Inhibition of nitric oxide synthase activity and nitric oxide-dependent calcium influx in renal epithelial cells by cyclic adenosine monophosphate: implications for cell injury.
Cell injury frequently occurs in the setting of tissue destruction and inflammation and is associated with a rise in intracellular calcium (Cai) and increased NO production. The mechanisms that trigger rises in Cai and NO during cell injury are not fully defined, but they may involve activation of G protein-coupled receptors for substances such as bradykinin, Ang II, thromboxane, and thrombin. These receptors act through G proteins from different families that have distinct functions. Receptors for bradykinin and Ang II act through members of the G alpha i and G alpha q families, whereas receptors for thrombin and thromboxane act through members of the G alpha i, G alpha q, and G alpha 12/13 families. These G proteins cooperate to regulate Cai and NO in epithelial cells through distinct mechanisms. In a number of experimental settings, activators of the adenylyl cyclase system reduce the severity of cell injury. To understand the mechanisms by which G protein-dependent signaling systems may contribute to cell injury and to define the role of adenylyl cyclase in ameliorating cell injury, the effects of adenylyl cyclase on bradykinin-stimulated Ca influx and NO in cultured renal epithelial cells that stably overexpress G alpha q and G alpha 13 were studied. This system allowed for the separation of different components of the signals initiated by receptors for thromboxane and thrombin. G alpha 13 increased bradykinin-stimulated Ca influx by a mechanism that depends on NO and cGMP. The increased Ca influx was blocked by inhibitors of NO synthase and guanylyl cyclase and by activation of adenylyl cyclase. NO production was inhibited by activators of cAMP-dependent protein kinase, which indicated that cAMP blocks Ca influx by inhibiting NO production. Expression of G alpha q, the G protein that regulates phospholipase C, also increased bradykinin-stimulated Ca influx, but by an NO, cGMP-independent mechanism that was insensitive to inhibition by adenylyl cyclase. The authors conclude that Ca influx is modulated by NO-dependent and independent mechanisms, and that to the extent that increased NO production contributes to increased Ca influx and cell injury, cell injury may be reduced by agents that activate adenylyl cyclase.
Kitamura
K
K
Third Department of Internal Medicine, Kumamoto University, Japan.
Tomita
K
K
Miller
R T
RT
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D001693:Biological Transport, Active; D002118:Calcium / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism*; D004789:Enzyme Activation; D004790:Enzyme Induction; D004847:Epithelial Cells / Q000201:enzymology / Q000378:metabolism*; D005453:Fluorescence; D005470:Fluorometry; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D009569:Nitric Oxide / Q000037:antagonists & inhibitors*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D052247:Nitric Oxide Synthase Type II
D009569:Nitric Oxide; D000242:Cyclic AMP; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D000262:Adenylyl Cyclases; D006152:Cyclic GMP; D002118:Calcium