TP53INP1 exerts neuroprotection under ageing and Parkinson's disease-related stress condition.

TP53INP1 is a stress-induced protein, which acts as a dual positive regulator of transcription and of autophagy and whose deficiency has been linked with cancer and metabolic syndrome. Here, we addressed the unexplored role of TP53INP1 and of its Drosophila homolog dDOR in the maintenance of neuronal homeostasis under chronic stress, focusing on dopamine (DA) neurons under normal ageing- and Parkinson's disease (PD)-related context. Trp53inp1-/- mice displayed additional loss of DA neurons in the substantia nigra compared to wild-type (WT) mice, both with ageing and in a PD model based on targeted overexpression of α-synuclein. Nigral Trp53inp1 expression of WT mice was not significantly modified with ageing but was markedly increased in the PD model. Trp53inp2 expression showed similar evolution and did not differ between WT and Trp53inp1-/- mice. In Drosophila, pan-neuronal dDOR overexpression improved survival under paraquat exposure and mitigated the progressive locomotor decline and the loss of DA neurons caused by the human α-synuclein A30P variant. dDOR overexpression in DA neurons also rescued the locomotor deficit in flies with RNAi-induced downregulation of dPINK1 or dParkin. Live imaging, confocal and electron microscopy in fat bodies, neurons, and indirect flight muscles showed that dDOR acts as a positive regulator of basal autophagy and mitophagy independently of the PINK1-mediated pathway. Analyses in a mammalian cell model confirmed that modulating TP53INP1 levels does not impact mitochondrial stress-induced PINK1/Parkin-dependent mitophagy. These data provide the first evidence for a neuroprotective role of TP53INP1/dDOR and highlight its involvement in the regulation of autophagy and mitophagy in neurons.

Enhanced differentiation of human induced pluripotent stem cells toward the midbrain dopaminergic neuron lineage through GLYPICAN-4 downregulation.

Enhancing the differentiation potential of human induced pluripotent stem cells (hiPSC) into disease-relevant cell types is instrumental for their widespread application in medicine. Here, we show that hiPSCs downregulated for the signaling modulator GLYPICAN-4 (GPC4) acquire a new biological state characterized by increased hiPSC differentiation capabilities toward ventral midbrain dopaminergic (VMDA) neuron progenitors. This biological trait emerges both in vitro, upon exposing cells to VMDA neuronal differentiation signals, and in vivo, even when transplanting hiPSCs at the extreme conditions of floor-plate stage in rat brains. Moreover, it is compatible with the overall neuronal maturation process toward acquisition of substantia nigra neuron identity. HiPSCs with downregulated GPC4 also retain self-renewal and pluripotency in stemness conditions, in vitro, while losing tumorigenesis in vivo as assessed by flank xenografts. In conclusion, our results highlight GPC4 downregulation as a powerful approach to enhance generation of VMDA neurons. Outcomes may contribute to establish hiPSC lines suitable for translational applications.

Progressive Parkinsonism by acute dysfunction of excitatory amino acid transporters in the rat substantia nigra.

Parkinson's disease (PD) is characterized by the progressive degeneration of substantia nigra (SN) dopamine neurons, involving a multifactorial cascade of pathogenic events. Here we explored the hypothesis that dysfunction of excitatory amino acid transporters (EAATs) might be involved. Acutely-induced dysfunction of EAATs in the rat SN, by single unilateral injection of their substrate inhibitor l-trans-pyrrolidine-2,4-dicarboxylate (PDC), triggers a neurodegenerative process mimicking several PD features. Dopamine neurons are selectively affected, consistent with their sustained excitation by PDC measured by slice electrophysiology. The anti-oxidant N-acetylcysteine and the NMDA receptor antagonists ifenprodil and memantine provide neuroprotection. Besides oxidative stress and NMDA receptor-mediated excitotoxicity, glutathione depletion and neuroinflammation characterize the primary insult. Most interestingly, the degeneration progresses overtime with unilateral to bilateral and caudo-rostral evolution. Transient adaptive changes in dopamine function markers in SN and striatum accompany cell loss and axonal dystrophy, respectively. Motor deficits appear when neuron loss exceeds 50% in the most affected SN and striatal dopamine tone is dramatically reduced. These findings outline a functional link between EAAT dysfunction and several PD pathogenic mechanisms/pathological hallmarks, and provide a novel acutely-triggered model of progressive Parkinsonism.

Impact of surgery targeting the caudal intralaminar thalamic nuclei on the pathophysiological functioning of basal ganglia in a rat model of Parkinson's disease.

There is accumulating evidence that the centre median-parafascicular (CM/Pf) complex of the thalamus is implicated in basal ganglia-related movement disorders and notably in Parkinson's disease. However, the impact of the changes affecting CM/Pf on the pathophysiological functioning of basal ganglia in parkinsonian state remains poorly understood. To address this issue, we have examined the effects of excitotoxic lesion of CM/Pf and of 6-hydroxydopamine-induced lesion of nigral dopamine neurons, separately or in association, on gene expression of markers of neuronal activity in the rat basal ganglia (striatal neuropeptide precursors, GAD67, cytochrome oxidase subunit I) by quantitative in situ hybridization histochemistry. CM/Pf lesion prevented the changes produced by the dopamine denervation in the components of the indirect pathway connecting the striatum to the output structures (striatopallidal neurons, globus pallidus, subthalamic nucleus), and among the output structures, in the entopeduncular nucleus. Preliminary data on the effects of deep brain stimulation of CM/Pf in rats with nigral dopamine lesion show that this surgical approach produces efficient anti-akinetic effect associated with partial reversal of the dopamine lesion-induced increase in striatal preproenkephalin A mRNA levels, a marker of the striatopallidal neurons. These data, which provide substrates for the potential of CM/Pf surgery in the treatment of movement disorders, are discussed in comparison with the effects of lesion or deep brain stimulation of the subthalamic nucleus, the currently preferred target for the surgical treatment of PD.

