Ethanol Activation of Protein Kinase A Regulates GABAA Receptor Subunit Expression in the Cerebral Cortex and Contributes to Ethanol-Induced Hypnosis

Protein kinases are implicated in neuronal cell functions such as modulation of ion channel function, trafficking, and synaptic excitability. Both protein kinase C (PKC) and A (PKA) are involved in regulation of γ-aminobutyric acid type A (GABAA) receptors through phosphorylation. However, the role of PKA in regulating GABAA receptors (GABAA-R) following acute ethanol exposure is not known. The present study investigated the role of PKA in the effects of ethanol on GABAA-R α1 subunit expression in rat cerebral cortical P2 synaptosomal fractions. Additionally, GABA-related behaviors were examined. Rats were administered ethanol (2.0–3.5 g/kg) or saline and PKC, PKA, and GABAA-R α1 subunit levels were measured by western blot analysis. Ethanol (3.5 g/kg) transiently increased GABAA-R α1 subunit expression and PKA RIIβ subunit expression at similar time points whereas PKA RIIα was increased at later time points. In contrast, PKC isoform expression remained unchanged. Notably, lower ethanol doses (2.0 g/kg) had no effect on GABAA-R α1 subunit levels, although PKA type II regulatory subunits RIIα and RIIβ were increased at 10 and 60 min when PKC isozymes are also known to be elevated. To determine if PKA activation was responsible for the ethanol-induced elevation of GABAA-R α1 subunits, the PKA antagonist H89 was administered to rats prior to ethanol exposure. H89 administration prevented ethanol-induced increases in GABAA-R α1 subunit expression. Moreover, increasing PKA activity intracerebroventricularly with Sp-cAMP prior to a hypnotic dose of ethanol increased ethanol-induced loss of righting reflex (LORR) duration. This effect appears to be mediated in part by GABAA-R as increasing PKA activity also increased the duration of muscimol-induced LORR. Overall, these data suggest that PKA mediates ethanol-induced GABAA-R expression and contributes to behavioral effects of ethanol involving GABAA-R

Behavioral and Neurochemical Characterization of the Effects of the Novel Adenosine Antogonist MSX-4

Organisms frequently make effort-related decisions based upon assessments of motivational value and response costs. Dopamine (DA) and DA receptors (DA-Rs), particularly in nucleus accumbens (NAc), are known to regulate such effort-related choice behavior processes. For instance, DA-R antagonists as well as NAc DA depletion can decrease choice behavior. Moreover, previous studies have shown that systemic administration of adenosine A2A antagonists reverses the behavioral effects of DA-R D2 antagonists in rats responding on the concurrent choice task. MSX-3, a selective adenosine A2A antagonist, is a prodrug of MSX-2 with better water solubility. MSX-3 produces a substantial dose-related reversal of the effects of DA-R D2 antagonists on lever pressing and chow intake. MSX-4 is a novel A2A antagonist that is an amino acid ester prodrug of MSX-2, and is thought to have better oral bioavailability than MSX-3. In the present studies, we investigated the effects of MSX-4 on choice behavior using the commonly employed fixed ratio 5 (FR5)/chow feeding procedure. MSX-4 (2.0-8.0 mg/kg IP) produced a dose-related reversal of the effects of 0.08 mg/kg eticlopride on lever pressing and chow intake, with a similar time course as MSX-3. Following oral administration, both MSX-3 and MSX-4 showed a reversal of motivational impairments induced by eticlopride at comparable doses. Furthermore, eticlopride-induced increases in c-Fos expression in NAc were reversed by co-administration of MSX-3 and MSX-4. Overall, these data suggest that both adenosine A2A antagonist pro-drugs induce similar behavioral effects, and similar actions on cellular markers of neural activity in NAc. Additionally, this research further supports the hypothesis that DA and adenosine interact in the regulation of effort-related choice behavior. Further understanding this interaction between DA and adenosine and may have implications for understanding the energy-related motivational dysfunctions, such as psychomotor slowing, fatigue, and apathy, all of which are seen in patients with depression, Parkinson’s disease, and other central nervous system disorders

The role of dopamine D1 receptor transmission in effort-related choice behavior: Effects of D1 agonists

Mesolimbic dopamine (DA), particularly in the nucleus accumbens, is a critical component of the brain circuitry involved in behavioral activation and effort-related processes. Although much is known about the characteristics of DA D2 receptor antagonism on effort-related choice behavior, less is known about the effects of D1 antagonism, and agonist/antagonist interactions. The highly selective D1 antagonist ecopipam was studied for its effects on effort-related choice behavior using the concurrent fixed ratio (FR) 5/chow feeding choice and T-maze barrier choice procedures. In rats tested on the FR5/chow feeding choice task, ecopipam shifted choice behavior, decreasing lever pressing for preferred high carbohydrate pellets but increasing consumption of lab chow. Also, ecopipam decreased selection of the high effort option (i.e., climbing the barrier to obtain a larger reward) in rats tested on the T-maze task, but did not disrupt arm preference or discrimination when no barrier was present. The D1 agonists SKF38393, SKF81297 and A77636 were assessed for their ability to reverse the effects of ecopipam, and in each case the D1 agonist significantly attenuated the effects of ecopipam, typically with an inverted-u shaped dose/response curve. SKF81297 also was able to reverse the effects of the catecholamine depleting agent tetrabenazine on T-maze performance. In summary, the present results implicate DA D1 receptors in the regulation of behavioral activation and effort-related functions, and demonstrate the utility of using tests of effort-related choice behavior for assessing the effects of D1 agonists.This work was supported by grants to J.S. from the National Institute of Mental Health (MH094966) and Pfizer, and to Merce Correa from Fundació Bancaixa/ U. Jaume I. (P1.1B2010-43). Many thanks to Christian Thompson for his assistance

The novel adenosine A_2A antagonist prodrug MSX-4 is effective in animal models related to motivational and motor functions

Adenosine A2A and dopamine D2 receptors interact to regulate diverse aspects of ventral and dorsal striatal functions related to motivational and motor processes, and it has been suggested that adenosine A2A antagonists could be useful for the treatment of depression, parkinsonism and other disorders. The present experiments were performed to characterize the effects of MSX-4, which is an amino acid ester prodrug of the potent and selective adenosine A2A receptor antagonist MSX-2, by assessing its ability to reverse pharmacologically induced motivational and motor impairments. In the first group of studies, MSX-4 reversed the effects of the D2 antagonist eticlopride on a concurrent lever pressing/chow feeding task that is used as a measure of effort-related choice behavior. MSX-4 was less potent after intraperitoneal administration than the comparison compound, MSX-3, though both were equally efficacious. With this task, MSX-4 was orally active in the same dose range as MSX-3. MSX-4 also reversed the locomotor suppression induced by eticlopride in the open field, but did not induce anxiogenic effects as measured by the relative amount of interior activity. Behaviorally active doses of MSX-4 also attenuated the increase in c-Fos and pDARPP-32(Thr34) expression in nucleus accumbens core that was induced by injections of eticlopride. In addition, MSX-4 suppressed the oral tremor induced by the anticholinesterase galantamine, which is consistent with an antiparkinsonian profile. These actions of MSX-4 indicate that this compound could have potential utility as a treatment for parkinsonism, as well as some of the motivational symptoms of depression and other disorders