doi: 10.1523/JNEUROSCI.22-22-09905.2002.
Conditional rescue of protein kinase C epsilon regulates ethanol preference and hypnotic sensitivity in adult mice
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- PMID:12427847
- PMCID: PMC6757853
- DOI: 10.1523/JNEUROSCI.22-22-09905.2002
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Conditional rescue of protein kinase C epsilon regulates ethanol preference and hypnotic sensitivity in adult mice
Doo-Sup Choi et al. J Neurosci..
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Abstract
Conventional gene targeting is a powerful tool to study the influence of specific genes on behavior. However, conclusions relevant for adult animals are limited by consequences of gene loss during development. Mice lacking protein kinase C epsilon (PKCepsilon) consume less alcohol and show greater acute sensitivity to alcohol than do wild-type mice. There are no selective inhibitors of PKCepsilon that can be administered systemically and cross the blood-brain barrier to test whether these phenotypes result from loss of PKCepsilon during development or in adulthood. Here we used conditional expression of PKCepsilon in the basal forebrain, amygdala, and cerebellum to rescue wild-type responses to alcohol in adult PKCepsilon(-/-) mice. Subsequent suppression of transgenic PKCepsilon restored PKCepsilon(-/-) behaviors. These findings establish that PKCepsilon signaling in the adult brain regulates alcohol consumption and sensitivity. If this extends to humans, then PKCepsilon inhibitors might prove useful as novel therapeutics for alcoholism.
Figures
Regulated expression of PKCε.A, Schematic drawing of thePrnp-tTA transgene of line TgPrnp-tTA/F959 and thePtet-PKCεtransgene of line TgPtet-PKCε/T9.B, Western blot of brain homogenate (40 μg perlane) from WT mice and PKCε-deficient mice carrying only thePtet-PKCε transgene (SKO) or bothPtet-PKCε andPrnp-tTA transgenes (DKO), before (−) or during (+) treatment with Dox.C, Immunofluorescence detection of the neuronal marker NeuN (red) and of PKCε (green) in the nucleus accumbens (NAc) and amygdala (Amy).Whitearrows, Several PKCε-expressing neurons with NeuN immunoreactive nuclei surrounded by PKCε immunoreactivity in the cell soma. Scale bar, 50 μm.D, Immunohistochemical staining of brain tissue from WT and DKO mice fed normal chow (−Dox) or chow containing doxycycline (+Dox) for 2 weeks. Sections shown are from the nucleus accumbens (NAc), ventral pallidum (VP), amygdala (Amy), ventral tegmental area (VTA), bed nucleus of the stria terminalis (BST), caudate putamen (CPu), hippocampus (Hp), cingulate cortex (Cg), lateral hypothalamic area (LH), and cerebellum (Cbl). Labeled subregions are the nucleus accumbens core (AcC), nucleus accumbens shell (AcS), islands of Calleja (ICj), lateral division of the central amygdaloid nucleus (CeL), anterior portion of the basolateral amygdaloid nucleus (BL), caudate putamen (CPu), anterior commissure (ac), corpus callosum (cc), medial globus pallidus (GPm), optic tract (ot), and the molecular layer (ML), Purkinje cell layer (PCL), and granule cell layer (GCL) of the cerebellum.Asterisksindicate the locations of the VTA, BST, and LH. Scale bar: Cbl, 50 μm; Hp, LH, 200 μm; other sections, 100 μm. Images are from representative experiments, each repeated two times using different mice, with similar results.
Doxycycline-regulated ethanol consumption and preference (mean ± SEM values) examined in 10 WT (●), 13 SKO (○), and 15 DKO (■) mice before (pre-Dox,A,D), during (Dox,B,E), and after (post-Dox,C,F) treatment with doxycycline. All mice were experimentally naive at the beginning of the experiment.A–C, Doxycycline altered ethanol consumption in a genotype and ethanol concentration-dependent manner [F(8,140) (genotype × doxycycline treatment × ethanol concentration) = 3.29;p = 0.0018].D–F, Doxycycline also altered ethanol preference (100 × milliliters of ethanol per total milliliters consumed) in a genotype and ethanol concentration-dependent manner [F(8,140)(genotype × doxycycline treatment × ethanol concentration) = 2.27;p = 0.026]. Shown are the results of Tukey's tests for the three-way interaction between genotype, doxycycline treatment, and ethanol concentration (*p < 0.05 relative to wild-type and DKO mice in trials 1 and 3;§p < 0.05 relative to wild-type mice only in trial 3;†p < 0.05 relative to SKO and DKO mice in trial 2;#p < 0.05 relative to WT mice in trial 1). Results for the two-way interaction between genotype and doxycycline treatment are given in Results.
