Circadian Rhythm Abnormalities in Parkinson's Disease from Humans to Flies and Back

Abstract

Clinical and research studies have suggested a link between Parkinson's disease (PD) and alterations in the circadian clock. Drosophila melanogaster may represent a useful model to study the relationship between the circadian clock and PD. Apart from the conservation of many genes, cellular mechanisms, signaling pathways, and neuronal processes, Drosophila shows an organized central nervous system and well-characterized complex behavioral phenotypes. In fact, Drosophila has been successfully used in the dissection of the circadian system and as a model for neurodegenerative disorders, including PD. Here, we describe the fly circadian and dopaminergic systems and report recent studies which indicate the presence of circadian abnormalities in some fly PD genetic models. We discuss the use of Drosophila to investigate whether, in adults, the disruption of the circadian system might be causative of brain neurodegeneration. We also consider approaches using Drosophila, which might provide new information on the link between PD and the circadian clock. As a corollary, since PD develops its symptomatology over a large part of the organism's lifespan and given the relatively short lifespan of fruit flies, we suggest that genetic models of PD could be used to perform lifelong screens for drug-modulators of general and/or circadian-related PD traits.

Keywords: Drosophila; PD models; Parkison’s disease; circadian clock; dj-1; dopaminergic system.

Figure 1

Locomotor activity and organization of the circadian clock and dopaminergic system in the brain of D. melanogaster (A,B) Locomotor activity (mean ± SEM) of ~50 wild-type flies in (A) 12:12 light-dark (LD) and (B) constant darkness (DD) conditions, as a function of time. In (A) red arrows indicate the morning and evening anticipatory activities; ZT: Zeitgeber Time. In (B) CT: Circadian Time; (C) Schematic representation of the adult fly brain, in which the relative positions of the circadian neurons (right), the dopaminergic neurons (left), the mushroom bodies (MB), the central complex (CC), the Pars Intercerebralis (PI) are reported. Right hemisphere: lLNv: large ventral lateral neurons; sLNv: small LNvs; 5th sLNv: the fifth PDF-negative sLNv; LPN: lateral posterior neuron; LNd: dorsal LNs; DN1: dorsal neurons group 1; DN2: DN group 2; DN3: DN group 3; ey: relative position of the compound eye. Left hemisphere: PAM: Protocerebral Anterior Medial group; PAL: Protocerebral Anterior Lateral group; PPM1-4: Protocerebral Posterior Medial groups 1–4; PPL1–5: Protocerebral Posterior Lateral groups 1–5; VUM: Ventral Unpaired Median group; open circles in the left optic lobe indicate the TH positive cells of the medulla. (D) The two major TTLs of the circadian molecular clock in Drosophila. In the first Drosophila TTL, dCLK and dCYC form a dimer which binds the Enhancer boxes (E-boxes) in the promoter of the dper and dtim clock genes. dPER and dTIM proteins interact to form a complex, enter into the nucleus, and inhibit dCLK-dCYC activity. A second TTL modulates dClk expression: the dCLK-dCYC dimer induces the transcription of dvri and dPdp1 δ/ε genes. dVRI and dPDP1 δ/ε compete for the same element (D-box) in the dClk promoter, controlling dClk transcription. Post-translational modifications mediated by the kinases dDBT and dSGG modulate clock protein activities, regulating protein–protein interactions, nuclear translocation and degradation. In some clock neurons, light activates the internal photoreceptor dCRY which associates with dTIM and mediates its degradation. CLOCK: Circadian Locomotor Output Cycles Kaput; CRY: Cryptochrome; CYC: Cycle; PER: Period; TIM: Timeless; VRI: Vrille; PDP1: Par Domain Protein 1; DBT: Doubletime; SGG: Shaggy. Black arrows indicate the flow of the process; black sinusoidal lines and red arrows indicate transcriptional activity; blue arrows indicate the protein translation process (A,B) Graphs derived from data from our laboratory; (C,D) Modified from [54].

Figure 2

Locomotor activity in 12:12 LD and circadian periodicity in DD conditions of young (3–5-day-old) and old (30-day-old) ddj-1α⁻; ddj-1β⁻ double knock-out (DKO) males. ddj-1α⁻; ddj-1β⁻ DKO and w¹¹¹⁸ control flies were reared at 23 °C in 12:12 LD conditions (light intensity ~800 lux). 3–5 or ~30-day-old adult males were transferred into vials (~30 individuals each) containing normal or 1 mM Paraquat (PQ)-supplemented medium for 4 days. Subsequently, individuals were singly placed in glass tubes, containing normal or oxidative stress-inducing food. Locomotor activity was recorded as 1 min bins for 10 days in 12:12 LD conditions, or for 3 days in 12:12 LD followed by 7 days in DD regimes, using the Trikinetics Activity Monitor (DAM2). Circadian periodicity was analyzed as described in [124]. At least 3 replicates for each experiment were performed. (A–D) Locomotor activity profiles (mean ± SEM) of 50 young w¹¹¹⁸ (A), 61 young ddj-1α⁻; ddj-1β⁻ DKO (B), 38 old w¹¹¹⁸ (C), 79 old ddj-1α⁻; ddj-1β⁻ DKO (D) males for 3 days in 12:12 LD conditions. (E) Free-running periodicity of young and old ddj-1α⁻; ddj-1β⁻ DKO and w¹¹¹⁸ control flies in normal or mild oxidative stress conditions (1 mM PQ). N: number of analyzed flies; %R: percentage of rhythmic flies; tau: periodicity in DD. (F) Survival analysis to evaluate the effectiveness of the chronic exposure to 1 mM PQ on vitality in young flies. Only PQ-treated ddj-1α⁻; ddj-1β⁻ DKO males showed a significant decrease in vitality (***: p < 0.0001 for all paired comparisons in Mantel-Cox test; for each genotype and treatment at least 90 individuals were monitored in 3 independent replicates).

Figure 3

Evaluation of the relationship between PD and circadian clock alterations using D. melanogaster. Clock gene mutants (green) can be used to determine the possible presence of clock-related neurodegeneration, while circadian abnormalities might be evaluated in genetic PD models (red), at the behavioral, physiological, and molecular levels. Similar analyses could be performed to screen for modulating drugs (magenta), in order to ameliorate circadian alterations.