Activation of phospholipase D-2 by P2X7 agonists in rat submandibular gland acini

• DOI: 10.1194/jlr.m100372-jlr200
• Publication Date: 2020-02-24T18:31:07Z
• Authors: Encarnación Pérez...; María Fernández-R...; Mónica González; Ana Zubiaga; Ainara Vallejo; Itxaso García; Carlos Matute; Stéphanie Pochet; Jean Paul Dehaye; Miguel Trueba; Aida Marino; Antonio Gómez-Muñoz
• Keywords: Male; Purinergic P2 Receptor Agonists; 0301 basic medicine; Adenosine Triphosphate -- pharmacology; Submandibular Gland; QD415-436; Purinergic P2 -- agonists; Ceramides; Biochemistry; Cell Line; Rats, Sprague-Dawley; Mice; 03 medical and health sciences; Adenosine Triphosphate; Phospholipase A2; Phospholipase C; Protein kinase C; Receptors; Submandibular Gland -- enzymology; Phospholipase D; phospholipase D; Animals; Submandibular Gland -- cytology; phospholipase C; DNA Primers; calcium; ceramides; Base Sequence; Reverse Transcriptase Polymerase Chain Reaction; Sciences bio-médicales et agricoles; Submandibular Gland -- drug effects; Rats; ATP; Enzyme Activation; Phospholipase D -- metabolism; Sphingosine 1-phosphate; Calcium; phospholipase A2; Sprague-Dawley; Receptors, Purinergic P2X7; Purinergic receptors

Exogenous ATP stimulated phospholipase D (PLD), but not sphingomyelinase in rat submandibular gland (SMG) acini. PLD activation was dependent upon extracellular Ca(2+) and did not involve intracellular Ca(2+) mobilization or phosphoinositide-specific phospholipase C activation. ATP-stimulated PLD was attenuated by inhibition or downregulation of protein kinase C (PKC). PLD activation was fully blocked by the cytosolic phospholipase A(2) (PLA(2)) inhibitor ONO-RS-082 and partially attenuated by the selective Ca(2+)-dependent cytosolic PLA(2) inhibitor, arachidonyl trifluoromethylketone (AACOCF(3)), or by bromoenol lactone, an inhibitor of Ca(2+)-independent cytosolic PLA(2). Magnesium, which decreases the concentration of ATP(4-), and nickel, which blocks nonspecific cation channels coupled to purinergic receptors, inhibited PLD activation by ATP. Using reverse transcription-polymerase chain reaction and Northern blotting techniques, we demonstrated that the PLD isoform stimulated by ATP was PLD-2. Among various ATP analogs, only the P2Z/P2X(7) purinergic receptor agonist benzoyl-benzoyl ATP stimulated PLD-2. The response to ATP was inhibited by the nonselective P2X purinergic antagonist suramin and by oxidized ATP, a potent P2Z/P2X(7) receptor antagonist. It is concluded that in rat SMG acinar cells, PLD-2 is upregulated by exogenous ATP through a mechanism involving Ca(2+) influx, cytosolic PLA(2), and PKC. Also, the data suggest an involvement of P2X(7) receptors in PLD-2 stimulation by ATP.