Several lines of evidence suggest that cannabinoid compounds are anticonvulsant. However, the anticonvulsant potential of cannabinoids and, moreover, the role of the endogenous cannabinoid system in regulating seizure activity has not been tested in an in vivo model of epilepsy that is characterized by spontaneous, recurrent seizures. Here, using the rat pilocarpine model of epilepsy, we show that the marijuana extract ⌬ 9 -tetrahydrocannabinol (10 mg/kg) as well as the cannabimimetic, 4, significantly increased both seizure duration and frequency. In some animals, CB 1 receptor antagonism resulted in seizure durations that were protracted to a level consistent with the clinical condition status epilepticus. Furthermore, we determined that during an short-term pilocarpine-induced seizure, levels of the endogenous CB 1 ligand 2-arachidonylglycerol increased significantly within the hippocampal brain region. These data indicate not only anticonvulsant activity of exogenously applied cannabinoids but also suggest that endogenous cannabinoid tone modulates seizure termination and duration through activation of the CB 1 receptor. Furthermore, Western blot and immunohistochemical analyses revealed that CB 1 receptor protein expression was significantly increased throughout the CA regions of epileptic hippocampi. By demonstrating a role for the endogenous cannabinoid system in regulating seizure activity, these studies define a role for the endogenous cannabinoid system in modulating neuroexcitation and suggest that plasticity of the CB 1 receptor occurs with epilepsy.

No abstract

No abstract

Ethanol enhances ␥-aminobutyrate (GABA) signaling in the brain, but its actions are inconsistent at GABA A receptors, especially at low concentrations achieved during social drinking. We postulated that the ⑀ isoform of protein kinase C (PKC⑀) regulates the ethanol sensitivity of GABA A receptors, as mice lacking PKC⑀ show an increased behavioral response to ethanol. Here we developed an ATP analog-sensitive PKC⑀ mutant to selectively inhibit the catalytic activity of PKC⑀. We used this mutant and PKC⑀ ؊/؊ mice to determine that PKC⑀ phosphorylates ␥2 subunits at serine 327 and that reduced phosphorylation of this site enhances the actions of ethanol and benzodiazepines at ␣1␤2␥2 receptors, which is the most abundant GABA A receptor subtype in the brain. Our findings indicate that PKC⑀ phosphorylation of ␥2 regulates the response of GABA A receptors to specific allosteric modulators, and, in particular, PKC⑀ inhibition renders these receptors sensitive to low intoxicating concentrations of ethanol. ␥-Aminobutyrate type A (GABA A )3 receptors mediate the majority of rapid inhibitory neurotransmission in the brain and are an important target for ethanol, the most widely abused drug (1). Ethanol modulation of GABA A receptors was first identified in synaptosomal preparations where intoxicating concentrations (10 -30 mM) enhanced receptor function as measured by 36 Cl uptake assays (2, 3). However, after 30 years of investigation, it is apparent that ethanol enhancement of synaptic GABA A receptors is variable and in some preparations cannot be detected even at anesthetic concentrations (1, 4).GABA A receptors are pentameric protein complexes of subunits from eight classes (␣1-6, ␤1-3, ␥1-3, ␦, ⑀, , , and 1-3) (5). Most receptors are composed of two ␣ subunits and two ␤ subunits that co-assemble with one ␥2 subunit, which anchors these receptors at synapses where they mediate phasic inhibition (6). A minority contain a ␦ subunit instead of ␥2; these receptors are extrasynaptic and mediate tonic inhibition in neurons (7). Because earlier ethanol studies that focused on GABA currents carried by ␥2-containing receptors produced variable results, recent attention has turned to ethanol effects at receptors containing ␦ subunits. Reports from three laboratories found these receptors enhanced by low (Յ30 mM) intoxicating concentrations of ethanol (8 -10). However, two recent studies were unable to demonstrate low dose ethanol sensitivity of ␦-containing GABA A receptors (11, 12), indicating that, like synaptic GABA A receptors, ethanol modulation of extrasynaptic receptors is also variable.The reasons for this high degree of variability are unknown. One possibility is that intracellular signaling pathways may regulate ethanol sensitivity of GABA A receptors, and the activity of such pathways was not controlled for in these studies. This hypothesis is consistent with our findings in mice lacking protein kinase C⑀ (PKC⑀), which show an increased behavioral response to ethanol (13). Ethanol modulation of GABA A receptors is a...

