Bidirectional sex‐dependent regulation of α6 and β3 nicotinic acetylcholine receptors by protein kinase Cε

Abstract

Nicotine and alcohol are the most commonly abused substances worldwide and are frequently coabused. Nicotinic acetylcholine receptors (nAChRs) containing the α6 and β3 subunits are expressed in neural reward circuits and are critical for nicotine and alcohol reward. nAChRs are dynamically regulated by signaling molecules such as protein kinase C epsilon (PKCε), which impact transcription of α6 and β3 subunit mRNA (Chrna6 and Chrnb3, respectively). Previous work found decreased expression of Chrna6 and Chrnb3 transcripts in the ventral midbrain of male PKCε−/− mice, who also consume less nicotine and alcohol compared with wild‐type (WT) littermates. Using RT‐qPCR, we show that female PKCε−/− mice have higher expression of Chrna6 and Chrnb3 transcripts in the ventral midbrain, which functionally impacts nAChR‐dependent behavior as female but not male PKCε−/− mice exhibit locomotor hypersensitivity to low‐dose (0.25 mg/kg i.p.) nicotine. Female PKCε−/− mice show no differences in alcohol‐induced sedation in the loss‐of‐righting reflex assay (4.0 g/kg i.p.) compared with WT littermates, whereas male PKCε−/− mice have enhanced sedation compared with WT mice. Female PKCε−/− mice also show reduced immobility time in response to varenicline (1.0 mg/kg i.p.) compared with WT littermates in the tail suspension test, and this effect was absent in male mice. Additionally, we found that female PKCε−/− mice show altered alcohol and nicotine consumption patterns in chronic voluntary two‐bottle choice assays. Our data reveal a bidirectional effect of sex in the transcriptional regulation of nicotinic receptors by PKCε, highlighting the importance of studying both sexes in preclinical animal models.

Addiction‐related neuroadaptations following chronic nicotine exposure

“…Furthermore, the PKCε knockout mice had lower expression levels of α6 and β3 nAChR subunit transcripts in ventral midbrain and striatum compared with wild-type mice (Lee & Messing, 2011). Most recently, it was shown that female PKCε knockout mice exhibit higher levels of α6 and β3 nAChR subunit transcripts in ventral midbrain, and show increased sensitivity to the locomotor stimulating effects of nicotine compared to female wild-type and male PKCε knockout mice (Moen et al, 2020). Furthermore, female PKCε knockout mice consumed more nicotine during the first week of access in a twobottle choice procedure than wild-type mice (Moen et al, 2020).…”

Sex Differences in the Nicotinic Acetylcholine Receptor System of Rodents: Impacts on Nicotine and Alcohol Reward Behaviors

“…As α6 * nAChRs in the ventral midbrain impact the addictive properties of both alcohol and nicotine, an effect of sex in their regulatory mechanism has important implications for drug development. Notably, we did not observe a sex difference in transcript expression levels of α6, β3, or PKCε itself in PKCε WT animals (Moen et al, 2021). It is possible that the regulation of α6 and β3 nAChR transcripts occurs through distinct PKCε-medicated mechanisms in male and female animals that arise following sexual differentiation of the brain.…”

“…One candidate regulatory mechanism involves signaling by protein kinase C epsilon (PKCε), as our group has previously shown that genetic ablation of PKCε results in decreased α6 and β3 nAChR mRNA expression in male mice ( Lee and Messing, 2011 ). However, further characterization of female mice revealed an oppositional effect of sex, with female PKCε knockout animals showing a threefold increase in α6 and β3 transcript expression in the ventral midbrain compared with WT littermates ( Moen et al, 2021 ). Our data also showed a strong correlation between α6 and β3 transcript expression levels in the ventral midbrain across both WT and KO animals, further suggesting shared transcriptional mechanisms involving the CHRNB3-CHRNA6 gene cluster.…”

“…The PKCε WT animals are maintained on a 129SvJae/J background with a single generation cross to C57BL/6J ( Khasar et al, 1999 ; Moen et al, 2021 ). C57BL/6J mice have unique phenotypes compared with the 129SvJae/J line, including decreased sensitivity to several drug classes and decreased anxiety-like behaviors ( Homanics et al, 1999 ).…”

“…While bidirectional sex differences in reward-related behaviors is uncommon, our work examining the influence of PKCε on nAChR expression revealed a similar effect on nicotine consumption in male and female PKCε knockout mice. Male PKCε knockouts have decreased expression of α6 and β3 transcript and showed decreased nicotine consumption during the first week of a 4-week nicotine 2-bottle choice assay compared with wild-type littermates ( Lee and Messing, 2011 ), whereas female PKCε knockouts showed increased consumption during the first week of the same procedure ( Moen et al, 2021 ).…”