BIOCHEMICAL-CHARACTERIZATION OF THE STIMULATORY EFFECTS OF HALOTHANE AND PROPOFOL ON PURIFIED BRAIN PROTEIN-KINASE-C

Halothane and propofol stimulate activation of protein kinase C (PKC) in the presence of physiologically relevant lipid bilayer vesicles in vitro. The mechanism of this stimulation was characterized by analyzing the effects of halothane and propofol on the activation of purified rat brain PKC by its three essential activators, phosphatidylserine, diacylglycerol, and Ca2+, each of which is known to interact with the regulatory domain. Clinically relevant concentrations of halothane (2.4 vol%) and propofol (200 mu M) increased the V-max without affecting the K-m for phosphorylation of the artificial substrate histone H1 by PKC, and increased the sensitivity of PKC to activation by phosphatidylserine, diacylglycerol, and Ca2+. Halothane reduced the EC(50) values for phosphatidylserine from 18 +/- 2.5 to 11 +/- 0.6 mol% (P < 0.05), for diacylglycerol from 1.6 +/- 0.3 to 0.87 +/- 0.2 mol% (P < 0.05) and for free Ca2+ from 4.5 +/- 1.0 to 2.8 +/- 0.4 mu M (P < 0.05). Propofol reduced the EC(50) values for phosphatidylserine from 18 +/- 1.9 to 11 +/- 1.2 mol% (P < 0.01), for diacylglycerol from 2.5 +/- 0.3 to 1.2 +/- 0.4 mol% (P < 0.01) and for free Ca2+ from 2.8 +/- 0.7 to 1.9 +/- 0.2 mu M (P < 0.05). The IC50 values for inhibition of PKC activity by the regulatory domain-specific PKC inhibitor sphingosine were increased from 20 +/- 1.5 to 26 +/- 0.6 mu M (P < 0.01) by halothane and from 24 +/- 4.8 to 34 +/- 4.8 mu M (P < 0.05) by propofol. These data suggest that halothane and propofol stimulate brain PKC activity by stabilizing its active conformation through an interaction with its regulatory domain. Given the diverse role of PKC in physiologic regulation, alterations in PKC activity may be a relevant mechanism for general anesthetic effects.