ucn-1028-c and sphingosine-1-phosphate

Interstitial cells of Cajal (ICC) are the pacemaker cells that generate the rhythmic oscillation responsible for the production of slow waves in gastrointestinal smooth muscle. Spingolipids are known to present in digestive system and are responsible for multiple important physiological and pathological processes. In this study, we are interested in the action of sphingosine 1-phosphate (S1P) on ICC. S1P depolarized the membrane and increased tonic inward pacemaker currents. FTY720 phosphate (FTY720P, an S1P(1,3,4,5) agonist) and SEW 2871 (an S1P(1) agonist) had no effects on pacemaker activity. Suramin (an S1P(3) antagonist) did not block the S1P-induced action on pacemaker currents. However, JTE-013 (an S1P(2) antagonist) blocked the S1P-induced action. RT-PCR revealed the presence of the S1P(2) in ICC. Calphostin C (a protein kinase C inhibitor), NS-398 (a cyclooxygenase-2 inhibitor), PD 98059 (a p42/44 inhibitor), or SB 203580 (a p38 inhibitor) had no effects on S1P-induced action. However, c-jun NH(2)-terminal kinase (JNK) inhibitor II suppressed S1P-induced action. External Ca(2+)-free solution or thapsigargin (a Ca(2+)-ATPase inhibitor of endoplasmic reticulum) suppressed action of S1P on ICC. In recording of intracellular Ca(2+) ([Ca(2+)](i)) concentration using fluo-4/AM S1P increased intensity of spontaneous [Ca(2+)](i) oscillations in ICC. These results suggest that S1P can modulate pacemaker activity of ICC through S1P(2) via regulation of external and internal Ca(2+) and mitogenactivated protein kinase activation.

Topics: Animals; Antinematodal Agents; Calcium; Cells, Cultured; Enzyme Inhibitors; Female; Interstitial Cells of Cajal; Intestine, Small; Ion Channels; Lysophospholipids; Male; Membrane Potentials; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; Naphthalenes; Organophosphates; Patch-Clamp Techniques; Pyrazoles; Pyridines; Receptors, Lysosphingolipid; Signal Transduction; Sphingosine; Sphingosine-1-Phosphate Receptors; Suramin

In rat type I astrocytes and C6 glioma cells, sphingosine 1-phosphate (S1P) clearly induced the expression of fibroblast growth factor-2 (FGF-2) mRNA to an extent comparable to that achieved by platelet-derived growth factor (PDGF) and endothelin. In C6 cells, Western blotting showed that S1P also induced expression of early growth response-1 (Egr-1), one of the immediate early gene products and an essential transcriptional factor for FGF-2 expression. On the other hand, sphingosine, a substrate for sphingosine kinase which forms intracellular S1P, was a very weak activator for the expression of either FGF-2 or Egr-1. The S1P-induced Egr-1 expression was partially inhibited by treatment of the cells with either calphostin C, an inhibitor of protein kinase C (PKC), or pertussis toxin (PTX), and completely inhibited by the combination of these agents. Essentially, the same inhibitory pattern by these agents has been observed for S1P-induced extracellular signal-regulated kinase (ERK) activation. The S1P-induced expression of Egr-1 was also completely inhibited in association with complete inhibition of ERK by PD 98059, an ERK kinase inhibitor. Thus, the S1P-induced activation of the Egr-1/FGF-2 system may be mediated through ERK activation, which may involve at least two signaling pathways, i.e., a PTX-sensitive G-protein-dependent pathway and a PKC-dependent pathway.

Topics: Animals; Animals, Newborn; Astrocytes; Cells, Cultured; DNA-Binding Proteins; Early Growth Response Protein 1; Endothelins; Fibroblast Growth Factor 2; Flavonoids; Gene Expression Regulation; Glioma; GTP-Binding Proteins; Immediate-Early Proteins; Lysophospholipids; Mitogen-Activated Protein Kinases; Naphthalenes; Pertussis Toxin; Platelet-Derived Growth Factor; Protein Kinase C; Rats; RNA, Messenger; Sphingosine; Tetradecanoylphorbol Acetate; Transcription Factors; Tumor Cells, Cultured; Virulence Factors, Bordetella