Dopamine Cytotoxicity on SH-SY5Y Cells: Involvement of α-Synuclein and Relevance in the Neurodegeneration of Sporadic Parkinson's Disease.

The cytotoxicity of dopamine on cultured cells of neural origin has been used as a tool to explore the mechanisms of dopaminergic neurodegeneration in Parkinson's disease. In the current study, we have shown that dopamine induces a dose-dependent (10-40 µM) and time-dependent (up to 96 h) loss of cell viability associated with mitochondrial dysfunction and increased intra-cellular accumulation of α-synuclein in cultured SH-SY5Y cells. Dopamine-induced mitochondrial dysfunction and the loss of cell viability under our experimental conditions could be prevented by cyclosporine, a blocker of mitochondrial permeability transition pore, as well as the antioxidant N-acetylcysteine. Interestingly, the dopamine effects on cell viability and mitochondrial functions were significantly prevented by knocking down α-synuclein expression by specific siRNA. Our results suggest that dopamine cytotoxicity is mediated by α-synuclein acting on the mitochondria and impairing its bioenergetic functions.

Synaptopathies: synaptic dysfunction in neurological disorders - A review from students to students.

Synapses are essential components of neurons and allow information to travel coordinately throughout the nervous system to adjust behavior to environmental stimuli and to control body functions, memories, and emotions. Thus, optimal synaptic communication is required for proper brain physiology, and slight perturbations of synapse function can lead to brain disorders. In fact, increasing evidence has demonstrated the relevance of synapse dysfunction as a major determinant of many neurological diseases. This notion has led to the concept of synaptopathies as brain diseases with synapse defects as shared pathogenic features. In this review, which was initiated at the 13th International Society for Neurochemistry Advanced School, we discuss basic concepts of synapse structure and function, and provide a critical view of how aberrant synapse physiology may contribute to neurodevelopmental disorders (autism, Down syndrome, startle disease, and epilepsy) as well as neurodegenerative disorders (Alzheimer and Parkinson disease). We finally discuss the appropriateness and potential implications of gathering synapse diseases under a single term. Understanding common causes and intrinsic differences in disease-associated synaptic dysfunction could offer novel clues toward synapse-based therapeutic intervention for neurological and neuropsychiatric disorders. In this Review, which was initiated at the 13th International Society for Neurochemistry (ISN) Advanced School, we discuss basic concepts of synapse structure and function, and provide a critical view of how aberrant synapse physiology may contribute to neurodevelopmental (autism, Down syndrome, startle disease, and epilepsy) as well as neurodegenerative disorders (Alzheimer's and Parkinson's diseases), gathered together under the term of synaptopathies. Read the Editorial Highlight for this article on page 783.

The Oral Iron Chelator, Deferasirox, Reverses the Age-Dependent Alterations in Iron and Amyloid-ß Homeostasis in Rat Brain: Implications in the Therapy of Alzheimer's Disease.

The altered metabolism of iron impacts the brain function in multiple deleterious ways during normal aging as well as in Alzheimer's disease. We have shown in this study that chelatable iron accumulates in the aged rat brain along with overexpression of transferrin receptor 1 (TfR1) and ferritin, accompanied by significant alterations in amyloid-ß (Aß) peptide homeostasis in the aging brain, such as an increased production of the amyloid-ß protein precursor, a decreased level of neprilysin, and increased accumulation of Aß42. When aged rats are given daily the iron chelator, deferasirox, over a period of more than 4 months starting from the 18th month, the age-related accumulation of iron and overexpression of TfR1 and ferritin in the brain are significantly prevented. More interestingly, the chelator treatment also considerably reverses the altered Aß peptide metabolism in the aging brain implying a significant role of iron in the latter phenomenon. Further, other results indicate that iron accumulation results in oxidative stress and the activation of NF-κB in the aged rat brain, which are also reversed by the deferasirox treatment. The analysis of the results together suggests that iron accumulation and oxidative stress interact at multiple levels that include transcriptional and post-transcriptional mechanisms to bring about changes in the expression levels of TfR1 and ferritin and also alterations in Aß peptide metabolism in the aging rat brain. The efficacy of deferasirox in preventing age-related changes in iron and Aß peptide metabolism in the aging brain, as shown here, has obvious therapeutic implications for Alzheimer's disease.

α-Synuclein-induced mitochondrial dysfunction in isolated preparation and intact cells: implications in the pathogenesis of Parkinson's disease.

