Circadian-Dependent and Sex-Dependent Increases in Intravenous Cocaine Self-Administration inNpas2Mutant Mice

Abstract

Substance use disorder (SUD) is associated with disruptions in circadian rhythms. The circadian transcription factor neuronal PAS domain protein 2 (NPAS2) is enriched in reward-related brain regions and regulates reward, but its role in SU is unclear. To examine the role of NPAS2 in drug taking, we measured intravenous cocaine self-administration (acquisition, dose-response, progressive ratio, extinction, cue-induced reinstatement) in wild-type (WT) and Npas2 mutant mice at different times of day. In the light (inactive) phase, cocaine self-administration, reinforcement, motivation and extinction responding were increased in all Npas2 mutants. Sex differences emerged during the dark (active) phase with Npas2 mutation increasing self-administration, extinction responding, and reinstatement only in females as well as reinforcement and motivation in males and females. To determine whether circulating hormones are driving these sex differences, we ovariectomized WT and Npas2 mutant females and confirmed that unlike sham controls, ovariectomized mutant mice showed no increase in self-administration. To identify whether striatal brain regions are activated in Npas2 mutant females, we measured cocaine-induced DFosB expression. Relative to WT, DFosB expression was increased in D11 neurons in the nucleus accumbens (NAc) core and dorsolateral (DLS) striatum in Npas2 mutant females after dark phase self-administration. We also identified potential target genes that may underlie the behavioral responses to cocaine in Npas2 mutant females. These results suggest NPAS2 regulates reward and activity in specific striatal regions in a sex and time of day (TOD)-specific manner. Striatal activation could be augmented by circulating sex hormones, leading to an increased effect of Npas2 mutation in females.

Sex-specific role of the circadian transcription factor NPAS2 in opioid tolerance, withdrawal, and analgesia

“…Our results are also consistent with prior studies which examined intersectional consequences between sex and circadian genes such as Clock or Npas2 44,75 . Interestingly, Npas2 deletion had higher impact on cocaine reward and self-administration behaviors in female mice 44 .…”

“…Our results are also consistent with prior studies which examined intersectional consequences between sex and circadian genes such as Clock or Npas2 44,75 . Interestingly, Npas2 deletion had higher impact on cocaine reward and self-administration behaviors in female mice 44 . More profound consequences on females than males could be explained by levels of circulating hormones, as sex differences in cocaine self-administration were abolished in ovariectomized females 44 .…”

“…Interestingly, Npas2 deletion had higher impact on cocaine reward and self-administration behaviors in female mice 44 . More profound consequences on females than males could be explained by levels of circulating hormones, as sex differences in cocaine self-administration were abolished in ovariectomized females 44 . This was consistent with the fact that circulating estrogens were shown to be essential in orchestrating rhythms of circadian genes in the SCN 76 .…”

“…Previous studies implicated circadian regulation of pain, tolerance, and physical dependence 11,12 , suggesting bidirectional interactions between circadian rhythms and opioids 9 . NPAS2 is a circadian gene enriched in brain regions and spinal cord involved in pain and opioids, and previously shown to be involved in psychostimulant reward 44,45 . However, the role of NPAS2 in opioid tolerance and dependence had yet to be established.…”

The Ventral Tegmental Area and Nucleus Accumbens as Circadian Oscillators: Implications for Drug Abuse and Substance Use Disorders

“…Notably, while CLOCK seems to play a role in regulating reward through its expression in the VTA, NPAS2 has been shown to regulate reward through its expression in the NAc. The use of Npas2 mutant mice or knockdown of Npas2 in the NAc decreases cocaine conditioned place preference through its expression in D 1 MSNs ( Ozburn et al, 2015 ; Parekh et al, 2019 ; Becker-Krail et al, 2022b ); Npas2 mutant mice actually show significant increases in locomotor response to novelty, exploratory drive, cocaine self-administration and self-administration motivation ( Ozburn et al, 2017 ; DePoy et al, 2020 ). In addition to its enriched expression in the D 1 MSNs of the NAc, this disrupted reward regulation can also be attributed to NPAS2’s transcriptional regulation of reward-related transcripts and downstream regulation of excitatory synaptic transmission and plasticity ( Parekh et al, 2019 ; Becker-Krail et al, 2022b ).…”