A major problem with drugs of abuse is the return to drug use after a period of abstinence (relapse). A contributing factor to relapse is the withdrawal-induced anxiety and depression, which stimulates re-administration as a form of self-medication. Supersensitivity of 5-HT2A receptors is associated with withdrawal from several drugs of abuse such as cocaine, MDMA and methamphetamine. Recent evidence also suggests that withdrawal from marijuana-related compounds (cannabinoids) is associated with supersensitivity of 5-HT2A receptors. 5-HT2A receptors in the amygdala and hypothalamic paraventricular nucleus (PVN) are important for the regulation of mood, impulse control and responses to stress. Therefore, withdrawal-induced increases in 5-HT2A receptor function in these limbic regions may be clinically related to the depressed mood and increased anxiety clinically observed during marijuana withdrawal. We hypothesize that desensitization/downregulation of synaptic cannabinoid receptors mediates the cannabinoid withdrawal-induced supersensitivity of 5-HT2A receptors. Aim 1 will determine the minimum number of cannabinoid injection days that will produce supersensitivity of 5-HT2A receptor signaling. These studies will identify potential changes in the parameters and mechanisms of cannabinoid and 5-HT2A receptor function in adult male rats withdrawn from chronic cannabinoid exposure. Aim 2 will determine whether down-regulation of synaptic cannabinoid receptors will produce supersensitivity of 5-HT2A receptor signaling. These studies will examine fundamental mechanisms regulating 5-HT2A receptor signaling in the forebrain of adult rats withdrawn from cannabinoid exposure. We contend that the proposed studies: (1) will provide a unique opportunity to systematically investigate the signaling mechanisms by which cannabinoid receptors regulate the sensitivity of 5-HT2A receptor systems in vivo, and (2), will further our understanding of the neurobiological mechanisms associated with withdrawal from cannabinoids, in order to refine therapy to reduce craving and relapse. PUBLIC HEALTH RELEVANCE Withdrawal from marijuana-related compounds (cannabinoids) induces increases in the activity of the neurotransmitter serotonin in brain areas that could be mechanistically involved in mediating propensity for relapse, and thereby stimulate re- administration as a form of self-medication. The proposed studies will provide a unique opportunity to systematically investigate the signaling mechanisms by which cannabinoid receptors regulate the sensitivity of 5-HT2A receptor systems in vivo. These studies will further our understanding of the neurobiological mechanisms associated with withdrawal from cannabinoids, in order to refine therapy to reduce craving and relapse.