Abstract
Abnormalities of alpha-synuclein (ASN), the main component of protein deposits (Lewy bodies), were observed in Parkinson’s disease (PD), dementia with Lewy bodies, Alzheimer’s disease, and other neurodegenerative disorders. These alterations include increase in the levels of soluble ASN oligomers in the extracellular space. Numerous works have identified several mechanisms of their toxicity, including stimulation of the microglial P2X7 receptor leading to oxidative stress. While the significant role of purinergic signaling—particularly, P2 family receptors—in neurodegenerative disorders is well known, the interaction of extracellular soluble ASN with neuronal purinergic receptors is yet to be studied. Therefore, in this study, we have investigated the effect of ASN on P2 purinergic receptors and ATP-dependent signaling. We used neuroblastoma SH-SY5Y cell line and rat synaptoneurosomes treated with exogenous soluble ASN. The experiments were performed using spectrofluorometric, radiochemical, and immunochemical methods. We found the following: (i) ASN-induced intracellular free calcium mobilization in neuronal cells and nerve endings depends on the activation of purinergic P2X7 receptors; (ii) activation of P2X7 receptors leads to pannexin 1 recruitment to form an active complex responsible for ATP release; and (iii) ASN greatly decreases the activity of extracellular ecto-ATPase responsible for ATP degradation. Thus, it is concluded that purinergic receptors might be putative pharmacological targets in the molecular mechanism of extracellular ASN toxicity. Interference with P2X7 signaling seems to be a promising strategy for the prevention or therapy of PD and other neurodegenerative disorders.
Highlights
Both oligomerization and accumulation of alpha-synuclein (ASN) are the key molecular processes involved in the pathophysiology of neurodegenerative diseases such as Parkinson’s disease (PD), Alzheimer’s disease (AD), and other synucleinopathies [1]
We used human neuroblastoma SH-SY5Y cell line, and because these cells are able to express a number of features characteristic for catecholaminergic neurons, including tyrosine hydroxylase and dopamine-β-hydroxylase activities [43], they express various P2 receptors belonging to both P2X and P2Y families as well [44]
In order to determine whether the increase of [Ca2 +]i is related to metabotropic P2Y receptor-mediated Ca2+ release from endoplasmic reticulum (ER), SH-SY5Y cells were treated with ASN in a calcium-free medium containing additional Ca2+-chelator, EGTA
Summary
Both oligomerization and accumulation of alpha-synuclein (ASN) are the key molecular processes involved in the pathophysiology of neurodegenerative diseases such as Parkinson’s disease (PD), Alzheimer’s disease (AD), and other synucleinopathies [1]. Previous data have indicated that the key process responsible for the propagation and expansion of neurodegeneration in the brain is the release of ASN to extracellular space [1, 3]. ASN released to extracellular compartment (by exocytosis or as an effect of neurodegeneration) changes the structure to β-sheet and acts outside the cell or penetrates the neuronal or glial cells in a manner dependent on the concentration and aggregation stage [4]. When ASN reaches the increased concentration inside the neuron, oligomerization that leads to cell death starts. The molecular mechanisms of extracellular ASN-mediated toxicity include Ca2+ deregulation, enhancement of the reactive oxygen and nitrogen species production, mitochondria dysfunction, deregulation of dopaminergic and glutamatergic neurotransmission, and activation of brain neuroinflammation [5,6,7,8,9,10,11,12]
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