Bombesin-stimulated ceramide production and MAP kinase activation in rabbit rectosigmoid smooth muscle cells.
We have investigated the hypotheses that 1) bombesin activation of protein kinase C (PKC) results in the hydrolysis of sphingolipids and the production of ceramide and that 2) ceramide produced on activation by bombesin mediates sustained contraction of smooth muscle cells by activation of PKC and mitogen-activated protein (MAP) kinase. Ceramide production was assessed using a technique that involved benzoylation of purified ceramide extracts, followed by reverse-phase high-performance liquid chromatography. Contraction of smooth muscle cells isolated from the rabbit rectosigmoid and stimulated with bombesin gave a significant increase in newly formed ceramide (38 +/- 3.5%). 12-O-tetradecanoylphorbol-13-acetate also induced production of ceramide, which was blocked by calphostin C. The short-chain permeable C2 ceramide induced a sustained contraction and activation of MAP kinase, which was blocked by calphostin C. The increase in MAP kinase activity was maximal at 30 s and declined at 2 min. The data suggest that stimulation of smooth muscle cells by bombesin results in a functional coupling between sn-1,2-diacylglycerol (DAG)/ PKC and a sphingomyelinase, whereby DAG activates the hydrolysis of sphingomyelin to produce ceramide. Ceramide in turn activates PKC, which then activates MAP kinase. This could be the basis for the sustained contraction observed with bombesin.
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Department of Pediatrics, University of Michigan Medical Center, Ann Arbor 48109-0658, USA.
Yamada
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Hajra
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D001003:Anal Canal / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D001839:Bombesin / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002518:Ceramides / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004075:Diglycerides / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007700:Kinetics; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology*; D009281:Naphthalenes / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism; D011817:Rabbits; D013109:Sphingomyelins / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors
D002518:Ceramides; D004075:Diglycerides; D004791:Enzyme Inhibitors; D009281:Naphthalenes; D013109:Sphingomyelins; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C058819:calphostin C; D013755:Tetradecanoylphorbol Acetate; D001839:Bombesin