D. Sbrissa et al., BOMBESIN-STIMULATED CERAMIDE PRODUCTION AND MAP KINASE ACTIVATION IN RABBIT RECTOSIGMOID SMOOTH-MUSCLE CELLS, American journal of physiology: Gastrointestinal and liver physiology, 35(6), 1997, pp. 1615-1625
We have investigated the hypotheses that 1) bombesin activation of pro
tein 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 act
ivation of PKC and mitogen-activated protein (MAP) kinase. Ceramide pr
oduction 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 fr
om the rabbit rectosigmoid and stimulated with bombesin gave a signifi
cant increase in newly formed ceramide (38 +/- 3.5%). 12-O-tetradecano
ylphorbol-13-acetate also induced production of ceramide, which was bl
ocked by calphostin C. The short-chain permeable C-2 ceramide induced
a sustained contraction and activation of MAP kinase, which was blocke
d 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 smoot
h muscle cells by bombesin results in a functional coupling between sn
-1,2-diacylglycerol (DAG)/PKC and a sphingomyelinase, whereby DAG acti
vates the hydrolysis of sphingomyelin to produce ceramide. Ceramide in
turn activates PKC, which then activates MAP kinase. This could be th
e basis for the sustained contraction observed with bombesin.