Involvement of the dorsal raphe serotonin system in basal and punished heroin self-administration and stress-induced reinstatement in male and female rats

Rationale Opioid use disorders are marked by both drug use despite negative consequences and drug use to avoid negative physiological and psychological states. The dorsal raphe nucleus (DRN) serotonin (5-HT) system modulates reward- and stress-related responses, but its role in both male and female rats remains incompletely understood. Objective To investigate the role of the DRN 5-HT system in heroin-motivated behaviors in male and female rats, and, in parallel, to examine the impacts of stress-induced reinstatement of heroin seeking on DRN 5-HT cell electrophysiological profiles. Methods To probe DRN 5-HT contributions to basal, punished, and stress-driven heroin consumption, we performed chemogenetic manipulations in transgenic Tph2-iCre Sprague-Dawley rats. Twenty-two- and 50-kHz ultrasonic vocalizations (USVs), reflecting negative and positive affective state, respectively, were recorded during basal and punished heroin self-administration and a novel dual stressor-induced reinstatement model to delineate affective profiles. In a separate cohort of conventional Sprague-Dawley rats, ex vivo electrophysiology in DRN 5-HT cells examined the impact of stress-induced reinstatement of heroin seeking on spontaneous inhibitory postsynaptic currents (sIPSCs). Results Activation of the DRN 5-HT system increased heroin consumption under basal and punished conditions but did not alter stress-induced reinstatement or affective measures. Although females showed higher active lever pressing and consumption of heroin intake during acquisition of self-administration, responses to DRN 5-HT activation did not differ by sex. Ex vivo recordings revealed elevated sIPSCs in DRN 5-HT neurons from post-reinstatement females, but not males, indicating increased inhibitory drive. Notably, chemogenetic viral expression was densest in the dorsal DRN (DRD), associated with anxiety-related behavior, whereas electrophysiological recordings targeted ventrolateral/ventral DRN (DRVL/DRV) regions linked to anxiolysis. Conclusions These findings demonstrate a role for the DRN 5-HT system in basal and punished heroin self-administration and a link between stress-induced heroin reinstatement and enhanced GABAergic inhibition of DRN 5-HT neurons in females. Serotonergic involvement in heroin consumption and sex differences in stress-driven heroin seeking patterns may reflect differential engagement of anatomically and functionally distinct DRN subdivisions.