Activation of PPAR gamma receptors reduces levodopa-induced dyskinesias in 6-OHDA-lesioned rats

Abstract

Long-term administration of l-3,4-dihydroxyphenylalanine (levodopa), the mainstay treatment for Parkinson's disease (PD), is accompanied by fluctuations in its duration of action and motor complications (dyskinesia) that dramatically affect the quality of life of patients. Levodopa-induced dyskinesias (LID) can be modeled in rats with unilateral 6-OHDA lesions via chronic administration of levodopa, which causes increasingly severe axial, limb, and orofacial abnormal involuntary movements (AIMs) over time. In previous studies, we showed that the direct activation of CB1 cannabinoid receptors alleviated rat AIMs. Interestingly, elevation of the endocannabinoid anandamide by URB597 (URB), an inhibitor of endocannabinoid catabolism, produced an anti-dyskinetic response that was only partially mediated via CB1 receptors and required the concomitant blockade of transient receptor potential vanilloid type-1 (TRPV1) channels by capsazepine (CPZ) (Morgese et al., 2007). In this study, we showed that the stimulation of peroxisome proliferator-activated receptors (PPAR), a family of transcription factors activated by anandamide, contributes to the anti-dyskinetic effects of URB+CPZ, and that the direct activation of the PPARγ subtype by rosiglitazone (RGZ) alleviates levodopa-induced AIMs in 6-OHDA rats. AIM reduction was associated with an attenuation of levodopa-induced increase of dynorphin, zif-268, and of ERK phosphorylation in the denervated striatum. RGZ treatment did not decrease striatal levodopa and dopamine bioavailability, nor did it affect levodopa anti-parkinsonian activity. Collectively, these data indicate that PPARγ may represent a new pharmacological target for the treatment of LID.

Keywords: Cannabinoid; Dyskinesia; Levodopa; PPARγ; Parkinson's disease; Rosiglitazone.

Figure 1

Effect of systemic administration of vehicle (5% PEG + 5% Tween-80 + 90% saline, i.p., 15 min before levodopa; open bar) on total levodopa-induced AIMs (axial+orofacial+limb) in 6-OHDA-lesioned rats (n=8/group). The FAAH inhibitor URB597 (URB 0.3 mg/kg, i.p., day 11, 15 min before levodopa) was co-administered with the TRPV1 antagonist capsazepine (CPZ, 10 mg/kg, i.p., 15 min before URB). The CB1 antagonist AM251 (1 mg/kg, i.p.) or the non-selective PPAR antagonist BADGE (BDG, 30 mg/kg, i.p.) were administered 15 min before CPZ (followed by URB) on day 12 and 13 of levodopa treatment, respectively. Levodopa effects were analyzed by Kruskal-Wallis followed by Dunn’s multiple comparison test (H=17.15, 3 d.f., p=0.0001; ***p<0.001 and *p<0.05 vs. levodopa+veh; ^#p<0.05 vs. URB+CPZ+BDG). AM251 and BDG effects were analyzed by Friedman test followed by Dunn’s multiple comparison test [Friedman statistic (chi-square)=9.33, 2 d.f., p=0.0055]. Data are expressed as median ± interquartile range.

Figure 2

Effect of chronic vehicle (saline, s.c.; open bars) or levodopa (6 mg/kg, s.c.; filled bars) on PPARα (a) and PPARγ (b) protein in the striatum of 6-OHDA-lesioned rats 1 h after last drug injection. (PPARα: t=0.2586, 6 d.f., p=0.8046; PPARγ: t=0.6088, 5 d.f., p=0.5692). Data are expressed as mean ± S.E.M. (C) PPARγ (green), dynorphin (red) or enkephalin (red) expression and respective colocalization (yellow) in striatal neurons of 6-OHDA-lesioned rats. Nuclei are visualized by Hoechst (blue). Scale bar, 50 µm.

