Cytisine is neuroprotective in female but not male 6‐hydroxydopamine lesioned parkinsonian mice and acts in combination with 17‐β‐estradiol to inhibit apoptotic endoplasmic reticulum stress in dopaminergic neurons

Apoptotic endoplasmic reticulum (ER) stress is a major mechanism for dopaminergic (DA) loss in Parkinson's disease (PD). We assessed if low doses of the partial α4β2 nicotinic acetylcholine receptor agonist, cytisine attenuates apoptotic ER stress and exerts neuroprotection in substantia nigra pars compacta (SNc) DA neurons. Alternate day intraperitoneal injections of 0.2 mg/kg cytisine were administered to female and male mice with 6‐hydroxydopamine (6‐OHDA) lesions in the dorsolateral striatum, which caused unilateral degeneration of SNc DA neurons. Cytisine attenuated 6‐OHDA‐induced PD‐related behaviors in female, but not in male mice. We also found significant reductions in tyrosine hydroxylase (TH) loss within the lesioned SNc of female, but not male mice. In contrast to female mice, DA neurons within the lesioned SNc of male mice showed a cytisine‐induced pathological increase in the nuclear translocation of the pro‐apoptotic ER stress protein, C/EBP homologous protein (CHOP). To assess the role of estrogen in cytisine neuroprotection in female mice, we exposed primary mouse DA cultures to either 10 nM 17‐β‐estradiol and 200 nM cytisine or 10 nM 17‐β‐estradiol alone. 17‐β‐estradiol reduced expression of CHOP, whereas cytisine exposure reduced 6‐OHDA‐mediated nuclear translocation of two other ER stress proteins, activating transcription factor 6 and x‐box‐binding protein 1, but not CHOP. Taken together, these data show that cytisine and 17‐β‐estradiol work in combination to inhibit all three arms (activating transcription factor 6, x‐box‐binding protein 1, and CHOP) of apoptotic ER stress signaling in DA neurons, which can explain the neuroprotective effect of low‐dose cytisine in female mice. Hyperactive ER stress is a convergent apoptotic mechanism in Parkinson's disease (PD), characterized by the activation of three proteins, ATF6, XBP1, and CHOP. Using a 6‐OHDA model of PD in mice, we found that the nicotinic ligand, cytisine attenuated motor deficits and reduced dopaminergic degeneration only in female mice. 17‐β‐estradiol reduced CHOP expression in dopaminergic neurons, whereas cytisine attenuated ATF6 and XBP1 activation. Thus, low‐dose cytisine combined with 17‐β‐estradiol inhibits all three arms of apoptotic ER stress signaling. This study has important implications for developing combinations of nicotinic ligands with 17‐β‐estradiol analogs as a novel neuroprotective treatment for PD.