Contribution of the striatum to the effects of 5-HT1A receptor stimulation in L-DOPA-treated hemiparkinsonian rats

Abstract

Clinical and experimental studies implicate the use of serotonin (5-HT)1A receptor agonists for the reduction of L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia (LID). Although raphe nuclei likely play a role in these antidyskinetic effects, an unexplored population of striatal 5-HT1A receptors (5-HT1AR) may also contribute. To better characterize this mechanism, L-DOPA-primed hemiparkinsonian rats received the 5-HT1AR agonist +/-8-OH-DPAT (0, 0.1, 1.0 mg/kg, i.p.) with or without cotreatment with the 5-HT1AR antagonist WAY100635 (0.5 mg/kg, i.p.) 5 min after L-DOPA, after which abnormal involuntary movements (AIMs), rotations, and forelimb akinesia were quantified. To establish the effects of 5-HT1AR stimulation on L-DOPA-induced c-fos and preprodynorphin (PPD) mRNA within the dopamine-depleted striatum, immunohistochemistry and real-time reverse transcription polymerase chain reaction, respectively, were used. Finally, to determine the contribution of striatal 5-HT1AR to these effects, L-DOPA-primed hemiparkinsonian rats received bilateral intrastriatal microinfusions of +/-8-OH-DPAT (0, 5, or 10 microg/side), WAY100635 (5 microg/side), or both (10 microg + 5 microg/side) 5 min after L-DOPA, after which AIMs and rotations were examined. Systemic +/-8-OH-DPAT dose- and receptor-dependently attenuated L-DOPA-mediated AIMs and improved forelimb akinesia. Striatal c-fos immunoreactivity and PPD mRNA ipsilateral to the lesion were strongly induced by L-DOPA, while +/-8-OH-DPAT suppressed these effects. Finally, intrastriatal infusions of +/-8-OH-DPAT reduced AIMs while coinfusion of WAY100635 reversed its antidyskinetic effect. Collectively, these results support the hypothesis that the cellular and behavioral properties of 5-HT1AR agonists are conveyed in part via a population of functional 5-HT1AR within the striatum.

Fig. 1

Systemic ±8-OH-DPAT dose- and receptor-dependently reduces ALO AIMs expression. In counterbalanced within-subjects designs, one group of L-DOPA-primed rats (n = 8) received pre-treatment with vehicle (VEH) or the 5-HT1AR agonist ±8-OH-DPAT (D-0.1 or 1.0 mg/kg, i.p.) 5 min after treatments of L-DOPA + benserazide (12 + 15 mg/kg, i.p.), after which (A) axial, limb, and orolingual AIMs (ALO AIMs) and (B) rotations were observed every 20 min for 2 hr. In a separate group of L-DOPA-primed rats (n = 7) 5 min after pretreatment with vehicle (VEH), 1.0 mg/kg of ± 8-OH-DPAT (D-1.0), 0.5 mg/kg of the 5-HT1AR antagonist WAY100635 (W-0.5 mg/kg, i.p.) or combined 8-OH-DPAT + WAY100635 (1.0 mg/kg + 0.5 mg/kg, i.p.; D+W), rats received treatments of L-DOPA + benserazide (12 + 15 mg/kg, i.p.), after which (C) ALO AIMs and (D) rotations were assessed every 20 min for 2 hr. Symbols represent averaged ALO AIMs and net rotations ± SEM observed at each time point of testing. ALO AIMs were analyzed by nonparametric Friedman ANOVAs. Two-way parametric ANOVAs were used for analysis of rotations. Post hoc comparisons denote significant differences between treatments as indicated. ^*P < 0.05 vs. VEH, +P < 0.05 vs. D-0.1, #P < 0.05 vs. D+W.

Fig. 2

±8-OH-DPAT improves the efficacy of L-DOPA on the FAS test. In a counterbalanced within-subjects design (n = 8), baseline motor disability was established by a pretreatment of vehicle followed 5 min later by another vehicle injection (VEH+VEH). The antiparkinsonian efficacy of L-DOPA was determined by injection of vehicle followed by L-DOPA + benserazide (12 + 15 mg/kg, i.p.; VEH+LD). Finally, the effects of 5-HT1AR stimulation on L-DOPA efficacy were examined by injection of ±8-OH-DPAT (0.1 or 1.0 mg/kg, i.p.) followed by L-DOPA + benserazide, abbreviated D(0.1) + LD or D(1.0) + LD, respectively. Bars show the effects of treatments on FAS performance expressed as mean percentages of nonlesioned (Intact) FAS ± SEM in that same treatment condition. Effects were analyzed by a one-way ANOVA and significant differences were established by appropriate post hoc comparisons. ^*P < 0.05 vs. VEH+VEH; +P < 0.05 vs. VEH+LD.

