Sleep alterations in an environmental neurotoxin-induced model of parkinsonism

Abstract

Parkinson's disease (PD) is classically defined as a motor disorder resulting from decreased dopamine production in the basal ganglia circuit. In an attempt to better diagnose and treat PD before the onset of severe motor dysfunction, recent attention has focused on the early, non-motor symptoms, which include but are not limited to sleep disorders such as excessive daytime sleepiness (EDS) and REM behavioral disorder (RBD). However, few animal models have been able to replicate both the motor and non-motor symptoms of PD. Here, we present a progressive rat model of parkinsonism that displays disturbances in sleep/wake patterns. Epidemiological studies elucidated a link between the Guamanian variant of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex (ALS/PDC) and the consumption of flour made from the washed seeds of the plant Cycas micronesica (cycad). Our study examined the effects of prolonged cycad consumption on sleep/wake activity in male, Sprague-Dawley rats. Cycad-fed rats exhibited an increase in length and/or number of bouts of rapid eye movement (REM) sleep and Non-REM (NREM) sleep at the expense of wakefulness during the active period when compared to control rats. This hypersomnolent behavior suggests an inability to maintain arousal. In addition, cycad-fed rats had significantly fewer orexin cells in the hypothalamus. Our results reveal a novel rodent model of parkinsonism that includes an EDS-like syndrome that may be associated with a dysregulation of orexin neurons. Further characterization of this early, non-motor symptom, may provide potential therapeutic interventions in the treatment of PD.

Fig. 1

Consumption of cycad induces a hypersomnolent behavior. During the 12-h active phase, cycad exposure decreased the total minutes spent in wake (A, *p =0.0056; t₍₁₄₎ =3.272), increased the amount of time spent in NREM sleep (B, *p =0.0152; t₍₁₄₎ =2.763) and REM sleep (C, *p =0.0047; t₍₁₄₎ =3.353) compared to flour-fed rats. No significant changes were observed between flour-fed and cycad-fed rats in Wakefulness (p =0.93), NREM sleep (p =0.65), or REM sleep (p =0.29) during the 12-h quiescent phase. Values are the means±SEM of 8 animals in each group.

Fig. 2

Total Sleep Time is increased in cycad-fed rats. (A) Cycad-fed rats showed a significant increase in the percentage of TST over the active phase (*; p =0.0053; t₍₁₄₎ =3.299). TST over the 24-h period also increased but did not reach statistical significance (p =0.061; t₍₁₄₎ =2.030). (B) Distribution of the percentage of TST into 2-h bins revealed a main effect of cycad on TST over the Active phase (F_(1,14) =11.1, p =0.005), but not in the Quiescent phase (p =0.99). Post-hoc analysis revealed that cycad-treated rats accumulated significantly more total sleep at ZT8 and ZT12 compared to the controls (p < 0.05). Values are the means±SEM of 8 animals in each group.

Fig. 3

Consumption of cycad induces a loss of Orexin-A neurons. (A) Representative images of ORX-A-ir cells in flour-fed and cycad-fed rats (left=4× magnification; right=20× magnification). (B) Analysis of ORX-A cell counts revealed a significant decrease in the mean number of orexin-A positive cells in anatomically matched tissue section of cycad-fed rats compared to flour-fed rats (*; p =0.0332; t₍₁₄₎ =2.362). (C) Representative images of MCH-ir cells in flour-fed and cycad-fed rats (left=4× magnification; right=20× magnification). (D) Analysis revealed no significant differences in cell counts of cycad-fed rats when compared with controls (p =0.25). Values are the means±SEM of 8 animals in each group.

Fig. 4

Loss of Orexin-A neurons causes a change in activation of remaining cells. (A) An example of co-labeled c-fos and ORX-A positive cells at low (4×) and high (100×) magnification. (B) Using c-fos as a nuclear marker of cell activation, analysis of ORX-A neurons revealed a significant increase in the percentage of activated ORX-A-ir cells in the lateral hypothalamus of cycad-fed rats when compared to flour-fed rats (*p=0.0273; t₍₁₄₎ =2.46). Values are the means±SEM of 8 animals in each group.