Amphetamine self‐administration alters the cholesterol content and G protein membrane compartmentalization in the rat striatum.
Authors: Hinshaw Tyler , Yeager Matthew , Luessen Deborah , Sun Haiguo , Chen Rong 
Published: November 2023

IntroductionChronic amphetamine use causes profound adaptations in the brain including desensitization and sensitization of many GPCRs and altered signaling of G proteins coupled to these receptors. These changes are associated with vulnerability to relapse. However, the mechanisms underlying changes in receptor‐mediated G protein signaling remain elusive.One possible mechanism is an alteration in the compartmentalization of G proteins in the plasma membrane. Lipid rafts are the subcompartment of the plasma membranes characterized by high cholesterol content. They act as microdomains to accommodate cellular signaling and protein trafficking. Changes in the lipid composition of the plasma membrane would alter protein localization and associated signaling. Little is known about the effects of psychostimulant treatments on G protein distribution in lipid raft and nonraft domains of striatal cell membranes.The purpose of this study was to use an animal model of amphetamine self‐administration to investigate the parallel changes in the brain cholesterol content and G protein membrane compartmentalization. This study may shed light on the contribution of membrane cholesterol content to amphetamine‐induced alteration in G protein signaling and receptor responses to stimuli.MethodsMale Sprague Dawley rats were anesthetized, implanted with catheters, and allowed to self‐administer amphetamine (0.189 mg/kg/infusion) on a fixed ratio one schedule. Animals were allowed to self‐administer up to 40 injections in 6 hrs per day over 5 or 14 days. Rats were sacrificed and the striata were dissected. Striatal tissues were fractionated using a sucrose density gradient and centrifugation to isolate lipid raft and non‐lipid raft fractions. Fractions were subjected to immunoblotting to determine G protein localization. Amplex Red Hydrogen Peroxide/Peroxidase assay kit was used to measure the striatal cholesterol levels.ResultsOur data showed both 5 and 14 days of amphetamine self‐administration significantly reduced the total cholesterol levels in the striatum. Interestingly, we found that amphetamine self‐administration differentially regulated the membrane compartmentalization of Gα subtypes. For Gαo and Gαs, there was a notable translocation from nonraft to raft fractions in both 5 and 14 days amphetamine self‐administering rats. For Gαi2 and Gαi3, there was no change in membrane compartmentalization following 5 days of amphetamine self‐administration; however, 14 days of amphetamine exposure caused these G‐proteins to translocate from raft to nonraft fractions. No significant membrane translocation of Gαq was observed. This study provides first time evidence that amphetamine self‐administration selectively alters the membrane compartmentalization of Gα subunits, which may be associated with changes in receptor function and addiction‐like behavior. Future research on the contribution of the membrane cholesterol content to receptor function and addiction behavior is warranted.Support or Funding InformationThis project is supported by NIH DA006634.