Dissociation of the pore-forming and phospholipase D activities stimulated via P2z purinergic receptors in BAC1.2F5 macrophages. Product inhibition of phospholipase D enzyme activity.
We have previously reported that the occupancy of P2z purinergic receptors in BAC1.2F5 macrophages stimulates both the formation of nonselective membrane pores/channels and a novel phospholipase D (PLD) activity. In the present study, the relationship between these two P2z receptor-mediated responses was investigated. In [3H]oleic acid-labeled macrophages, isoosmotic replacement of extracellular Na+ with choline completely abolished 3'-O-(4-benzoyl)benzoyl-ATP (BzATP)-stimulated [3H]phosphatidylethanol ([3H]PEt) accumulation. This inhibitory effect of choline was mimicked when Na+ was replaced with other amine-containing cations (N-methylglucamine, ethanolamine, Tris+), but not with inorganic cations (K+, Li+). In contrast, substitution of Na+ with choline (or other cationic amines) did not prevent BzATP-induced Ca2+ influx and membrane depolarization. Thus, choline, which enters the cells through the BzATP-induced pores, can act to inhibit P2z receptor-stimulated PLD activity but not pore formation. The two responses also exhibit differential sensitivities to the pH of the incubation medium, to depletion of intracellular ATP, and to chronic phorbol ester treatment. These observations indicate that P2z receptor-mediated pore formation can be functionally uncoupled from the activation of PLD. However, both pore formation and PLD activity are similarly inhibited when the cells are: 1) partially desensitized by previous treatment with P2z agonists and 2) preincubated with 2',3'-dialdehyde-ATP or 4',4'-diisothiocyanostilbene-2,2'-disulfonate. This suggests that these two distinct responses are mediated by the same, or similar, P2z receptors.
el-Moatassim
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Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106.
Dubyak
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D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002794:Choline / Q000378:metabolism; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007473:Ion Channels / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D010739:Phospholipase D / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011983:Receptors, Purinergic / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D007473:Ion Channels; D011983:Receptors, Purinergic; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D012964:Sodium; D010739:Phospholipase D; D002794:Choline; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium