European Journal of Pharmacology, 217 (1992) 215-216 © 1992 Elsevier Science Publishers B.V. All rights reserved 0014-2999/92/$05.00
215
EJP 0358R
Rapid communication
Isoflurane anesthesia is stereoselective Bradford Harris a, Eric Moody a,b and Phil Skolnick a Laborato~ of Neuroscience, NIDDK / NIH, Bethesda, MD 20892, USA and b Department of Anesthesiology and Critical Care Medicine, Johns Hopkins UniL,ersity, Baltimore, MD 21205, USA Received 25 May 1992, accepted 1 Juni 1992
The anesthetic effects of the (+) and ( - ) isomers of isoflurane were determined in mice. Like the clinically important racemate, both isomers produced dose-dependent increases in anesthetic sleep time. A statistically significant (P < 0.005, ANOVA) difference in the potencies of these isomers ((+)> (-)) was observed. To our knowledge, this is the first demonstration of a stereospecific action of a volatile anesthetic in vivo. Isoflurane; Anesthesia; Stereoisomers
The mechanism(s) responsible for the anesthetic actions of inhalational agents remains controversial, with both lipids and proteins representing putative targets (Miller, 1985; Franks and Lieb, 1991, Moody et al., 1991). The ability of pharmacological agents to produce a stereospecific action is strong evidence of a receptor-mediated event (Burt, 1985). While several clinically used inhalation anesthetics possess an asymmetric center, the lack of sufficient quantities of their stereoisomers has precluded an examination of stereoselectivity. The recent availability (Huang et al., 1992) of the stereoisomers of isoflurane made it feasible to determine if there were differences in the anesthetic potencies of these compounds in vivo. The stereoisomers of isoflurane were prepared and donated by Anaquest (Murray Hill, N J). Adult male NIH/Swiss mice (25-32 g) were housed under N I H / A A A L A C conditions. Mice were injected i.p. with 0.2 ml of a freshly prepared solution containing 10% DMSO (Sigma), 30% Alkamuls EL-620 (RhonePoulenc, Cranbury, NJ), isoflurane (5.4-10.8 mm/kg) and saline to final volume. Once asleep, the mice were placed supine on plexiglas troughs in a 34°C chamber. Sleep times were recorded as the difference between injection time and the return of the righting reflex (three times within 1 min). Two-way ANOVA (Genstat, Infallible Software, Research Triangle Park, NC) was used to evaluate statistical significance. Due to the
Correspondence to: B. Harris, NIDDK/NIH, Building 8, Room 111, 9000 Rockville Pike, Bethesda, MD 20892, USA. Tel. 1.301.496 0606, fax 1.301.402 2872.
limited quantities of isomers available, only three doses of 9-16 mice/group were used. The doses of isoflurane isomers selected were based on pilot experiments with racemic isoflurane (data not shown). Like the racemic mixture, both isomers of isoflurane produced a dose-dependent increase in sleep time in vivo (fig. 1). (+)-Isoflurane was significantly more potent than (-)-isoflurane at doses between 5.4 and 10.8 mmol/kg. Thus, two-way ANOVA revealed a highly significant effect for both dose dependence (P ( - )) at potassium channels in the right parietal ganglion of the mollusc Lynmaea stagnalis (Franks and Lieb, 1991). Moreover, other studies using the isomers of halothane failed to demonstrate stereoselectivity in either optical rotatory dispersion or electron spin resonance in lipid environments (Laasbery and Hedley-Whyte, 1971; Kendig et al., 1973). The in vivo studies reported here demonstrate that inhalation anesthetics are likely to exert their effects via a receptor-mediated process since both isomers should exhibit similar properties in lipid environments. The differences in anesthetic potency of (+)- and (-)-isoflurane, while highly statistically significant, were modest. This may be attributed in part to both the mode of administration and paradigm employed that were mandated by the limited availability of these
216
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including GABA A and nicotinic acetylcholine receptors (Moody et al., 1991; Miller 1985). The availability of greater quantities of these isomers witll permit the further examination of this phenomena using traditional administration techniques in other mammals.
30 c
~
25
.~
20
Acknowledgements
15
We thank Dr. Huang and Mr. Vernice of Anaquest for providing the isomers of isoflurane.
