Membrane Cholesterol Depletion Alters Striatal Dopamine Homeostasis and Dopamine-Associated Behavior

Abstract ID 87320 Poster Board 296 Background: Chronic cocaine use dysregulates dopamine transporter (DAT) function and synaptic dopamine (DA) transmission, leading to enhanced drug-taking behaviors. While the mechanisms underlying cocaine-induced DAT dysfunction remain elusive, emerging evidence suggests a link between DAT function and brain cholesterol content. Whether cocaine dysregulates DAT function by disrupting brain cholesterol content has yet to be explored. Therefore, this project investigated: 1) if cocaine self-administration (SA) altered brain cholesterol homeostasis, 2) if decreasing cholesterol ex vivo directly altered DAT function, and 3) if decreasing brain cholesterol in vivo altered dopamine-related behaviors. Method: Cocaine SA: Male Sprague-Dawley rats were allowed to self-administer 40 injections of cocaine (1.5 mg/kg/infusion, 6 h/d) or vehicle for 5 d. Striata were dissected 18 h after the last cocaine infusion. Striatal [³H]DA Uptake and Cocaine Inhibition: [³H]DA uptake was performed on striatal synaptosomes from cocaine SA and control animals to measure DAT function. To determine the potency of cocaine to inhibit DA uptake, [³H]DA uptake was performed in the presence of cocaine. Ex-Vivo Fast-Scan Cyclic Voltammetry (FSCV): Recording and stimulating electrodes were placed in the nucleus accumbens core, and a single, rectangular electrical pulse evoked dopamine release and reuptake. Once the evoked DA response was stable, methyl-β-cyclodextrin (MβCD, 0.3, 3, or 10 mM), a membrane cholesterol chelator, was perfused over brain slices for 1 h, followed by washout. Kinetics were determined using Michaelis-Menten-based modeling. Simvastatin Osmotic Minipump: To assess cholesterol depletion in vivo, male Sprague-Dawley rats were implanted with an osmotic minipump delivering the cholesterol synthesis inhibitor simvastatin (5 mg/kg/d) or vehicle for 14 d. Rats were then subjected to behavioral assessments of dopamine function, including novelty- and cocaine-induced locomotor activity. Results: First, we found that cocaine SA decreased [³H]DA uptake and the ability of cocaine to inhibit DAT function. In parallel, cocaine SA reduced striatal membrane cholesterol by accelerating cholesterol turnover. Second, we found that application of 3 mM and 10 mM, but not 0.3 mM MβCD to brain slices: 1) decreased evoked DA release, 2) reduced the maximal rate of DA uptake, and 3) reduced the ability of cocaine to inhibit DA release. Finally, brain cholesterol depletion via simvastatin administration significantly altered dopamine-related behaviors, suggesting the impact of disrupted brain cholesterol on dopamine transmission. Conclusions: Our data demonstrate that membrane cholesterol depletion disrupts striatal dopamine homeostasis and the interactions between DAT and cocaine. This study underscores the need for tools to monitor brain cholesterol metabolism and highlights brain cholesterol as a potential new pharmacotherapy for psychostimulant use disorder. This work was supported by the National Institute on Drug Abuse R21DA056857 and R01DA042862. Anna Neel is supported by the National Institute on Alcohol Abuse and Alcoholism training grant T32AA007565 and the National Science Foundation Graduate Research Fellowship Program grant 2102417.