Loren H. Parsons

Significance: Many CNS disorders result from dysregulation of the mesocorticolimbic dopamine projections arising from the ventral tegmental area. A mechanistic understanding of these dysregulations is critical for developing treatments for these diseases. Nicotine addiction is a global health concern, and results in part from dysregulated mesocorticolimbic dopamine function. Here we present evidence that nicotine-induced 2-arachidonoylglycerol formation in the ventral tegmental area is sensitized following chronic nicotine exposure, and that this results in the loss of GABA-mediated constraint of mesocorticolimbic dopamine activity. Potent and selective diacylglycerol lipase inhibitors are characterized and used in ex vivo electrophysiology and in vivo behavioral analyses to demonstrate that this mechanism plays a prominent role in the dysregulation of ventral tegmental dopamine neurons following nicotine exposure.

Cannabinoid receptors and their endogenous ligands are widely expressed throughout the brain, with a particularly strong presence and influence in neuronal circuits such as the mesocorticolimbic pathways highly implicated in reward and addiction. Cannabinoid 1 receptor (CB1R) signalling influences the motivation for both natural and drug rewards. In comparison to most drugs of abuse, CB1Rs exert only modest influence on psychostimulant intake. Brain endocannabinoid (eCB) levels are increased by most drugs of abuse, although the nature of this effect differs between classes of drugs and across brain regions. The response contingency of drug exposure (volitional versus response-independent) seems to influence brain eCB production, suggesting contributions both of drug-related pharmacological effects and of neural activity engaged by active drug-seeking behaviour. Chronic exposure to drugs of abuse generally results in impaired CB1R function, loss of eCB-mediated synaptic plasticity in addiction-related neural circuits, and negative affective states that can be ameliorated through pharmacologically enhanced eCB tone. The eCB system (ECS) has a strong role in modulating relapse-like behaviour induced by conditioned cues or reward priming, and this is evident for both natural and drug rewards. Recent investigations of CNR1 (which encodes CB1R) and fatty acid amide hydrolase (FAAH) variants generally suggest an association with endophenotypes implicated in addiction susceptibility, including reward sensitivity, impulsivity and negative affect. However, confounding factors, including restricted sample size, ethnicity and polysubstance use, limit interpretational power, and the functional consequences of the variants (causal or linked) are currently unknown.

In humans, long-term marijuana use is associated with impaired impulse control and attentional capacity, though it has been difficult to distinguish pre-existing cognitive deficits from possible consequences of prolonged marijuana exposure. To evaluate the effects of long-term exposure to Δ9-Tetrahydrocannabinol (Δ9-THC), the primary psychoactive constituent in marijuana, on indices of impulse control and attentional capacity using the rat 5-Choice Serial Reaction Time Task (5-CSRTT). Ten 14-day cycles of Δ9-THC dosing and 5-CSRTT testing were employed, each comprised of 5-day Δ9-THC dosing (0.3 or 3 mg/kg b.i.d.) and 5-CSRTT testing during the 9 days of drug abstinence. Subsequent 5-CSRTT testing continued during 5 weeks of protracted abstinence. Dose-dependent increases in motor impulsivity (premature responses) and behavioral disinhibition (perseverative responses) emerged following 5 cycles of Δ9-THC exposure that persisted for the remaining dosing and testing cycles. Δ9-THC-related disruptions in motor impulsivity and behavioral inhibition were most pronounced during cognitively challenging 5-CSRTT sessions incorporating varying novel inter-trial intervals (ITIs), and these disruptions persisted for at least 5 weeks of Δ9-THC abstinence. Δ9-THC-related impairments in attentional capacity (response accuracy) were also evident during variable ITI challenge tests, though these attentional disruptions abated within 3 weeks of Δ9-THC abstinence. These observations demonstrate that long-term intermittent exposure to clinically meaningful Δ9-THC doses induces persistent impairments in impulse control and attentional function. If present in humans, these disruptions may impact academic and professional performance.

Impaired cognitive processing is a hallmark of addiction. In particular, deficits in inhibitory control can propel continued drug use despite adverse consequences. Clinical evidence shows that detoxified alcoholics exhibit poor inhibitory control in the Continuous Performance Task (CPT) and related tests of motor impulsivity. Animal models may provide important insight into the neural mechanisms underlying this consequence of chronic alcohol exposure though pre-clinical investigations of behavioral inhibition during alcohol abstinence are sparse. The present study employed the rat 5 Choice-Continuous Performance Task (5C-CPT), a novel pre-clinical variant of the CPT, to evaluate attentional capacity and impulse control over the course of protracted abstinence from chronic intermittent alcohol consumption. In tests conducted with familiar 5C-CPT conditions EtOH-exposed rats exhibited impaired attentional capacity during the first hours of abstinence and impaired behavioral restraint (increased false alarms) during the first 5d of abstinence that dissipated thereafter. Subsequent tests employing visual distractors that increase the cognitive load of the task revealed significant increases in impulsive action (premature responses) at 3 and 5 weeks of abstinence, and the emergence of impaired behavioral restraint (increased false alarms) at 7 weeks of abstinence. Collectively, these findings demonstrate the emergence of increased impulsive action in alcohol-dependent rats during protracted alcohol abstinence and suggest the 5C-CPT with visual distractors may provide a viable behavioral platform for characterizing the neurobiological substrates underlying impaired behavioral inhibition resulting from chronic intermittent alcohol exposure.

