Alcohol during the adolescent period produces permanent maladaptive changes in the brain. Adolescent drinkers report greater prevalence of pain disorders in adulthood, and functional magnetic resonance imaging has shown greater amygdala activation compared to non-drinkers. The amygdala, and more specifically the central nucleus of the amygdala (CeA) is a key brain region in pain processing by glutamate transmission from pain-related brain regions. The CeA is sensitive to alcohol, however there are divergent results depending on timing of exposure. In adult rodent models, chronic alcohol increases CeA activity by upregulating N-methyl-D-Aspartate (NMDAR) subunits and AMPAR mediated excitatory transmission leading to increased pain processing. These effects are transient, as changes in NMDAR subunit expression and glutamate transmission normalize within 1-2 weeks following exposure. In an adolescent alcohol exposure model, increases in NMDAR subunit and glutamate signaling lasts into adulthood in the extended amygdala, suggesting adolescent alcohol effects are more persistent. The melanocortin system and neural specific melanocortin 4 receptor (MC4R) are highly expressed in the CeA and regulate glutamatergic transmission. MC4R increases AMPAR post-synaptically, modulates NMDAR function, and increases glutamate signaling in other pain-related regions. Melanocortin signaling in the amygdala modulates pain-related behaviors as intra-CeA MC4R antagonism reverses alcohol-induced hyperalgesia in adult rats. Adult alcohol exposure decreases MC4R expression, but adolescent alcohol exposure increases MC4R expression in adulthood. How the melanocortin system modulates glutamate transmission in the CeA, the longitudinal effects of adolescent alcohol on CeA glutamate and melanocortin systems, and how the melanocortin system modulates pain processing longitudinally have yet to be addressed. Using a rat model, we will test the effect of adolescent alcohol exposure on melanocortin and glutamatergic signaling in CeA, the effect of chronic MC4R antagonism on cellular effects during this period, and we will test MC4R antagonism during adulthood on hyperalgesia induced by an adolescent intermittent ethanol (AIE) vapor model. Male and female Wistar rats will receive alcohol vapor from postnatal day (PND) 28 to PND 56, with one cohort receiving chronic MC4R antagonism via intracerebroventricular minipumps during this period. We will use Western blots to measure AMPAR and NMDAR subunits, and melanocortin system protein expression acutely after AIE (PND 56) and during adulthood (PND 86). We will use slice electrophysiology to measure glutamatergic transmission in the CeA during adolescence and adulthood, and its modulation by MC4R drugs. Finally, we will test the effects of intra-CeA and intranasal MC4R antagonism on nociception in adulthood. Our hypothesis is that AIE produces lasting increases in melanocortin and glutamatergic signaling in the CeA that mediates AIE- induced hyperalgesia, that chronic MC4R blockade during adolescence reverses AIE-induced cellular effects, and that MC4R antagonism in adulthood will rescue AIE-induced hyperalgesia.
Adolescent alcohol exposure leads to long term detrimental effects on behavior, with recent evidence indicating exposure causes increased responsiveness to pain or hyperalgesia in adulthood, however the neural and biochemical alterations leading to this phenotype are not known. Here, we propose a fellowship training plan where a physician-scientist in training will investigate adolescent alcohol effects on amygdalar neuropeptide and glutamate systems, and will test the role of those systems in mediating adolescent alcohol induced hyperalgesia in adulthood. The findings from the proposed work will (1) characterize how the amygdala changes during and after adolescent alcohol exposure, (2) test and inform the development of translationally relevant therapies, and (3) foster the development of a promising M.D./Ph.D. student with training in alcohol and pain related neuroscience.