Necrotizing meningoencephalitis (NME) is a disorder of Pug Dogs that appears to have an immune etiology and high heritability based on population studies. The present study was undertaken to identify a genetic basis for the disease. A genome-wide association scan with single tandem repeat (STR) markers showed a single strong association near the dog leukocyte antigen (DLA) complex on CFA12. Fine resolution mapping with 27 STR markers on CFA12 further narrowed association to the region containing DLA-DRB1, -DQA1 and, -DQB1 genes. Sequencing confirmed that affected dogs were more likely to be homozygous for specific alleles at each locus and that these alleles were linked, forming a single high risk haplotype. The strong DLA class II association of NME in Pug Dogs resembles that of human multiple sclerosis (MS). Like MS, NME appears to have an autoimmune basis, involves genetic and nongenetic factors, has a relatively low incidence, is more frequent in females than males, and is associated with a vascularly orientated nonsuppurative inflammation. However, NME of Pug Dogs is more aggressive in disease course than classical human MS, appears to be relatively earlier in onset, and involves necrosis rather than demyelination as the central pathobiologic feature. Thus, Pug Dog encephalitis (PDE) shares clinical features with the less common acute variant forms of MS. Accordingly, NME of Pug Dogs may represent a naturally occurring canine model of certain idiopathic inflammatory disorders of the human central nervous system.

Alcoholism is a progressive disorder that involves the amygdala. Mice lacking protein kinase C epsilon (PKCe) show reduced ethanol consumption, sensitivity and reward. We therefore investigated whether PKCe signaling in the amygdala is involved in ethanol consumption. Local knockdown of PKCe in the amygdala reduced ethanol consumption and preference in a limited-access paradigm. Further, mice that are heterozygous for the PKCe allele consume less ethanol compared with wildtype mice in this paradigm. These mice have a >50% reduction in the abundance of PKCe in the amygdala compared with wild-type mice. We conclude that amygdala PKCe is important for ethanol consumption in mice.

A low level of response to ethanol is associated with increased risk of alcoholism. A major determinant of the level of response is the capacity to develop acute functional tolerance (AFT) to ethanol during a single drinking session. Mice lacking protein kinase C epsilon (PKCe) show increased signs of ethanol intoxication and reduced ethanol self-administration. Here, we report that AFT to the motorimpairing effects of ethanol is reduced in PKCe (À/À) mice when compared with wild-type littermates. In wild-type mice, in vivo ethanol exposure produced AFT that was accompanied by increased phosphorylation of PKCe and resistance of GABA A receptors to ethanol. In contrast, in PKCe (À/À) mice, GABA A receptor sensitivity to ethanol was unaltered by acute in vivo ethanol exposure. Both PKCe (À/À) and PKCe ( + / + ) mice developed robust chronic tolerance to ethanol, but the presence of chronic tolerance did not change ethanol preference drinking. These findings suggest that ethanol activates a PKCe signaling pathway that contributes to GABA A receptor resistance to ethanol and to AFT. AFT can be genetically dissociated from chronic tolerance, which is not regulated by PKCe and does not alter PKCe modulation of ethanol preference.

N-type calcium channels are modulated by acute and chronic ethanol exposure in vitro at concentrations known to affect humans, but it is not known whether N-type channels are important for behavioral responses to ethanol in vivo. Here, we show that in mice lacking functional N-type calcium channels, voluntary ethanol consumption is reduced and place preference is developed only at a low dose of ethanol. The hypnotic effects of ethanol are also substantially diminished, whereas ethanol-induced ataxia is mildly increased. These results demonstrate that N-type calcium channels modulate acute responses to ethanol and are important mediators of ethanol reward and preference.