Activation of PtdIns(4,5)P2-sensitive phospholipase C in rabbit tracheal epithelial cells.
A role for phospholipase C hydrolysis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] as a mechanism of alpha 2-adrenergic signal transduction in rabbit tracheal epithelial cells (tracheocytes) was investigated in isolated cells grown in in vitro culture and prelabeled with myo-[3H]inositol (3 microCi/ml) for 72 h. Breakdown of polyphosphoinositides was measured by using thin-layer chromatography to detect phosphatidylinositol, phosphatidylinositol 4-phosphate [PtdIns(4)P], and PtdIns(4,5)P2. Inositol phosphates were separated by ion-exchange column chromatography. The endogenous catecholamine l-epinephrine and alpha 2-adrenergic agonists clonidine and 1-(2,6-dichlorobenzylideneamino)guanidine (guanabenz) produced a rapid transient accumulation of inositol trisphosphate and inositol 4,5-bisphosphate and breakdown of [PtdIns(4)P] and PtdIns(4,5)P2. The alpha 2-adrenergic effects were not blocked by the beta-adrenergic antagonist DL-propranolol or by the alpha 1-adrenergic antagonists prazosin and methylurapidil but were inhibited by pertussis toxin and blocked by yohimbine, an alpha 2-adrenergic antagonist. The 50% effective concentration for guanabenz-stimulated inositol trisphosphate generation was right shifted from 0.3 to 0.9 microM by yohimbine. The results provide the first demonstration of alpha 2A-adrenergic activation of pertussis toxin-sensitive PtdIns(4,5)P2-dependent phospholipase C in mammalian tracheocytes. The findings are consistent with previous observations on alpha 2A-adrenergic-mediated activation of NaCl cotransport in these cells.
Liedtke
C M
CM
Department of Pediatrics, Case Western Reserve University, Cleveland, Ohio 44106.
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D004837:Epinephrine / Q000494:pharmacology; D004847:Epithelial Cells; D004848:Epithelium / Q000201:enzymology; D007294:Inositol / Q000378:metabolism; D007295:Inositol Phosphates / Q000378:metabolism; D050356:Lipid Metabolism; D008297:Male; D037342:Pertussis Toxin; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000494:pharmacology*; D011817:Rabbits; D014132:Trachea / Q000166:cytology / Q000201:enzymology*; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000316:Adrenergic alpha-Agonists; D000317:Adrenergic alpha-Antagonists; D007295:Inositol Phosphates; D010566:Virulence Factors, Bordetella; D007294:Inositol; D037342:Pertussis Toxin; D017853:Phosphotransferases (Alcohol Group Acceptor); C068961:phosphatidylinositol 4,5-biphosphate kinase; D010738:Type C Phospholipases; D004837:Epinephrine