Impulsivity is a risk factor for alcoholism, and long‐term alcohol exposure may further impair impulse control in a manner that propels problematic alcohol use. The present study employed the rat 5‐choice serial reaction time task (5‐CSRTT) to measure behavioral inhibition and attentional capacity during abstinence from repeated 5‐day cycles of alcohol liquid diet consumption. Task performance was not disrupted following the first cycle of alcohol exposure; however, evidence of impaired behavioral inhibition emerged following the third cycle of alcohol exposure. In comparison with controls, alcoholic rats exhibited deficits in inhibitory control during cognitively challenging 5‐CSRTT tests employing variable intertrial interval (varITI). This behavioral disruption was not present during early abstinence (3 days) but was evident by 7 days of abstinence and persisted for at least 34 days. Interestingly, renewed alcohol consumption ameliorated these disruptions in impulse control, although deficient behavioral inhibition re‐emerged during subsequent abstinence. Indices of increased impulsivity were no longer present in tests conducted after 49 days of abstinence. Alcohol‐related impairments in impulse control were not evident in sessions employing highly familiar task parameters regardless of the abstinence period, and control experiments confirmed that performance deficits during the challenge sessions were unlikely to result from alcohol‐related disruption in the adaptation to repeated varITI testing. Together, the current findings demonstrate that chronic intermittent alcohol consumption results in decreased behavioral inhibition in rats that is temporally similar to clinical observations of disrupted impulsive control in abstinent alcoholics performing tasks of behavioral inhibition.

Cannabinoid-1 receptors (CB₁) have an important role in nicotine reward and their function is disrupted by chronic nicotine exposure, suggesting nicotine-induced alterations in endocannabinoid (eCB) signaling. However, the effects of nicotine on brain eCB levels have not been rigorously evaluated. Volitional intake of nicotine produces physiological and behavioral effects distinct from forced drug administration, although the mechanisms underlying these effects are not known. This study compared the effects of volitional nicotine self-administration (SA) and forced nicotine exposure (yoked administration (YA)) on levels of eCBs and related neuroactive lipids in the ventral tegmental area (VTA) and other brain regions. Brain lipid levels were indexed both by in vivo microdialysis in the VTA and lipid extractions from brain tissues. Nicotine SA, but not YA, reduced baseline VTA dialysate oleoylethanolamide (OEA) levels relative to nicotine-naïve controls, and increased anandamide (AEA) release during nicotine intake. In contrast, all nicotine exposure paradigms increased VTA dialysate 2-arachidonoyl glycerol (2-AG) levels. Thus, nicotine differentially modulates brain lipid (2-AG, AEA, and OEA) signaling, and these modulations are influenced by the volitional nature of the drug exposure. Corresponding bulk tissue analysis failed to identify these lipid changes. Nicotine exposure had no effect on fatty acid amide hydrolase activity in the VTA, suggesting that changes in AEA and OEA signaling result from alterations in their nicotine-induced biosynthesis. Both CB₁ (by AEA and 2-AG) and non-CB₁ (by OEA) targets can alter the excitability and activity of the dopaminergic neurons in the VTA. Collectively, these findings implicate disrupted lipid signaling in the motivational effects of nicotine.

The present experiments employed in vivo microdialysis to characterize the effects of commonly used endocannabinoid clearance inhibitors on basal and depolarization-induced alterations in interstitial endocannabinoid levels in the nucleus accumbens of rat brain. Compounds targeting the putative endocannabinoid transporter and hydrolytic enzymes (FAAH and MAGL) were compared. The transporter inhibitor AM404 modestly enhanced depolarization-induced increases in 2-arachidonoyl glycerol (2-AG) levels but did not alter levels of N-arachidonoyl-ethanolamide (anandamide, AEA). The transport inhibitor UCM707 did not alter dialysate levels of either endocannabinoid. The FAAH inhibitors URB597 and PF-3845 robustly increased AEA levels during depolarization without altering 2-AG levels. The MAGL inhibitor URB602 significantly enhanced depolarization-induced increases in 2-AG, but did not alter AEA levels. In contrast, the MAGL inhibitor JZL184 did not alter 2-AG or AEA levels under any condition tested. Finally, the dual FAAH/MAGL inhibitor JZL195 significantly enhanced depolarization-induced increases in both AEA and 2-AG levels. In contrast to the present observations in rats, prior work in mice has demonstrated a robust JZL184-induced enhancement of depolarization-induced increases in dialysate 2-AG. Thus, to further investigate species differences, additional tests with JZL184, PF-3845, and JZL195 were performed in mice. Consistent with prior reports, JZL184 significantly enhanced depolarization-induced increases in 2-AG without altering AEA levels. PF-3845 and JZL195 produced profiles in mouse dialysates comparable to those observed in rats. These findings confirm that interstitial endocannabinoid levels in the brain can be selectively manipulated by endocannabinoid clearance inhibitors. While PF-3845 and JZL195 produce similar effects in both rats and mice, substantial species differences in JZL184 efficacy are evident, which is consistent with previous studies.