Dopamine induces mitochondrial depolarization without activating PINK1‐mediated mitophagy

Abstract

Parkinson's disease (PD) is one of the most prevalent neurodegenerative disorders, characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta. PD mostly occurs sporadically and its cause remains unknown, nevertheless the discovery of familiar forms of PD, characterized by mutations of genes encoding proteins associated with mitochondria homeostasis, suggests a strong implication of the mitochondrial quality control system in PD. We investigated the effect of dopamine cytosolic accumulation in undifferentiated SH-SY5Y cells, an in vitro model widely used to reproduce impairment of dopamine homeostasis, an early step in PD pathogenesis. A strong depolarization of the mitochondrial membrane was observed after dopamine exposure. Nevertheless, mitochondrial network resulted to assume a peculiar morphology with a distinct pattern of OPA1 and MFN1, key regulators of mitochondrial dynamics. Moreover, selective elimination of dysfunctional mitochondria did not take place, suggesting an impairment of the mitophagic machinery induced by dopamine. Indeed, PINK1 did not accumulate on the outer mitochondrial membrane, nor was parkin recruited to depolarized mitochondria. Altogether, our results indicate that an improper handling of dysfunctional mitochondria may be a leading event in PD pathogenesis.

Exploring the Impact of PARK2 Mutations on the Total and Mitochondrial Proteome of Human Skin Fibroblasts

“…After 30 min of incubation at 37 • C, cells were washed with PBS and fixed with 4% paraformaldehyde for 15 min. Images were acquired through a cooled CCD camera on an Olympus IX81 microscope (40×) and analyzed using the ImageJ software, as previously described (Bondi et al, 2016). Seven fields of view randomly taken from three independent experiments were evaluated for each subject.…”

“…Five randomly chosen fields of view were captured and analyzed for each coverslip. The analysis of images was performed using the "Analyze Particles" function of the ImageJ software, as previously described (Bondi et al, 2016). The analysis was blinded.…”

“…As expected, CCCP induced fragmentation of the network in all samples ( Figure 3A and Supplementary Figure 3). To give a statistical assessment to this qualitative evaluation, images were analyzed with the "Analyze Particles" function of Image J (Bondi et al, 2016). By employing this strategy, it was possible to measure several morphological parameters that describe the mitochondrial network shape.…”

Network Analysis Identifies Disease-Specific Pathways for Parkinson’s Disease

“…Although it is difficult to speculate about the link between a single protein and a complex, multifactorial disease, we may envisage a role of the pentose phosphate pathway in the regulation of mitochondrial homeostasis in the presence of enhanced oxidative stress [49]. Indeed, aberrant control of mitochondrial function has been reported in a cellular model that specifically recapitulates dopamine-linked oxidative stress [11].…”

“…Progressive impairment of motor control is caused by the specific loss of dopaminergic neurons in the substantia nigra (SN) and the formation of intra-neuronal proteinaceous inclusions, called Lewy bodies (LBs) [7,8]. The progressive deterioration of vulnerable SN dopaminergic neurons may arise from cellular disturbances caused by misfolding and aggregation of the synaptic protein alpha synuclein [9], the disruption of the autophagy-lysosome system [10], mitochondrial dysfunction [11], endoplasmic reticulum (ER) stress [12], and dysregulation of calcium homeostasis [13]. These pathways are not specific for PD but they are shared by Alzheimer Disease (AD) [14] and Amyotrophic Lateral Sclerosis (ALS) [2] and there is recent evidence supporting the view that ALS and PD share common pathogenetic mechanisms.…”