Abstract
GABA is synthesized by two isoforms of glutamate decarboxylase (GAD), GAD65, and GAD67. However, the relative contributions of GAD65-mediated GABA synthesis to the in vivo actions of anesthetics remain unknown. To address this issue, we used mice deficient in the 65-kDa isoform of GAD and tested the hypothesis that partial reduction of GABA content in GAD65-deficient mice [GAD65(-/-)] would contribute to hypnotic and immobilizing actions of the anesthetics. The open field test, loss of righting reflex (LORR), loss of tail-pinch withdrawal response (LTWR), and locomotor activity were compared between wild-type (WT) mice and GAD65(-/-) mice. Effects of general anesthetics on both phasic and tonic GABAergic currents were examined using the patch-clamp method in frontal cortex pyramidal neurons in brain slices. The duration of propofol (100 mg/kg i.p.)-induced LORR and the duration of propofol (150 mg/kg i.p.)-induced LTWR in GAD65(-/-) mice were significantly reduced compared with WT mice. In contrast, no difference was seen for ketamine. Preinjection of the GABA transporter 1 inhibitor, NO-711 (C21H22N2O3 · HCl) (0.75 mg/kg i.p.), reinstated diminished actions of propofol in GAD65(-/-) mice. Cortical pyramidal neurons in GAD65(-/-) mice had smaller tonic conductances, and propofol-induced enhancement of tonic inhibition was smaller than in WT mice, suggesting that genotype differences in GAD65-mediated GABAergic inhibitory tone may be, at least in part, a cellular basis underlying behavioral differences. In conclusion, GAD65(-/-) mice show a diminished response to propofol, but not ketamine, indicating that GAD65-mediated GABA synthesis plays an important role in hypnotic and immobilizing actions of propofol.
Footnotes
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This work was supported by the Ministry of Education, Culture, Sports, Science and Technology of Japan [Grants-in-Aid for Scientific Research 17390425, 17659483, 20390412, 187866, 19791059, 18300102, 19040002]; and a grant-in-aid from the Japan Medical Association (Tokyo, Japan).
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Parts of this work were previously presented at the following conference: Nishikawa K, Kubo K, Yamada M, Ishizeki J, and Saito S (2007) Altered responses to propofol, but not ketamine, in mice lacking glutamate decarboxylase 65. 2007 Annual Meeting of the American Society of Anesthesiologists; 2007 Oct 13-17; San Francisco, CA. American Society of Anesthesiologists, Park Ridge, IL.
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doi:10.1124/jpet.109.151456.
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ABBREVIATIONS: GAD, glutamate decarboxylase; WT, wild type; LORR, loss of righting reflex; LTWR, loss of tail-pinch withdrawal response; LHWR, loss of hind-limb withdrawal reflex; ACSF, artificial cerebrospinal fluid; fEPSP, field excitatory postsynaptic potential; PS, population spike; ANOVA, analysis of variance; DMSO, dimethyl sulfoxide; GAT, GABA transporter; SR95531, 2-(3-carboxypropyl)-3-amino-6-(4 methoxyphenyl)pyridazinium bromide; BIC, bicuculline; mIPSC, miniature inhibitory postsynaptic current; PIC, picrotoxin; NO-711, C21H22N2O3 · HCl.
- Received January 25, 2009.
- Accepted February 19, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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