Adenosine signaling has been implicated in the pathophysiology of many psychiatric disorders including alcoholism. Striatal adenosine A2A receptors (A2AR) play an essential role in both ethanol drinking and the shift from goal-directed action to habitual behavior. However, direct evidence for a role of striatal A2AR signaling in ethanol drinking and habit development has not been established. Here, we identified that decreased A2AR-mediated CREB activity in the dorsomedial striatum (DMS) enhanced initial behavioral acquisition of goal-directed behaviors and the vulnerability to progress to excessive ethanol drinking during operant conditioning in mice lacking ethanol-sensitive adenosine transporter ENT1 (ENT1−/−). Utilizing mice expressing β-galactosidase (lacZ) under the control of seven-repeated CRE sites in both genotypes (CRE-lacZ/ENT1+/+ mice and CRE-lacZ/ENT1−/− mice) as well as dnCREB (dominant negative form of CREB), we found that reduced CREB activity in the DMS is causally associated with decreased A2AR signaling and increased goal-directed ethanol drinking. Finally, we demonstrated that A2AR antagonist (ZM241385) dampened PKA-activity mediated signaling in the DMS and promoted excessive ethanol drinking in ENT1+/+ mice, but not in ENT1−/− mice. Taken together, our studies indicate that A2AR-mediated CREB signaling in the DMS is a key determinant to enhance the development of goal-directed ethanol drinking in mice.

Background Chronic ethanol abuse is associated with disrupted circadian rhythms and sleep. Ethanol administration impairs circadian clock phase-resetting, suggesting a mode for the disruptive effect of alcohol abuse on circadian timing. Here we extend previous studies to explore the effects of chronic forced ethanol on photic phase-resetting, photic entrainment and daily locomotor activity patterns in C57BL/6J mice. Methods First, microdialysis was used to characterize the circadian patterns of ethanol uptake in the suprachiasmatic (SCN) circadian clock, and correlate this with systemic ethanol levels and episodic drinking of 10% or 15% ethanol. Second, the effects of chronic forced ethanol drinking and withdrawal on photic phase-delays of the circadian activity rhythm were assessed. Third, the effects of chronic ethanol drinking on entrainment to a weak photic zeitgeber (1 min of 25 lux intensity light per day) were assessed. This method was used to minimize any masking actions of light that could obscure ethanol effects on clock entrainment. Results Peak ethanol levels in the SCN and periphery occurred during the dark phase, and coincided with the time when light normally induces phase-delays in mice. These delays were dose-dependently inhibited by chronic ethanol and its withdrawal. Chronic ethanol did not impede re-entrainment to a shifted light cycle, but produced unstable entrainment under the weak photic zeitgeber and disrupted the daily pattern of locomotor activity. Conclusions These results confirm that chronic ethanol consumption and withdrawal markedly impair circadian clock photic phase-resetting. Ethanol also disrupts the temporal structure of nighttime locomotor activity, and photic entrainment. Collectively these results suggest a direct action of ethanol on the SCN clock.

Alcohol withdrawal syndrome (AWS) is a potentially fatal outcome of severe alcohol dependence that presents a significant challenge to treatment. Although AWS is thought to be driven by a hyperglutamatergic brain state, benzodiazepines, which target the GABAergic system, comprise the first line of treatment for AWS. Using a rat model of ethanol withdrawal, we tested whether ceftriaxone, a b-lactam antibiotic known to increase the expression and activity of glutamate uptake transporter EAAT2, reduces the occurrence or severity of ethanol withdrawal manifestations. After a 2-week period of habituation to ethanol in two-bottle choice, alcohol-preferring (P) and Wistar rats received ethanol (4.0 g/kg) every 6 h for 3-5 consecutive days via gavage. Rats were then deprived of ethanol for 48 h during which time they received ceftriaxone (50 or 100 mg/kg, IP) or saline twice a day starting 12 h after the last ethanol administration. Withdrawal manifestations were captured by continuous video recording and coded. The evolution of ethanol withdrawal was markedly different for P rats vs Wistar rats, with withdrawal manifestations occurring 412 h later in P rats than in Wistar rats. Ceftriaxone 100 mg/ kg per injection twice per day (200 mg/kg/day) reduced or abolished all manifestations of ethanol withdrawal in both rat variants and prevented withdrawal-induced escalation of alcohol intake. Finally, ceftriaxone treatment was associated with lasting upregulation of ethanol withdrawal-induced downregulation of EAAT2 in the striatum. Our data support the role of ceftriaxone in alleviating alcohol withdrawal and open a novel pharmacologic avenue that requires clinical evaluation in patients with AWS.

Disrupted circadian rhythmicity is associated with ethanol (EtOH) abuse, yet little is known about how EtOH affects the mammalian circadian clock of the suprachiasmatic nucleus (SCN). Clock timing is regulated by photic and nonphotic inputs to the SCN involving glutamate release from the retinohypothalamic tract and serotonin (5-HT) from the midbrain raphe, respectively. Our recent in vitro studies in the SCN slice revealed that EtOH blocks photic phase-resetting action of glutamate and enhances the nonphotic phase-resetting action of the 5-HT1A,7 agonist, 8-OH-DPAT. To explore the basis of these effects in the whole animal, we used microdialysis to characterize the pharmacokinetics of intraperitoneal injection of EtOH in the hamster SCN extracellular fluid compartment and then studied the effects of such EtOH treatment on photic and serotonergic phase resetting of the circadian locomotor activity rhythm. Peak EtOH levels (approximately 50 mM) from a 2 g/kg injection occurred within 20-40 min with a half-life of approximately 3 h. EtOH treatment dose-dependently attenuated photic phase advances but had no effect on phase delays and, contrary to in vitro findings, markedly attenuated 8-OH-DPAT-induced phase advances. In a complementary experiment using reverse microdialysis to deliver a timed SCN perfusion of EtOH during a phase-advancing light pulse, the phase advances were blocked, similar to systemic EtOH treatment. These results are evidence that acute EtOH significantly affects photic and nonphotic phase-resetting responses critical to circadian clock regulation. Notably, EtOH inhibition of photic signaling is manifest through direct action in the SCN. Such actions could underlie the disruption of circadian rhythmicity associated with alcohol abuse.