High-frequency stimulation of the subthalamic nucleus potentiates L-DOPA-induced neurochemical changes in the striatum in a rat model of Parkinson's disease.

This study examined the cellular changes produced in the striatum by chronic L-DOPA treatment and prolonged subthalamic nucleus high-frequency stimulation (STN-HFS) applied separately, successively, or in association, in the 6-hydroxydopamine-lesioned rat model of Parkinson's disease (PD). Only animals showing severe L-DOPA-induced dyskinesias (LIDs) were included, and STN-HFS was applied for 5 d at an intensity efficient for alleviating akinesia without inducing dyskinesias. L-DOPA treatment alone induced FosB/deltaFosB immunoreactivity, exacerbated the postlesional increase in preproenkephalin, reversed the decrease in preprotachykinin, and markedly increased mRNA levels of preprodynorphin and of the glial glutamate transporter GLT1, which were respectively decreased and unaffected by the dopamine lesion. STN-HFS did not affect per se the postlesion changes in any of these markers. However, when applied in association with L-DOPA treatment, it potentiated the positive modulation exerted by L-DOPA on all of the markers examined and tended to exacerbate LIDs. After 5 d of L-DOPA withdrawal, the only persisting drug-induced responses were an elevation in preprodynorphin mRNA levels and in the number of FosB/deltaFosB-immunoreactive neurons. Selective additional increases in these two markers were measured when STN-HFS was applied subsequently to L-DOPA treatment. These data provide the first evidence that STN-HFS exacerbates the responsiveness of striatal cells to L-DOPA medication and suggest that STN-HFS acts specifically through an L-DOPA-modulated signal transduction pathway associated with LIDs in the striatum. They point to striatal cells as a primary site for the complex interactions between these two therapeutic approaches in PD and argue against a direct anti-dyskinetic action of STN-HFS.

Differential effects of prolonged high frequency stimulation and of excitotoxic lesion of the subthalamic nucleus on dopamine denervation-induced cellular defects in the rat striatum and globus pallidus.

This study examined the effects of prolonged (4 days) high frequency stimulation (HFS) of the subthalamic nucleus (STN), in comparison with those of STN lesion, on the dopamine denervation-mediated cellular changes in the basal ganglia in a Wistar rat model of Parkinson's disease. STN HFS counteracted the dopamine lesion-induced increase in GAD67 mRNA expression in the output structures of the basal ganglia, as shown previously after STN lesion, providing cellular support for the similar antiparkinsonian benefits produced by the two surgical procedures. The dopamine denervation-induced increase in GAD67 mRNA levels in the globus pallidus was partially antagonized after HFS and totally reversed after ibotenate-induced STN lesion. The overexpression of striatal enkephalin mRNA tended to be further increased by HFS but was antagonized by STN lesion. The decrease in striatal substance P mRNA levels was affected neither by STN HFS nor lesion. As STN HFS for two hours was previously found not to interfere with the effects of dopamine lesion in the globus pallidus and striatum, the present data provide strong evidence that the effects of STN surgery in these structures involve long-term adaptive processes and that the rearrangements mediated by HFS and lesion are, at least in part, different.

High-frequency stimulation of the subthalamic nucleus selectively reverses dopamine denervation-induced cellular defects in the output structures of the basal ganglia in the rat.

High-frequency stimulation (HFS) of the subthalamic nucleus (STN) is now recognized as an effective treatment for advanced Parkinson's disease, but the molecular basis of its effects remains unknown. This study examined the effects of unilateral STN HFS (2 hr of continuous stimulation) in intact and hemiparkinsonian awake rats on STN neuron metabolic activity and on neurotransmitter-related gene expression in the basal ganglia, by means of in situ hybridization histochemistry and immunocytochemistry. In both intact and hemiparkinsonian rats, this stimulation was found to induce c-fos protein expression but to decrease cytochrome oxidase subunit I mRNA levels in STN neurons. STN HFS did not affect the dopamine lesion-mediated overexpression of enkephalin mRNA or the decrease in substance P in the ipsilateral striatum. The lesion-induced increases in intraneuronal glutamate decarboxylase 67 kDa isoform (GAD67) mRNA levels on the lesion side were reversed by STN HFS in the substantia nigra, partially antagonized in the entopeduncular nucleus but unaffected in the globus pallidus. The stimulation did not affect neuropeptide or GAD67 mRNA levels in the side contralateral to the dopamine lesion or in intact animals. These data furnish the first evidence that STN HFS decreases the metabolic activity of STN neurons and antagonizes dopamine lesion-mediated cellular defects in the basal ganglia output structures. They provide molecular substrate to the therapeutic effects of this stimulation consistent with the current hypothesis that HFS blocks STN neuron activity. However, the differential impact of STN HFS on the effects of dopamine lesion among structures receiving direct STN inputs suggests that this stimulation may not cause simply interruption of STN outflow.