Multiple phospholipase activation by OX(1) orexin/hypocretin receptors.
We investigated coupling of OX(1) receptors to phospholipase activation and diacylglycerol generation in Chinese hamster ovary (CHO) cells using both biochemical and fluorescence "real-time" methods. The results indicate that at lowest orexin-A concentrations (highest potency), diacylglycerol generated results from phospholipase D activity. At 10-100-fold higher orexin-A concentrations, phospholipase C is activated, likely hydrolyzing phosphatidylinositol (PI) or phosphatidylinositol monophosphate (PIP) but not phosphatidylinositol bisphosphate (PIP(2)). At further 7-fold higher orexin-A concentrations, PIP(2) is hydrolyzed, releasing both diacylglycerol and inositol-1,4,5-trisphosphate. Thus, OX(1) orexin receptors connect to multiple phospholipase activities, apparently composed of at least one phospholipase D and two different phospholipase C activities. At low agonist concentrations, diacylglycerol and phosphatidic acid are the preferred products, and interestingly, it seems that even the primarily activated phospholipase C mainly works to increase diacylglycerol and not inositol-1,4,5-trisphosphate.
Johansson
L
L
Department of Neuroscience, Unit of Physiology, Uppsala University, Uppsala, Sweden.
Ekholm
M E
ME
Kukkonen
J P
JP
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004075:Diglycerides / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000494:pharmacology; D009479:Neuropeptides / Q000494:pharmacology; D064446:Orexin Receptors; D000068797:Orexins; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D018013:Receptors, Neuropeptide / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism*
D004075:Diglycerides; C000596229:HCRT protein, human; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides; D064446:Orexin Receptors; D000068797:Orexins; D019269:Phosphatidylinositol 4,5-Diphosphate; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide; D010738:Type C Phospholipases; D010739:Phospholipase D