Halothane regulates G-protein-dependent phospholipase C activity in turkey erythrocyte membranes.
The ability of halothane to stimulate phospholipase C (PLC) was examined in turkey erythrocyte membranes prepared from [3H]inositol-labeled turkey erythrocytes by measuring [3H]inositol phosphate formation ([3H]InsP) in the presence and absence of G-protein activation. In the presence of guanosine 5'-3-O-(thio)triphosphate) (GTP gamma S), halothane (0.5-10 mM) caused a dose-dependent activation of PLC. The EC50 value for halothane-induced PLC activation was 2.8 +/- 0.3 mM. Halothane (0.1-30 mM) had no effect on PLC activity in the absence of G-protein activation and did not affect Ca(2+)-dependent PLC activity. The activation of PLC by GTP gamma S occurred after an initial lag period of 60 s which was followed by a linear increase in [3H]InsP. Halothane dose-dependently decreased the lag period for GTP gamma S-induced PLC activation (minimal value 15 s) and increased the rate of [3H]InsP formation at all time points following this lag. As a result, halothane shifted the EC50 value for GTP gamma S-induced PLC activation to the left (4-fold) and increased its maximal response. Halothane also caused a dose-dependent activation of PLC in the presence of AlF4-. Half-maximal stimulation of AlF4(-)-activated PLC occurred with an EC50 value of 2.9 +/- 0.4 mM halothane, which is similar to the halothane dose giving half-maximal stimulation of PLC in the presence of GTP gamma S. At low doses (0.1-0.3 mM) halothane inhibited both isoproterenol- and adenosine 5'-O-(2-thiodiphosphate) (ADP beta S)-induced [3H]InsP formation, whereas at higher concentrations it stimulated PLC independent of the presence of these agonists. At concentrations chosen to reflect their different membrane/buffer partition coefficients, both hexanol (5 mM) and benzyl alcohol (20 mM) fluidized turkey erythrocyte membranes to the same degree as halothane (5 mM). However, these agents had no effect on GTP gamma S- or AlF(4-)-induced PLC activity, indicating that halothane-induced PLC activation was not secondary to changes in bulk lipid fluidity properties. Halothane also stimulated [3H]inositol bisphosphate and [3H]inositol trisphosphate formation in intact erythrocytes. These data demonstrate that the anesthetic halothane can stimulate G-protein-dependent PLC activity and modify the responsiveness of this signaling system to activation by receptor-linked agonists.
Rooney
T A
TA
Department of Pathology and Cell Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Hager
R
R
Stubbs
C D
CD
Thomas
A P
AP
D000535:Aluminum / Q000494:pharmacology; D017607:Aluminum Compounds; D000818:Animals; D004789:Enzyme Activation; D004910:Erythrocyte Membrane / Q000201:enzymology*; D005459:Fluorides / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006221:Halothane / Q000494:pharmacology*; D007295:Inositol Phosphates / Q000096:biosynthesis; D007700:Kinetics; D008563:Membrane Lipids / Q000378:metabolism; D015398:Signal Transduction; D014422:Turkeys; D010738:Type C Phospholipases / Q000378:metabolism*
D017607:Aluminum Compounds; D007295:Inositol Phosphates; D008563:Membrane Lipids; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000535:Aluminum; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D005459:Fluorides; D006221:Halothane; C032311:aluminum fluoride