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Regulation by hypoxia of endothelin‐1‐stimulated phospholipase D activity in sheep pulmonary artery cultured smooth muscle cells
Author(s) -
Plevin Robin,
Kellock Nicola A.,
Wakelam Michael J.O.,
Wadsworth Roger
Publication year - 1994
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1994.tb13070.x
Subject(s) - protein kinase c , endocrinology , medicine , phospholipase c , stimulation , hypoxia (environmental) , biology , endothelin 1 , activator (genetics) , agonist , endothelins , phospholipase d , chemistry , phosphorylation , receptor , biochemistry , signal transduction , organic chemistry , oxygen
1 The aim of the study was to characterize the effects of hypoxia on agonist‐stimulated phospholipase D (PLD) and phospholipase C activity of sheep pulmonary artery cultured smooth muscle cells. 2 Endothelin‐1 (ET‐1), 5‐hydroxytryptamine (5‐HT) and the protein kinase C (PKC) activator tetradecanoylphorbol acetate (TPA), stimulated a time‐ and concentration‐dependent increase in [ 3 H]‐phosphatidylbutanol accumulation. This was abolished by pretreatment of the cells with the PKC inhibitor, Ro‐318220, suggesting that agonist‐stimulated phospholipase D activity is dependent upon the activation of PKC. 3 Hypoxia (Po 2 20 mmHg for 30 min) stimulated basal [ 3 H]‐phosphatidylbutanol accumulation by approximately 2 fold and this activity was abolished by preincubation of the cells with 10 μ m Ro‐318220. 4 In cells preincubated in low O 2 containing medium for 30 min, the subsequent agonist‐stimulated accumulation of [ 3 H]‐phosphatidylbutanol was reduced. However, the decrease in stimulation was greater for ET‐1 and 5‐HT than for TPA. 5 ET‐1 and TPA stimulated a time‐dependent increase in protein kinase C‐ mediated psuedosubstrate phosphorylation. Following preincubation for 30 min in low O 2 containing media, basal pseudosubstrate phosphorylation increased whilst the fold stimulation by TPA and ET‐1 decreased. 6 In cells preincubated in low O 2 containing medium, ET‐1‐stimulated [ 3 H]‐inositol phosphate accumulation was reduced by approximately 30– 40%. This reduction was reversed by preincubation of the cells with Ro‐318220. 7 These results suggest a role for PKC in the effects of hypoxia on PLD in pulmonary artery smooth muscle cells.