Article date: August 1996
By: Pierre Pacaud, Erick Feolde, Christian Frelin, Gervaise Loirand, in Volume 118, Issue 8, pages 2213-2219
The P2‐purinoceptor subtype and the intracellular signalling mechanism(s) involved in the rise in the free cytosolic Ca2+ concentration ([Ca2+]i) induced by ATP and analogues were analyzed in myocytes isolated from the longitudinal muscle layer of rat ileum by means of molecular and physiological techniques.
The P2‐purinoceptor expressed by ileal smooth muscle cells shared 100% amino acid identity with the rat P2Y1‐receptor.
Short applications of the purinoceptor agonists induced a transient rise in [Ca2+]i in an all‐or‐nothing manner. The rank order of potency of the analogues of ATP and ADP, determined by measuring the percentage of responding cells was 2‐methylthioATP = 2‐chloro‐ATP > ADP > ATP, with concentrations giving [Ca2+]i response in 50% of cells ranging between 3 nM and 0.6 μm. The concentration‐response curves to ADP and ATP were shifted to the right by 10 μm pyridoxal phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS).
Although the rise in [Ca2+]i induced by stimulation of the ileal P2Y‐purinoceptor was inhibited by heparin (5 mg ml−1), we were not able to detect stimulation of phospholipase C under conditions (37°C) where muscarinic cholinoceptor activation markedly increased inositol phosphate (InsP) accumulation. However, the carbachol (CCh)‐induced increase in InsP accumulation was suppressed when the agonist was applied at 20°C while a CCh‐induced [Ca2+]i rise similar to that obtained in response to the P2‐purinoceptor agonist was still observed.
Our results indicate that the rat ileal myocytes express a PPADS‐sensitive P2‐purinoceptor similar to the P2Y1‐receptor subtype. Although there 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, indicating that they couple to phospholipase C.
The P2‐purinoceptor subtype and the intracellular signalling mechanism(s) involved in the rise in the free cytosolic Ca2+ concentration ([Ca2+]i) induced by ATP and analogues were analyzed in myocytes isolated from the longitudinal muscle layer of rat ileum by means of molecular and physiological techniques.
The P2‐purinoceptor expressed by ileal smooth muscle cells shared 100% amino acid identity with the rat P2Y1‐receptor.
Short applications of the purinoceptor agonists induced a transient rise in [Ca2+]i in an all‐or‐nothing manner. The rank order of potency of the analogues of ATP and ADP, determined by measuring the percentage of responding cells was 2‐methylthioATP = 2‐chloro‐ATP > ADP > ATP, with concentrations giving [Ca2+]i response in 50% of cells ranging between 3 nM and 0.6 μm. The concentration‐response curves to ADP and ATP were shifted to the right by 10 μm pyridoxal phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS).
Although the rise in [Ca2+]i induced by stimulation of the ileal P2Y‐purinoceptor was inhibited by heparin (5 mg ml−1), we were not able to detect stimulation of phospholipase C under conditions (37°C) where muscarinic cholinoceptor activation markedly increased inositol phosphate (InsP) accumulation. However, the carbachol (CCh)‐induced increase in InsP accumulation was suppressed when the agonist was applied at 20°C while a CCh‐induced [Ca2+]i rise similar to that obtained in response to the P2‐purinoceptor agonist was still observed.
Our results indicate that the rat ileal myocytes express a PPADS‐sensitive P2‐purinoceptor similar to the P2Y1‐receptor subtype. Although there 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, indicating that they couple to phospholipase C.
DOI: 10.1111/j.1476-5381.1996.tb15665.x
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