This study has shown that purified recombinant human α-synuclein (20 µM) causes membrane depolarization and loss of phosphorylation capacity of isolated purified rat brain mitochondria by activating permeability transition pore complex. In intact SHSY5Y (human neuroblastoma cell line) cells, lactacystin (5 µM), a proteasomal inhibitor, causes an accumulation of α-synuclein with concomitant mitochondrial dysfunction and cell death. The effects of lactacystin on intact SHSY5Y cells are, however, prevented by knocking down α-synuclein expression by specific siRNA. Furthermore, in wild-type (non-transfected) SHSY5Y cells, the effects of lactacystin on mitochondrial function and cell viability are also prevented by cyclosporin A (1 µM) which blocks the activity of the mitochondrial permeability transition pore. Likewise, in wild-type SHSY5Y cells, typical mitochondrial poison like antimycin A (50 nM) produces loss of cell viability comparable to that of lactacystin (5 µM). These data, in combination with those from isolated brain mitochondria, strongly suggest that intracellularly accumulated α-synuclein can interact with mitochondria in intact SHSY5Y cells causing dysfunction of the organelle which drives the cell death under our experimental conditions. The results have clear implications in the pathogenesis of sporadic Parkinson's disease. α-Synuclein is shown to cause mitochondrial impairment through interaction with permeability transition pore complex in isolated preparations. Intracellular accumulation of α-synuclein in SHSY5Y cells following proteasomal inhibition leads to mitochondrial impairment and cell death which could be prevented by knocking down α-synuclein gene. The results link mitochondrial dysfunction and α-synuclein accumulation, two key pathogenic mechanisms of Parkinson's disease, in a common damage pathway.

Multiple mechanisms of iron-induced amyloid beta-peptide accumulation in SHSY5Y cells: protective action of negletein.

The increased accumulation of iron in the brain in Alzheimer's disease (AD) is well documented, and excess iron is strongly implicated in the pathogenesis of the disease. The adverse effects of accumulated iron in AD brain may include the oxidative stress, altered amyloid beta-metabolism and the augmented toxicity of metal-bound amyloid beta 42. In this study, we have shown that exogenously added iron in the form of ferric ammonium citrate (FAC) leads to considerable accumulation of amyloid precursor protein (APP) without a corresponding change in the concerned gene expression in cultured SHSY5Y cells during exposure up to 48 h. This phenomenon is also associated with increased ß-secretase activity and augmented release of amyloid beta 42 in the medium. Further, the increase in ß-secretase activity, in SHSY5Y cells, upon exposure to iron apparently involves reactive oxygen species (ROS) and NF-κB activation. The synthetic flavone negletein (5,6-dihydroxy-7-methoxyflavone), which is a known chelator for iron, can significantly prevent the effects of FAC on APP metabolism in SHSY5Y cells. Further, this compound inhibits the iron-dependent formation of ROS and also blocks the iron-induced oligomerization of amyloid beta 42 in vitro. In concentrations used in this study, negletein alone appears to have only marginal toxic effects on cell viability, but, on the other hand, the drug is capable of ameliorating the iron-induced loss of cell viability considerably. Our results provide the initial evidence of potential therapeutic effects of negletein, which should be explored in suitable animal models of AD.

Combination of N-acetylcysteine, α-lipoic acid and α-tocopherol substantially prevents the brain synaptosomal alterations and memory and learning deficits of aged rats.

This study has compared several synaptosomal parameters in three groups of rats: young (46 months), aged (22-24 months) and antioxidant supplemented aged rats (antioxidant supplementation given with the diet as a combination of N-acetylcysteine, α-lipoic acid and α-tocopherol from 18 months onwards till 22-24 months). The synaptosomes from aged rat brain, in comparison to those of young animals, exhibit an increased membrane potential with altered contents of Na(+) and K(+) under basal incubation condition and in the presence of depolarizing agents. The intrasynaptosomal Ca(2+) is also higher in aged than in young rat. These age-dependent changes in synaptosomal parameters are prevented markedly in the antioxidant supplemented group. When examined on T-maze, the aged animals are noticeably impaired in learning and memory functions, but the deficit is remarkably prevented in the antioxidant supplemented aged animals. It is suggested that the synaptosomal alterations partly contribute to the cognitive deficits of aged animals, and both are rescued by long-term antioxidant supplementation.