Contribution of two ionotropic purinergic receptors to ATP responses in submandibular gland ductal cells.
The effect of extracellular ATP on salivary gland function was compared in wild-type (WT) and P2X(7) knockout (KO) mice. The increase in the intracellular concentration of calcium ([Ca(2+)](i)) in response to carbachol was similar in submandibular ductal cells of WT and KO mice. ATP and its analog, benzoyl-ATP, induced a sustained increase in the [Ca(2+)](i) in WT animals. In KO mice, ATP slightly and transiently increased the [Ca(2+)](i) and benzoyl-ATP had no effect. The response to ATP of WT but not KO mice was blocked by KN-62, Coomassie blue and magnesium. The small response of ATP observed in KO mice was completely blocked in the absence of extracellular calcium, unchanged by U73122 and potentiated by ivermectin indicating the probable involvement of a P2X(4) receptor. A RT-PCR and a Western blot confirmed the presence of these receptors in ducts of both WT and KO mice. ATP increased the permeability of the cells to ethidium bromide and stimulated a phospholipase A(2) activity in WT but not KO mice. Mice submandibular gland cells secreted IL-1beta but this secretion was not modified by ATP and was similar in both groups of animals. The volume of saliva provoked by pilocarpine and the concentration of proteins, sodium and chloride in this saliva was similar in both groups of animals. The concentration of potassium was higher in KO mice. We can conclude that the major purinergic receptors expressed in mice submandibular ductal cells are P2X(7) receptors but that P2X(4) receptors are also involved in some ATP effects.
Pochet
Stéphanie
S
Laboratoire de Biochimie et de Biologie Cellulaire, Institut de Pharmacie, C.P. 205/3, Université libre de Bruxelles (ULB), Campus de la Plaine, Boulevard du Triomphe B1050, Brussels, Belgium. spochet@ulb.ac.be
Garcia-Marcos
Mikel
M
Seil
Michèle
M
Otto
Anne
A
Marino
Aida
A
Dehaye
Jean-Paul
JP
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D004996:Ethidium / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D054467:Phospholipases A2 / Q000378:metabolism; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000502:physiology*; D058848:Receptors, Purinergic P2X4; D058486:Receptors, Purinergic P2X7; D012463:Saliva / Q000737:chemistry; D013363:Submandibular Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D053583:Interleukin-1beta; C548227:P2rx4 protein, mouse; C546469:P2rx7 protein, mouse; D018048:Receptors, Purinergic P2; D058848:Receptors, Purinergic P2X4; D058486:Receptors, Purinergic P2X7; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; C063302:KN 62; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate; D002217:Carbachol; D054467:Phospholipases A2; D004996:Ethidium; D002118:Calcium