP > ADP > ATP, with concentrations giving [Ca2+](i) response in 50
% of cells ranging between 3 nM and 0.6 mu M. The concentration-respon
se curves to ADP and ATP were shifted to the right by 10 mu M pyridoxa
l phosphate-6-azopheny1-2',4'-disulphonic acid (PPADS). 4 Although the
rise in [Ca2+](i) induced by stimulation of the ileal P-2Y-purinocept
or was inhibited by heparin (5 mg ml(-1)), we were not able to detect
stimulation of phospholipase C under conditions (37 degrees C) where m
uscarinic cholinoceptor activation markedly increased inositol phospha
te (InsP) accumulation. However, the carbachol (CCh)-induced increase
in InsP accumulation was suppressed when the agonist was applied at 20
degrees C while a CCh-induced [Ca2+](i) rise similar to that obtained
in response to the P-2-purinoceptor agonist was still observed. 5 Our
results indicate that the rat ileal myocytes express a PPADS-sensitiv
e P-2-purinoceptor similar to the P-2Yl-receptor subtype. Although the
re is no detectable increase in InsP production, stimulation of these
receptors leads to a rise in [Ca2+](i) by activation of the inositol 1
,4,5-trisphosphate receptor-channel of the intracellular Ca2+ store, i
ndicating that they couple to phospholipase C.