Figure 3

Time course of the effects of systemic administration of levodopa (6 mg/kg, s.c., 1 injection per day; filled circles) measured at 60 (a) and 90 (b) minutes after levodopa injection. The selective PPARγ agonist, rosiglitazone (RGZ, 5 and 10 mg/kg, i.p.; open triangles and open squares, respectively) was administered 15 minutes before levodopa from day 9 through day 11. Panel a: RGZ 5 mg/kg: Limb day 10, H=9.948, 25 d.f., p=0.0069. RGZ 10 mg/kg: Limb day 9, H=10.97, 23 d.f., p=0.0042; Limb day 10, H=9.948, 25 d.f., p=0.0069; Limb day 11, H=11.08, 23 d.f., p=0.0039; Axial day 10, H=9.986, 19 d.f., p=0.0065; Axial day 11, H=8.933, 20 d.f., p=0.0115; Orofacial day 11, H=13.24, 24 d.f., p=0.0013. Panel b: RGZ 5mg/kg: Limb day 9, H=9.869, 17 d.f., p=0.0072; Limb day 10, H=15.27, 19 d.f., p=0.0005; Axial day 10, H=8.932, 14 d.f., p=0.0135; Orofacial day 9, H=13.24, 11 d.f., p=0.0399; Orofacial day 10, H=7.169, 11 d.f., p=0.0278. RGZ 10mg/kg: Limb day 9, H=9.869, 17 d.f., p=0.0072; Limb day 10, H=15.27, 19 d.f., p=0.0005; Limb day 11, H=13.98, 19 d.f., p=0.0009; Axial day 11, H=7.918, 13 d.f., p=0.0191; Orofacial day 11, H=10.97, 11 d.f., p=0.0088. ^# and * refer to RGZ 5mg/kg and RGZ 10mg/kg respectively. ^#*p<0.05, ^##, **p<0.01, ***p<0.001, Kruskal-Wallis followed by Dunn’s multiple comparison test. Data are expressed as median. The interquartile ranges were removed for clarity.

Figure 4

The selective PPARγ antagonist GW9662 (4 mg/kg, i.p.) reversed the anti-dyskinetic effect of rosiglitazone (RGZ, 10 mg/kg, i.p.) at 60 (a) and 90 (b) min after levodopa administration on day 12. GW9662 was administered 15 min before RGZ. Panel a: Limb, H=15.02, 34 d.f., p=0.0018; Axial, H=8.507, 36 d.f., p=0.04. Panel b: Limb, H=18.69, 28 d.f., p=0.0003; Axial, H=7.675, 34 d.f., p=0.05. *p<0.05; **p<0.01 and ***p<0.001 vs. veh+LD, Kruskal-Wallis followed by Dunn’s multiple comparison test. Data are expressed as median ± interquartile range.

Figure 5

Effect of 6-OHDA vehicle (Veh) plus saline (gray bars), or vehicle plus levodopa (Veh+LD, empty bars) or levodopa plus rosiglitazone (LD+RGZ, 10 mg/kg, i.p.; filled bars) on zif-268 and dynorphin mRNA levels (a), and p-ERK 1/2 protein levels (b) in the ipsilateral striatum of 6-OHDA-lesioned rats 90 min after last levodopa injection (day 12). Panel a: one-way ANOVA [zif-268: F_group(1,12)=15.55, p=0.0005; dynorphin: F_group (1,12)=16.19, p=0.0004] followed by Tukey post hoc test (*p<0.01 vs. veh+saline; ^#p<0.05 vs. veh+LD). Panel b: one-way ANOVA [F_group (1,22)=4.6418, p=0.042415] followed by Newman-Keuls post hoc (*p<0.05 vs. veh+saline; ^#p<0.05 vs. veh+LD). Data are expressed as mean ± S.E.M.

Figure 6

Effect of vehicle (5% PEG + 5% Tween-80 + 90% saline; open circles) or RGZ (10 mg/kg, i.p.; open squares) on levodopa (a) and dopamine (b) output in the ipsilateral striatum of 6-OHDA-lesioned rats (n = 5–7/group) 90 min after the last levodopa injection on day 12. ANOVA, F_time(17,170)=3.22, p=0.00005; F_group(1,10)=0.07, P=0.7 (a and b, respectively). Data are expressed as mean ±S.E.M.

Figure 7

Effect of vehicle (5% PEG + 5% Tween-80 + 90% saline; dark grey bar) or RGZ (10 mg/kg, i.p.; filled bar) on FAST (a) and VSFPT (b). Drugs were administered 15 min before levodopa (6 mg/kg, s.c). Panel a: one-way ANOVA [F_group (3, 20)=7.188, p=0.0018] followed by Newman-Keuls post hoc (^###p<0.001 vs. baseline; *p<0.05 vs. saline). Panel b: one-way ANOVA [F_group (3, 20)=43.94, p<0.0001] followed by Newman-Keuls post hoc (^###p<0.0001 vs. baseline; *p<0.05, vs. saline). Data are expressed as mean ± S.E.M

Scheme 1