Abstract
The 6-Hydroxydopamine (6-OHDA) rat model of Parkinson's disease is essential for a better understanding of the pathological processes underlying the human disease and for the evaluation of promising therapeutic interventions. This work evaluated whether a single striatal injection of 6-OHDA causes progressive apoptosis of dopamine (DA) neurons and activation of glycogen synthase kinase 3β (GSK-3β) and caspase-3 in the substantia nigra compacta (SNc). The loss of DA neurons was shown by three neuron markers; tyrosine hydroxylase (TH), NeuN, and β-III tubulin. Apoptosis activation was determined using Apostain and immunostaining against cleaved caspase-3 and GSK-3β pY216. We also explored the possibility that cleaved caspase-3 is produced by microglia and astrocytes. Our results showed that the 6-OHDA caused loss of nigral TH(+) cells, progressing mainly in rostrocaudal and lateromedial directions. In the neostriatum, a severe loss of TH(+) terminals occurred from day 3 after lesion. The disappearance of TH(+) cells was associated with a decrease in NeuN and β-III tubulin immunoreactivity and an increase in Apostain, cleaved caspase-3, and GSK-3β pY216 in the SNc. Apostain immunoreactivity was observed from days 3 to 21 postlesion. Increased levels of caspase-3 immunoreactivity in TH(+) cells were detected from days 1 to 15, and the levels then decreased to day 30 postlesion. The cleaved caspase-3 also collocated with microglia and astrocytes indicating its participation in glial activation. Our results suggest that caspase-3 and GSK-3β pY216 activation might participate in the DA cell death and that the active caspase-3 might also participate in the neuroinflammation caused by the striatal 6-OHDA injection.
Highlights
The unilateral 6-hydroxydopamine (6-OHDA) lesion of the nigrostriatal dopamine (DA) system in the rat has become an essential model for the understanding of the neuropathology of Parkinson’s disease (PD) [1,2,3,4,5], the pharmacological characterization of new antiparkinsonian drugs [6], and the evaluation of promising therapies for PD [7,8,9]
This work shows for the first time that 6-OHDA injected into the neostriatum leads to a progressive loss of cytoskeleton integrity of the tyrosine hydroxylase (TH)(+) cells in the substantia nigra compacta (SNc)
The increased immunoreactivity of GSK3b pY216 found in this study in nigral DA neurons at days 3 and 15 shows activation of GSK3b pY216 and suggests participation in the 6-OHDA-triggered DA neuron death in vivo
Summary
The unilateral 6-hydroxydopamine (6-OHDA) lesion of the nigrostriatal dopamine (DA) system in the rat has become an essential model for the understanding of the neuropathology of Parkinson’s disease (PD) [1,2,3,4,5], the pharmacological characterization of new antiparkinsonian drugs [6], and the evaluation of promising therapies for PD [7,8,9]. Though 6-OHDA has been used since the early 1960s [15], the molecular and cellular mechanisms of 6-OHDA cytotoxicity in the DA nigrostriatal system have not been fully characterized Elucidation of those mechanisms is of critical importance to gain an insight into the mechanisms of neurotrophic factor therapy and those of drugs targeting the celldeath signaling pathway [16,17]. Some studies have suggested apoptosis in the SNc using silver staining or the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay after a 6-OHDA striatal injection [19,20,21,22] These studies did not accompany the TUNEL assays with other apoptosis markers such as caspases and key proteins in the signal transduction pathways of caspase activation to soundly support the participation of apoptosis in the effect of 6-OHDA. These antecedents suggest that cleaved caspase-3 might be associated with apoptosis of DA neurons and neuroinflammation, but in different stages of the neurodegenerative process
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