endothelin-1 and mastoparan

Previous studies have observed that endothelin-1 (ET-1) concentration is elevated in CSF and contributes to impaired cerebral hemodynamics following fluid percussion brain injury (FPI) in an age-dependent manner. This study was designed to characterize the effects of FPI on the vascular activity of two activators of a pertussin toxin-sensitive G protein, mastoparan and mastoparan-7, as a function of age and the role of ET-1 in such effects in newborn (1-5 days old) and juvenile (3-4 weeks old) pigs equipped with a closed cranial window. Mastoparan (10(-8), 10(-6) M) elicited pial artery dilation that was blunted more by FPI in newborn versus juvenile pigs (9 +/- 1 and 16 +/- 1 vs. 3 +/- 1 and 5 +/- 1%, newborn; 9 +/- 1 and 15 +/- 1 vs. 6 +/- 1 and 9 +/- 1%, juvenile). Similar results were observed for mastoparan-7, but the inactive analogue mastoparan-17 had no effect on pial diameter. BQ123 (10(-6) M), an ET-1 antagonist, partially restored impaired mastoparan dilation after FPI in the newborn but not in the juvenile (3 +/- 1 and 5 +/- 1 vs. 7 +/- 1 and 11 +/- 1%, newborn; 6 +/- 1 and 9 +/- 1 vs. 6 +/- 1 and 10 +/- 1%, juvenile). These data show that G protein activation elicits cerebrovasodilation that is blunted following FPI in an age-dependent manner. These data suggest that ET-1 contributes to the impairment of G protein-mediated vasodilation in an age-dependent manner after FPI.

Topics: Age Factors; Animals; Animals, Newborn; Brain Injuries; Carbon Dioxide; Cerebrovascular Circulation; Endothelin-1; Female; GTP-Binding Proteins; Hydrogen-Ion Concentration; Intercellular Signaling Peptides and Proteins; Male; Oxygen; Peptides; Pertussis Toxin; Pia Mater; Signal Transduction; Swine; Vasodilation; Wasp Venoms

We have shown previously that cytosolic phospholipase A(2) (cPLA(2)) is responsible for endothelin-1-induced release of arachidonic acid for prostaglandin synthesis in cat iris sphincter smooth muscle (CISM) cells [Husain and Abdel-Latif (1998) Biochim. Biophys. Acta 1392, 127-144]. Here we show that p38 mitogen-activated protein (MAP) kinase, but not p42/p44 MAP kinases, plays an important role in the phosphorylation and activation of cPLA(2) in endothelin-1-stimulated CISM cells. This conclusion is supported by the following findings. Both p38 MAP kinase and p42/p44 MAP kinases were present in the CISM cells and both were activated by endothelin-1. SB203580, a potent specific inhibitor of p38 MAP kinase, but not the p42/p44 MAP kinases specific inhibitor, PD98059, markedly suppressed endothelin-1-enhanced cPLA(2) phosphorylation, cPLA(2) activity and arachidonic acid release. The addition of endothelin-1 resulted in the phosphorylation and activation of cPLA(2). Endothelin-1 stimulated p38 MAP kinase activity in a time- and concentration-dependent manner, and these effects were mediated through the endothelin-A receptor subtype. The protein kinase C (PKC) inhibitor, RO 31-8220, had no inhibitory effect on endothelin-1-induced p38 MAP kinase activation, suggesting that endothelin-1 activation of p38 MAP kinase is independent of PKC. Pertussis toxin inhibited both endothelin-1 and mastoparan stimulation of p38 MAP kinase activity and arachidonic acid release. The inhibitory effects of pertussis toxin are not mediated through cAMP formation. Mastoparan-stimulated [(3)H]arachidonic acid release and cPLA(2) activation was inhibited by SB203580, but not by RO 31-8220. These data suggest that endothelin-1 binds to the endothelin-A receptor to activate the Gi-protein which, through a series of kinases, leads to the activation of p38 MAP kinase and subsequently to phosphorylation and activation of cPLA(2). Activation of cPLA(2) leads to the liberation of arachidonic acid from membrane phospholipids. The ability of the activated endothelin-A receptor, which is coupled to both Gq- and Gi-proteins, to recruit and activate this complex signal transduction pathway remains to be elucidated. Further studies on the mechanism of these relationships could provide important information about the functions of p38 MAP kinase in smooth muscle.

Topics: Animals; Arachidonic Acid; Calcium-Calmodulin-Dependent Protein Kinases; Cats; Cells, Cultured; Cyclic AMP; Cytosol; Dose-Response Relationship, Drug; Endothelin Receptor Antagonists; Endothelin-1; Enzyme Activation; Flavonoids; GTP-Binding Proteins; Imidazoles; Indoles; Intercellular Signaling Peptides and Proteins; Iris; Mitogen-Activated Protein Kinases; Muscle, Smooth; p38 Mitogen-Activated Protein Kinases; Peptides; Pertussis Toxin; Phospholipases A; Phospholipases A2; Phosphorylation; Pyridines; Receptors, Endothelin; Virulence Factors, Bordetella; Wasp Venoms

The presence of functional endothelin receptors and their signal transduction mechanism has not been determined so far in the pineal gland. We examined the effect of endothelin-1 (ET-1) on phosphoinositide turnover in whole pineal gland. Endothelin-1 increased monophosphate accumulation in a dose-dependent manner. The phosphoinositide (PI) response elicited by ET-1 was dependent on the presence of extracellular Ca (++) since its chelation resulted in a marked decrease in ET-1-stimulated InsP(1) accumulation. On the contrary, phosphoinositide hydrolysis was not changed by the calcium blocker amlodipine. ET-1 induced PI breakdown was inhibited by neomycin, an inhibitor of phospholipase C. However, mastoparan 7, a G protein activator via Gi/Go s timulation, did not alter ET-1-induced InsP(1) accumulation. Our data indicate that stimulation of PI turnover constitutes one of the signaling pathways of ET in rat pineal gland through the stimulation of a receptor-coupled phospholipase C. And they demonstrate, for the first time, the presence of functional binding sites for endothelin in the pineal gland.

Topics: Amlodipine; Animals; Calcium; Calcium Channel Blockers; Chelating Agents; Egtazic Acid; Endothelin-1; Enzyme Inhibitors; Inositol Phosphates; Intercellular Signaling Peptides and Proteins; Male; Neomycin; Peptides; Phosphatidylinositol Diacylglycerol-Lyase; Phosphatidylinositols; Pineal Gland; Rats; Rats, Sprague-Dawley; Receptors, Endothelin; Second Messenger Systems; Type C Phospholipases; Wasp Venoms