Preference for saccharin and quinine and total fluid intake. Data inA andB are mean ± SEM values from 10 WT, 13 SKO, and 15 DKO mice. Shown are preference ratios for solutions containing 0.03 and 0.06% saccharin (A) and 0.015 and 0.030 mm quinine (B) during 2 d of access to each solution before (white bars) and during (black bars) treatment with doxycycline.
Drug-induced LORR. Duration of drug-induced LORR was examined before (white bars) or during (black bars) treatment with doxycycline. All data are mean ± SEM values.A, Ethanol-induced LORR in nine WT, 11 SKO, and 11 DKO mice not exposed to doxycycline and in 10 WT, 12 SKO, and 12 DKO mice treated with doxycycline for 2 weeks. Doxycycline altered LORR duration in a genotype-specific manner [F(2,59) (genotype) = 9.05,p = 0.0004;F(1,59)(doxycycline treatment), NS;F(2,59)(genotype × doxycycline treatment) = 3.56,p = 0.034].B, Plasma ethanol concentrations measured using the ethanol diagnostic kit 332-UV (Sigma, St Louis, MO) did not differ among three WT, five SKO, and five DKO mice after acute administration of ethanol (3.6 gm/kg, i.p.).C, Pentobarbital-induced LORR examined in seven WT, seven SKO, and 11 DKO mice not exposed to doxycycline and in seven WT, nine SKO, and nine DKO mice treated with doxycycline for 2 weeks. Doxycycline altered LORR duration in a genotype-specific manner [F(2,45) (genotype) = 8.66,p < 0.001;F(1,45)(doxycycline treatment) = 7.08,p = 0.011;F(2,45) (genotype × doxycycline treatment) = 3.56,p = 0.034].D, Ketamine-induced LORR duration examined in seven WT, seven SKO, and eight DKO mice not exposed to doxycycline and in seven WT, eight SKO, and eight DKO mice treated with doxycycline was similar in all genotypes before and during doxycycline treatment. Data are mean ± SEM values. *p < 0.05 compared with WT in the absence of doxycycline and DKO in the absence of doxycycline;†p < 0.05 compared with DKO plus doxycycline;§p < 0.05 compared with WT plus doxycycline (Newman–Keuls tests).
References
- Browman KE, Crabbe JC. Quantitative trait loci affecting ethanol sensitivity in BXD recombinant inbred mice. Alcohol Clin Exp Res. 2000;24:17–23. - PubMed
- Chen X-N, Knauf JA, Gonsky R, Wang M, Lai EH, Chissoe S, Fagin JA, Korenberg JR. From amplification to gene in thyroid cancer: a high-resolution mapped bacterial-artificial-chromosome resource for cancer chromosome aberrations guides gene discovery after comparative genome hybridization. Am J Hum Genet. 1998;63:625–637. - PMC - PubMed
- Coe IR, Yao L, Diamond I, Gordon AS. The role of protein kinase C in cellular tolerance to ethanol. J Biol Chem. 1996;271:29468–29472. - PubMed
- Crabbe JC, Phillips TJ, Feller DJ, Hen R, Wenger CD, Lessov CN, Schafer GL. Elevated alcohol consumption in null mutant mice lacking 5-HT1B serotonin receptors. Nat Genet. 1996;14:98–101. - PubMed
- Crabbe JC, Phillips TJ, Buck KJ, Cunningham CL, Belknap JK. Identifying genes for alcohol and drug sensitivity: recent progress and future directions. Trends Neurosci. 1999;22:173–179. - PubMed
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