Antagonism of the Ethanol-Like Discriminative Stimulus Effects of Ethanol, Pentobarbital, and Midazolam in Cynomolgus Monkeys Reveals Involvement of Specific GABA_AReceptor Subtypes

Abstract

The ␥-aminobutyric acid (GABA) A receptors mediating the discriminative stimulus effects of ethanol were studied by comparing the potency of ethyl-8-azido-5,6-dihydro-5-methyl-6-oxo-4H-imidazol(1,5-a)benzodiazepine-3-carboxylate (Ro15-4513) and ethyl 8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazol(1,5-a)-benzodiazepine-3-carboxylate (flumazenil, Ro15-1788) to antagonize ethanol, pentobarbital (PB), and midazolam substitution for ethanol. Ro15-4513 has high affinity for receptors containing ␣ 4/6 and ␣ 5 subunits and lower affinity for ␣ 1 , ␣ 2 , and ␣ 3 subunits. Flumazenil is nonselective for GABA A receptors containing ␣ 1 , ␣ 2 , ␣ 3 , and ␣ 5 subunits and has low affinity for ␣ 4/6 -containing receptors. Male (n ϭ 9) and female (n ϭ 8) cynomolgus monkeys (Macaca fascicularis) were trained to discriminate ethanol (1.0 or 2.0 g/kg i.g., 30-min pretreatment) from water. Ethanol, PB, and midazolam dose-dependently substituted for ethanol (80% ethanol-appropriate responding). Ro15-4513 (0.003-0.56 mg/kg i.m., 5-min pretreatment) shifted the ethanol, PB, and midazolam doseresponse functions rightward in a vast majority of monkeys tested (15/15, 16/17, and 11/12, respectively). In contrast, flumazenil (0.30 -10.0 mg/kg i.m., 5-min pretreatment) shifted the ethanol, PB, and midazolam dose-response functions rightward in 9 of 16, 12 of 16, and 7 of 9 monkeys tested, respectively. In the monkeys showing antagonism with both Ro15-4513 and flumazenil, ethanol and PB substitution were antagonized more potently by Ro15-4513 than by flumazenil, whereas midazolam substitution was antagonized with similar potency. There were no sex or training dose differences, with the exception that flumazenil failed to antagonize ethanol substitution in males trained to discriminate 2.0 g/kg ethanol. GABA A receptors with high affinity for Ro15-4513 (i.e., containing ␣ 4/6 and ␣ 5 subunits) may be particularly important mediators of the multiple discriminative stimulus effects of ethanol through GABA A receptor systems.

Effects of ethanol on cocaine self-administration in monkeys responding under a second-order schedule of reinforcement

“…This pretreatment time was selected on the basis of the time course for the discriminative stimulus effects of i.g. EtOH in male cynomolgus monkeys (Helms et al, 2009). EtOH test sessions were typically conducted on Tuesdays and Fridays and vehicle (water) pretreatments were typically on Thursdays.…”

Cross-Species Translational Findings in the Discriminative Stimulus Effects of Ethanol

“…Specifically, studies in monkeys have examined subunit-selective ligands and antagonists at the GABA A receptor [75, 79–81], as well as neuroactive steroid activity [74, 78, 82, 83]. Additionally, cross-generalization analysis was possible by studies that trained ethanol-like GABA A ligands and examined ethanol in substitution tests [79, 84–86].…”

“…In contrast to rodents, however, GABA A modulators produce full substitution at low and high training doses (1.0–2.0 g/kg), rather than just predominantly at lower doses. Converging evidence from multiple studies suggests that α5 subunit-containing receptors are particularly important in ethanol’s discriminative stimulus effects [75, 80, 81], as well as some contribution of the α1 and α2/3 subunits. Alpha-5 and alpha-1 selective agonists substitute for ethanol, but only inverse agonists selective for α5 (L-655,708) and α5 + α4/6 (Ro-154513) are able to antagonize ethanol’s discriminative stimulus effects [75, 87].…”

“…Converging evidence from multiple studies suggests that α5 subunit-containing receptors are particularly important in ethanol’s discriminative stimulus effects [75, 80, 81], as well as some contribution of the α1 and α2/3 subunits. Alpha-5 and alpha-1 selective agonists substitute for ethanol, but only inverse agonists selective for α5 (L-655,708) and α5 + α4/6 (Ro-154513) are able to antagonize ethanol’s discriminative stimulus effects [75, 87]. Ro-154513 is also able to antagonize the substitution of benzodiazepines and barbiturates for ethanol, suggesting a shared action at the GABA A subunit level [76].…”

Activation of Group II Metabotropic Glutamate Receptors Inhibits the Discriminative Stimulus Effects of Alcohol via Selective Activity Within the Amygdala

“…As such, activation of mGlu2/3 receptors decreases the synaptic availability of glutamate, allowing for 'refinement' of glutamatergic neurotransmission (Schoepp, 2001;Pinheiro and Mulle, 2008). Given the functional role of mGlu2/3 receptors in modulating glutamate release and that the discriminative stimulus effects of alcohol are generally characterized by processes that reduce/inhibit glutamatergic neurotransmission (Kostowski and Bienkowski, 1999), such that N-methyl-Daspartic acid (NMDA) antagonists and g-aminobutyric acid type A (GABA A )-positive modulators produce alcohol-like discriminative stimulus effects (Jarbe and McMillan, 1983;Schechter et al, 1993;Ator et al, 1993;Bienkowski et al, 1997;Hundt et al, 1998;Grant et al, 2000;Shelton and Grant, 2002;Vivian et al, 2002;Helms et al, 2009), we hypothesized that mGlu2/3 receptors may have a modulatory role in the expression of the discriminative stimulus effects of alcohol. Further support for this hypothesis comes from studies showing that mGlu2/3 receptors are highly expressed in limbic brain regions (Petralia et al, 1996;Ohishi et al, 1998;Ferraguti and Shigemoto, 2006) known to modulate the discriminative stimulus effects of alcohol, such as the nucleus accumbens and the amygdala (Hodge and Aiken, 1996;Hodge and Cox, 1998;Hodge et al, 2001;Besheer et al, 2003).…”