Pharmacological characterization of P2X7 receptors in rat peritoneal cells.
OBJECTIVE
P2X(7) receptor activation by ATP results in the release of IL-1beta and IL-18. Prolonged stimulation can lead to pore formation and cell death. In this study we pharmacologically characterized P2X(7) receptors on rat peritoneal cells (RPC) and on 1321N1 cells transfected with rat P2X(7) receptor (1321rP2X(7)-11).


METHODS
RPC were isolated from rats by lavage. P2X(7) agonist induced pore formation in RPC was measured by EtBr uptake. P2X(7)-stimulated pore formation and Ca(++) influx in 1321rP2X(7)-11 cells were measured by a fluorometric imaging plate reader. The effects of pyridoxal phosphate-6-azo phenyl -2'-4'-disulfonic acid (PPADS) on pore formation and Ca(++) influx were examined in both RPC and 1321rP2X(7)-11. P2X(7)-mediated IL-1beta release in RPC and the effect of PPADS were determined.


RESULTS
RPC express functional P2X(7) receptors that were activated by ATP analogs with a rank order of potency of 2'- 3'-O-(4-Benzoylbenzoyl) adenosine 5'-triphosphate (BzATP) > ATP > alpha,beta-methylene ATP. Activation of P2X(7) receptors by BzATP was inhibited by PPADS. Similar results were also obtained in 1321rP2X(7)-11 cells. Activation of P2X(7) receptors on RPC resulted in IL-1 beta secretion, which was inhibited by PPADS.


CONCLUSIONS
RPC express functional P2X(7) receptors that form pores and mediate the release of IL-1beta.
Chen
Y-W
YW
Global Pharmaceutical Research and Development, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064-6119, USA. yung-wu.chen@abbott.com
Donnelly-Roberts
D L
DL
Namovic
M T
MT
Gintant
G A
GA
Cox
B F
BF
Jarvis
M F
MF
Harris
R R
RR
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D017252:CD3 Complex / Q000096:biosynthesis; D002118:Calcium / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004996:Ethidium / Q000494:pharmacology; D005434:Flow Cytometry; D007249:Inflammation; D007375:Interleukin-1 / Q000378:metabolism; D020382:Interleukin-18 / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008024:Ligands; D008070:Lipopolysaccharides / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D010537:Peritoneum / Q000166:cytology*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000737:chemistry / Q000502:physiology*; D058486:Receptors, Purinergic P2X7; D013997:Time Factors; D014162:Transfection
D017252:CD3 Complex; D007375:Interleukin-1; D020382:Interleukin-18; D008024:Ligands; D008070:Lipopolysaccharides; C546470:P2rx7 protein, rat; D010975:Platelet Aggregation Inhibitors; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; C077792:pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D011732:Pyridoxal Phosphate; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D004996:Ethidium; D002118:Calcium