Phorbol ester and vasopressin activate phospholipase D in Leydig cells.
In the present study evidence is provided for the existence of phospholipase D (PLD) activity in rat Leydig cells. Leydig cells were cultured and labelled with [3H]myristic acid. In the presence of ethanol, phorbol 12-myristate 13-acetate (PMA) stimulated the formation of [3H]phosphatidylethanol ([3H]PEt) in a dose-dependent manner at the expense of [3H]phosphatidic acid ([3H]PA). In cells prelabelled with [3H]choline, PMA caused a rapid increase in intracellular free [3H]choline. The time course of [3H]PEt formation was similar to the time course of intracellular [3H]choline formation. The data taken together support the notion that PMA stimulates phosphatidylcholine (PC) hydrolysis by a mechanism, which principally involves PLD. Activation of PLD by PMA was inhibited by long-term pretreatment of cells with PMA to downregulate protein kinase C (PKC) and by pretreatment with staurosporine. These data support the notion that activation of PLD by PMA is dependent on PKC. Arginine vasopressin (AVP) caused a rapid stimulation of PLD activity in the cells. This activation was inhibited after down-regulation of PKC, indicating that the agonist acts by a mechanism similar to that of PMA.
Department of Biological Sciences, Royal Danish School of Pharmacy, Copenhagen.
A M
AM
Vinggaard
H S
HS
Hansen
D000818:Animals; D001127:Arginine Vasopressin / Q000494:pharmacology*; D002478:Cells, Cultured; D002794:Choline / Q000378:metabolism; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004789:Enzyme Activation; D007700:Kinetics; D007985:Leydig Cells / Q000187:drug effects / Q000201:enzymology*; D008297:Male; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology; D010712:Phosphatidic Acids / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D009227:Myristic Acids; D010703:Phorbol Esters; D010712:Phosphatidic Acids; D019814:Myristic Acid; D001127:Arginine Vasopressin; C008876:phorbol-12,13-didecanoate; D010739:Phospholipase D; D002794:Choline; D013755:Tetradecanoylphorbol Acetate; D004121:Dimethyl Sulfoxide