I-r~
oo
10 References 5 5.4 8.1 10,8 Isoflurane, mmoles/kg Fig. 1. Effect of isoflurane injection on sleep time in mice. Mice were injected with 0.2 ml of isoflurane suspension i.p. as described in the text. Values represent means +_S.E.M. of n = 9-16. Two-way ANOVA revealed a significant dose effect ( P
215
EJP 0358R
Rapid communication
Isoflurane anesthesia is stereoselective Bradford Harris a, Eric Moody a,b and Phil Skolnick a Laborato~ of Neuroscience, NIDDK / NIH, Bethesda, MD 20892, USA and b Department of Anesthesiology and Critical Care Medicine, Johns Hopkins UniL,ersity, Baltimore, MD 21205, USA Received 25 May 1992, accepted 1 Juni 1992
The anesthetic effects of the (+) and ( - ) isomers of isoflurane were determined in mice. Like the clinically important racemate, both isomers produced dose-dependent increases in anesthetic sleep time. A statistically significant (P < 0.005, ANOVA) difference in the potencies of these isomers ((+)> (-)) was observed. To our knowledge, this is the first demonstration of a stereospecific action of a volatile anesthetic in vivo. Isoflurane; Anesthesia; Stereoisomers
The mechanism(s) responsible for the anesthetic actions of inhalational agents remains controversial, with both lipids and proteins representing putative targets (Miller, 1985; Franks and Lieb, 1991, Moody et al., 1991). The ability of pharmacological agents to produce a stereospecific action is strong evidence of a receptor-mediated event (Burt, 1985). While several clinically used inhalation anesthetics possess an asymmetric center, the lack of sufficient quantities of their stereoisomers has precluded an examination of stereoselectivity. The recent availability (Huang et al., 1992) of the stereoisomers of isoflurane made it feasible to determine if there were differences in the anesthetic potencies of these compounds in vivo. The stereoisomers of isoflurane were prepared and donated by Anaquest (Murray Hill, N J). Adult male NIH/Swiss mice (25-32 g) were housed under N I H / A A A L A C conditions. Mice were injected i.p. with 0.2 ml of a freshly prepared solution containing 10% DMSO (Sigma), 30% Alkamuls EL-620 (RhonePoulenc, Cranbury, NJ), isoflurane (5.4-10.8 mm/kg) and saline to final volume. Once asleep, the mice were placed supine on plexiglas troughs in a 34°C chamber. Sleep times were recorded as the difference between injection time and the return of the righting reflex (three times within 1 min). Two-way ANOVA (Genstat, Infallible Software, Research Triangle Park, NC) was used to evaluate statistical significance. Due to the
Correspondence to: B. Harris, NIDDK/NIH, Building 8, Room 111, 9000 Rockville Pike, Bethesda, MD 20892, USA. Tel. 1.301.496 0606, fax 1.301.402 2872.
limited quantities of isomers available, only three doses of 9-16 mice/group were used. The doses of isoflurane isomers selected were based on pilot experiments with racemic isoflurane (data not shown). Like the racemic mixture, both isomers of isoflurane produced a dose-dependent increase in sleep time in vivo (fig. 1). (+)-Isoflurane was significantly more potent than (-)-isoflurane at doses between 5.4 and 10.8 mmol/kg. Thus, two-way ANOVA revealed a highly significant effect for both dose dependence (P ( - )) at potassium channels in the right parietal ganglion of the mollusc Lynmaea stagnalis (Franks and Lieb, 1991). Moreover, other studies using the isomers of halothane failed to demonstrate stereoselectivity in either optical rotatory dispersion or electron spin resonance in lipid environments (Laasbery and Hedley-Whyte, 1971; Kendig et al., 1973). The in vivo studies reported here demonstrate that inhalation anesthetics are likely to exert their effects via a receptor-mediated process since both isomers should exhibit similar properties in lipid environments. The differences in anesthetic potency of (+)- and (-)-isoflurane, while highly statistically significant, were modest. This may be attributed in part to both the mode of administration and paradigm employed that were mandated by the limited availability of these
216
35
including GABA A and nicotinic acetylcholine receptors (Moody et al., 1991; Miller 1985). The availability of greater quantities of these isomers witll permit the further examination of this phenomena using traditional administration techniques in other mammals.
30 c
~
25
.~
20
Acknowledgements
15
We thank Dr. Huang and Mr. Vernice of Anaquest for providing the isomers of isoflurane.
I-r~
oo
10 References 5 5.4 8.1 10,8 Isoflurane, mmoles/kg Fig. 1. Effect of isoflurane injection on sleep time in mice. Mice were injected with 0.2 ml of isoflurane suspension i.p. as described in the text. Values represent means +_S.E.M. of n = 9-16. Two-way ANOVA revealed a significant dose effect ( P