Fig. 3

L-DOPA-induced striatal c-fos expression is reduced by ±8-OH-DPAT. In a between-subjects design, groups of L-DOPA-primed rats with equivalent AIMs expression (n = 4 rats/group) were randomly assigned to receive either vehicle followed 5 min later by another vehicle injection (VEH+VEH), vehicle 5 min after L-DOPA + benserazide (12 +15 mg/kg, i.p.; VEH+LD) or ±8-OH-DPAT (1.0 mg/kg, i.p.) followed by L-DOPA + benserazide (D(1.0) + LD). Rats were killed 1 hr after L-DOPA treatments and immediately perfused, after which 50-μm sections were processed for c-fos immunohistochemistry. A: Schematic depiction of dorsolateral (DL), dorsomedial (DM), ventrolateral (VL), and ventromedial (VM) striatal regions analyzed for c-fos expression by ImageJ software (NIH) and photorepresentations of (B) VEH+VEH, (C) VEH+LD, and (D) D(1.0) + LD treatments on striatal c-fos induction in the DA-lesioned DL striatum are shown. Scale bar = 100 μm. E: Scale bars depict the effects of treatments on striatal c-fos induction by region, expressed as mean number of c-fos positive cells/mm² ± SEM. Main treatment effects were determined by one-way ANOVA. Significant differences within each striatal region were established by planned comparison post hoc comparisons. ^*P < 0.05 vs. VEH + VEH; +P < 0.05 vs. VEH + LD. Anatomic structures: Aca, anterior commissure; Cpu, caudate putamen; Cc, corpus callosum; Ctx, cortex; LV, lateral ventricle.

Fig. 4

5-HT1AR stimulation reverses L-DOPA-induced increases in striatal PPD mRNA. In a between-subjects design, groups of L-DOPA-naive (vehicle-treated) and L-DOPA-primed rats (n = 10 rats/group) were assigned treatments. L-DOPA-naive rats received either vehicle (VEH/VEH) or L-DOPA + benserazide (12 + 15 mg/kg, i.p.; VEH/LD) while L-DOPA-primed rats received either L-DOPA + benserazide (12 + 15 mg/kg, i.p.; LD/LD) or ±8-OH-DPAT (1.0 mg/kg, i.p.) followed by L-DOPA + benserazide (12 + 15 mg/kg, i.p.; LD/D(1.0) + LD). Rats were killed 2 hr after treatments, and striata were immediately dissected and placed in RNA-later for subsequent analysis of PPD mRNA with RT-PCR. White and black bars depict the effects of treatment on striatal PPD mRNA in intact and lesioned striata, respectively, graphed as percentage change from control (nonlesioned striata treated with vehicle) ± SEM. Main effects of treatment and lesion, as well as treatment-by-lesion interactions, were determined by two-way ANOVA. Post hoc comparisons established significant differences between conditions as indicated. ^*P < 0.05 vs. all; +P < 0.05 vs. LD/LD.

Fig. 5

Striatal cannulae placements. Schematic representations of coronal sections of the rat brain taken from Paxinos and Watson (1998). A: Representative striatal section portraying the dorsolateral (DL) striatal target sites for drug injection. B: Shaded ovals depict the location of striatal microinfusion sites for rats included in Figure 6A,B. C: Open ovals depict striatal injection sites for rats included in Figure 6C,D. Anatomic structures: Aca, anterior commissure; Cpu, caudate putamen; Cc, corpus callosum; Ctx, cortex; LV, lateral ventricle.

Fig. 6

Direct striatal infusions of ±8-OH-DPAT dose- and receptor-dependently reduce ALO AIMs expression. In counterbalanced within-subject designs, one group of L-DOPA-primed rats (n = 10) received intrastriatal microinfusions of vehicle (VEH) or ±8-OH-DPAT (D-5.0 or 10.0 μg/side) 5 min after treatments of L-DOPA + benserazide (12 + 15 mg/kg, i.p.), after which (A) ALO AIMs and (B) rotations were observed every 20 min for 2 hr. In a separate group of L-DOPA-primed rats (n = 8) 5 min after intrastriatal microinfusions of vehicle (VEH), 10.0 μg/side of ±8-OH-DPAT (D-10.0), 5.0 μg/side of WAY100635 (W-5.0), or combined 8-OH-DPAT + WAY100635 (10.0 + 5.0 μg/side; D+W), rats received treatments of L-DOPA + benserazide (12 + 15 mg/kg, i.p.), after which (C) ALO AIMs and (D) rotations were assessed every 20 min for 2 hr. Symbols represent averaged ALO AIMs and net rotations ± SEM at each time point of testing. ALO AIMs were analyzed by nonparametric Friedman ANOVAs. Two-way parametric ANOVAs were used for analysis of rotations. Post hoc comparisons denote significant differences between treatments as indicated. ^*P < 0.05 vs. VEH, +P < 0.05 vs. D-5.0, #P < 0.05 vs. D+W.