2--3--dialdehyde-atp and Hypoxia

Brain ischemia leading to stroke is a major cause of disability in developed countries. Therapeutic strategies have most commonly focused on protecting neurons from ischemic damage. However, ischemic damage to white matter causes oligodendrocyte death, myelin disruption, and axon dysfunction, and it is partially mediated by glutamate excitotoxicity. We have previously demonstrated that oligodendrocytes express ionotropic purinergic receptors. The objective of this study was to investigate the role of purinergic signaling in white matter ischemia. We show that, in addition to glutamate, enhanced ATP signaling during ischemia is also deleterious to oligodendrocytes and myelin, and impairs white matter function. Thus, ischemic oligodendrocytes in culture display an inward current and cytosolic Ca(2+) overload, which is partially mediated by P2X7 receptors. Indeed, oligodendrocytes release ATP after oxygen and glucose deprivation through the opening of pannexin hemichannels. Consistently, ischemia-induced mitochondrial depolarization as well as oxidative stress culminating in cell death are partially reversed by P2X7 receptor antagonists, by the ATP degrading enzyme apyrase and by blockers of pannexin hemichannels. In turn, ischemic damage in isolated optic nerves, which share the properties of brain white matter, is greatly attenuated by all these drugs. Ultrastructural analysis and electrophysiological recordings demonstrated that P2X7 antagonists prevent ischemic damage to oligodendrocytes and myelin, and improved action potential recovery after ischemia. These data indicate that ATP released during ischemia and the subsequent activation of P2X7 receptor is critical to white matter demise during stroke and point to this receptor type as a therapeutic target to limit tissue damage in cerebrovascular diseases.

Topics: Action Potentials; Adenosine Triphosphate; Animals; Animals, Newborn; Axons; Calcium; Cell Death; Connexins; Excitatory Amino Acid Antagonists; Glucose; Hypoxia; L-Lactate Dehydrogenase; Microscopy, Electron, Transmission; Nerve Tissue Proteins; Oligodendroglia; Optic Nerve; Optic Neuropathy, Ischemic; Patch-Clamp Techniques; Purinergic P2 Receptor Antagonists; Pyridoxal Phosphate; Rats; Reactive Oxygen Species; Receptors, Purinergic P2; Receptors, Purinergic P2X7

To investigate the hypoxia-induced death of rat retinal neurons and to determine whether P2X(7) activation is involved in this type of neuronal death.. Cultured retinal neurons from fetal rats were used. The effects and time course of various degrees of hypoxia (1%-5% O(2)) in the death of retinal neurons, were examined. The effects of P2X(7) antagonists, oxidized adenosine triphosphate (oxidized ATP; 30-100 microM), and brilliant blue G (BBG; 100 nM-10 microM) on hypoxia-induced neuronal death, including apoptosis, were assessed by using trypan blue exclusion, TUNEL assays, and cleaved caspase-3 immunoreactivity. Immunocytochemical analysis was performed to determine whether these neurons express P2X(7) receptors. The effects of P2X(7) receptor stimulation, induced by the P2X(7) agonist benzoyl- benzoyl-ATP (BzATP), on neuronal viability and intracellular Ca(2+) levels ([Ca(2+)](i)) were examined.. Retinal neuronal death increased according to the degree of hypoxia and became more severe after 12 hours. Both oxidized ATP and BBG significantly decreased hypoxia-induced neuronal death. Immunocytochemistry demonstrated that P2X(7) receptors were expressed by the cultured retinal neurons. ATP and BzATP caused P2X(7) receptor-dependent neuronal death in a dose-dependent manner and led to a sustained increase in [Ca(2+)](i), with BzATP being more effective than ATP. These effects were hypoxia-induced factor-1alpha- independent and were prevented by oxidized ATP.. The results suggest that the death of retinal neurons can be triggered by hypoxia and that P2X(7) activation is involved in the hypoxia-induced death of retinal neurons. P2X(7) antagonists can prevent hypoxia-induced damage in retinal neurons.

Topics: Adenosine Triphosphate; Animals; Animals, Newborn; Calcium; Caspase 3; Cell Death; Cells, Cultured; Dose-Response Relationship, Drug; Female; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; In Situ Nick-End Labeling; Pregnancy; Purinergic P2 Receptor Agonists; Purinergic P2 Receptor Antagonists; Rats; Rats, Wistar; Receptors, Purinergic P2; Receptors, Purinergic P2X7; Retinal Neurons; Rosaniline Dyes