Low dose pramipexole is neuroprotective in the MPTP mouse model of Parkinson's disease, and downregulates the dopamine transporter via the D3 receptor

BMC Biology Low dose pramipexole is neuroprotective in the MPTP mouse model of Parkinson's disease, and downregulates the dopamine transporter via the D3 receptor Jeffrey N Joyce 0 Cheryl Woolsey 0 Han Ryoo 0 Sabine Borwege 0 Diane Hagner 0 0 Address: Thomas H. Christopher Center for Parkinson's Disease Research, Sun Health Research Institute , 10515 West Santa Fe Dr., Sun City, AZ, 85352 , USA Background: Our aim was to determine if pramipexole, a D3 preferring agonist, effectively reduced dopamine neuron and fiber loss in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model when given at intraperitoneal doses corresponding to clinical doses. We also determined whether subchronic treatment with pramipexole regulates dopamine transporter function, thereby reducing intracellular transport of the active metabolite of MPTP, 1-methyl-4-phenylpyridinium (MPP+). Methods: Ten 12-month old C57BL/6 mice were treated with MPTP (or saline) twice per day at 20 mg/ kg s.c. (4 injections over 48 h). Mice were pretreated for 3 days and during the 2-day MPTP regimen with pramipexole (0.1 mg/kg/day) or saline. Stereological quantification of dopamine neuron number and optical density measurement of dopamine fiber loss were carried out at 1 week after treatment, using immunostaining for dopamine transporter (DAT) and tyrosine hydroxylase (TH). Additional wild-type (WT) and D3 receptor knockout (KO) mice were treated for 5 days with pramipexole (0.1 mg/kg/day) or vehicle. The kinetics of [3H]MPP+ and [3H]DA uptake (Vmax and Km) were determined 24 h later; and at 24 h and 14 days dopamine transporter density was measured by quantitative autoradiography. Results: Pramipexole treatment completely antagonized the neurotoxic effects of MPTP, as measured by substantia nigra and ventral tegmental area TH-immunoreactive cell counts. MPTP- induced loss of striatal innervation, as measured by DAT-immunoreactivity, was partially prevented by pramipexole, but not with regard to TH-IR. Pramipexole also reduced DAT- immunoreactivity in non-MPTP treated mice. Subchronic treatment with pramipexole lowered the Vmax for [3H]DA and [3H]MPP+ uptake into striatal synaptosomes of WT mice. Pramipexole treatment lowered Vmax in WT but not D3 KO mice; however, D3 KO mice had lower Vmax for [3H]DA uptake. There was no change in DAT number in WT with pramipexole treatment or D3 KO mice at 24 h post-treatment, but there was a reduction in WTpramipexole treated and not in D3 KO mice at 14 days post-treatment. Conclusion: These results suggest that protection occurs at clinically suitable doses of pramipexole. Protection could be due to a reduced amount of MPP+ taken up into DA terminals via DAT. D3 receptor plays an important role in this regulation of transporter uptake and availability. - Background An interesting development in the use of dopamine (DA) agonists for treatment of Parkinson's disease (PD) is that some of them have proven to be neuroprotective in animal models of PD. Antiparkinsonian agents that are direct DA agonists, such as apomorphine [1], bromocriptine [2], and pramipexole [3], are neuroprotective against 1methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)induced damage to the DA system in mice. Administration of MPTP, which is converted to 1-methyl-4-phenylpyridinium (MPP+) and intracellularly transported into DAergic neurons [4], provides a good model for studying neuroprotection in PD. MPTP produces Parkinsonism in humans and in subhuman species through selective loss of DAergic neurons of the substantia nigra (SN) [5,6], and a number of related compounds to MPTP also produce nigral cell loss in primates [7]. MPTP causes apoptosis associated with PD [8-10] ;MPTP produces progressive cell death in humans for decades after the initial insult [11]. Hence, drugs that reduce the neurotoxicity of compounds like MPTP may be neuroprotective in PD. In fact, it is now hypothesized that direct DA agonists may slow the loss of DAergic terminal function upon longterm administration to PD patients [12-15]. Dopaminergic neurons are tonically inhibited by dendritic and terminal autoreceptors, operating in interaction with DA transporters (DAT) and pharmacologically of the D2 receptor subtype [16-19]. However, Zapata et al [20] have reported that the D3 preferring agonist (+)-PD 128907 regulates extracellular DA levels via interactions with D3 autoreceptors. If D3 preferring agonists are potent autoreceptor agonists, then hypothetically long-term changes in expression of DAT or the functional properties of DAT might occur following subchronic treatment. Since intracellular accumulation of MPP+ following systemic injection of MPTP requires DAT [4], then when DAT is downregulated by D3 preferring agonists, this could result in lower intracellular accumulation of MPP+ and reduced neurotoxicity to MPTP. The D3 receptor preferring agonists, pramipexole and ropinirole, are the most potent of the DA agonists affording neuroprotection at 1 mg/kg for pramipexole against MPTP-induced neurodegeneration [3,21] and at 2 mg/kg for ropinirole against 6-OHDA lesions in rats [22]. Doses 1030 times higher of DA agonists with low D3 receptor affinity such as apomorphine [1] and bromocriptine [2,23] are needed against MPTP-induced neurodegeneration. Because neuroprotection by pramipexole is most evident with concurrent treatment with MPTP and not with post-MPTP treatment [24], i.e. when autoreceptor contributions should be most pronounced, regulation of DAT may be important. In addition, while the lowest effective dose reported is 1.0 mg/kg for mice, this is significantly greater than a clinically relevant dose in humans (1.5 mg t.i.d., p.o.[25]). Based on information from Pharmacia Corporation, equivalent plasma levels obtained with 1.5 mg t.i.d., p.o. in humans could be produced with 0.1 to 0.5 mg/kg in the mice. We tested whether 0.1 mg/kg pramipexole would be neuroprotective in aging mice against MPTP-induced neurodegeneration to the DA system, and if this effect could be due to regulation of DAT function. Results Neurohistopathology Male C57BL/6 mice of 810 months of age were pretreated with saline or pramipexole (0.1 mg/kg/day) followed by MPTP. At the end of the 7-day recovery period following the last injection of MPTP or vehicle were assessed for the degree of toxicity to the dopamine system by MPTP. MPTP produced a marked loss of tyrosine hydroxylase-immunoreactive (TH-IR) neurons in the substantia nigra (SN), but had less impact in the ventral tegmental area (VTA) (Figs 1 and 2), Unbiased stereological quantification of the number of Nissl-stained and TH-IR neurons in the SNpc and VTA was made in the midbrains of the treated groups. MPTP produced a 31% loss of THIR neurons in the SN and 17% loss in the VTA. Pramipexole administered once a day for 5 days (i.e. 3 days prior to and during the 2-day vehicle treatment) did not alter the total number of TH-IR neurons in the SN or VTA. Pramipexole a (...truncated)