2-phenyl-4-4-5-5-tetramethylimidazoline-1-oxyl-3-oxide and Hypoxia

Chronic intermittent hypoxia (CIH) is a concomitant of sleep apnea that produces a slowly developing chemosensory-dependent blood pressure elevation ascribed in part to NMDA receptor-dependent plasticity and reduced nitric oxide (NO) signaling in the carotid body. The hypothalamic paraventricular nucleus (PVN) is responsive to hypoxic stress and also contains neurons that express NMDA receptors and neuronal nitric oxide synthase (nNOS). We tested the hypothesis that extended (35 d) CIH results in a decrease in the surface/synaptic availability of the essential NMDA NR1 subunit in nNOS-containing neurons and NMDA-induced NO production in the PVN of mice. As compared with controls, the 35 d CIH-exposed mice showed a significant increase in blood pressure and an increased density of NR1 immunogold particles located in the cytoplasm of nNOS-containing dendrites. Neither of these between-group differences was seen after 14 d, even though there was already a reduction in the NR1 plasmalemmal density at this time point. Patch-clamp recording of PVN neurons in slices showed a significant reduction in NMDA currents after either 14 or 35 d exposure to CIH compared with sham controls. In contrast, NO production, as measured by the NO-sensitive fluorescent dye 4-amino-5-methylamino-2',7'-difluorofluorescein, was suppressed only in the 35 d CIH group. We conclude that CIH produces a reduction in the surface/synaptic targeting of NR1 in nNOS neurons and decreases NMDA receptor-mediated currents in the PVN before the emergence of hypertension, the development of which may be enabled by suppression of NO signaling in this brain region.

Topics: Analysis of Variance; Animals; Arginine; Blood Gas Analysis; Blood Pressure; Cyclic N-Oxides; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Free Radical Scavengers; Hydrogen-Ion Concentration; Hypoxia; Imidazoles; In Vitro Techniques; Male; Membrane Potentials; Mice; Mice, Inbred C57BL; Microscopy, Electron, Transmission; N-Methylaspartate; Neuronal Plasticity; Neurons; Nitric Oxide; Nitric Oxide Synthase Type I; Paraventricular Hypothalamic Nucleus; Receptors, N-Methyl-D-Aspartate; S-Nitroso-N-Acetylpenicillamine; Signal Transduction; Time Factors; Vasopressins

The objective of this work was to test the hypothesis that the limitation of nitric oxide (NO) availability accentuates microvascular reactivity to oxygen. The awake hamster chamber window model was rendered hypoxic and hyperoxic by ventilation with 10 and 100% oxygen. Systemic and microvascular parameters were determined in the two conditions and compared with normoxia in a group receiving the NO scavenger nitronyl nitroxide and a control group receiving only the vehicle (saline). Mean arterial blood pressure did not change with different gas mixtures during infusion of the vehicle, but it increased significantly in the NO-depleted group. NO scavenging increased the reactivity of microvessels to the changed oxygen supply, causing the arteriolar wall to significantly increase oxygen consumption. Tissue Po2 was correspondingly significantly reduced during NO scavenger infusion. The present findings support the hypothesis that microvascular oxygen consumption is proportional to oxygen-induced vasoconstriction. The effect of oxygen on vascular tone is modulated by NO. As a consequence, NO acts as a regulator of the vessel wall oxygen consumption. The vessel wall consumes oxygen in proportion to the local Po2, and an impairment of NO availability renders the circulation more sensitive to changes in the oxygen supply.

Topics: Animals; Blood Flow Velocity; Blood Pressure; Cricetinae; Cyclic N-Oxides; Dose-Response Relationship, Drug; Free Radical Scavengers; Hyperoxia; Hypoxia; Imidazoles; Microcirculation; Nitric Oxide; Oxygen; Skin; Vasoconstriction

Oxygen plays a key role in energy metabolism. However, there are organisms that survive severe shortfalls in oxygen. Drosophila embryos rapidly arrest development upon severe hypoxia and recover upon restoration of oxygen, even days later. Stabilization of the normally unstable engrailed RNA and protein preserved the localized striped pattern of this embryonic patterning gene during 3 days in hypoxia. Severe hypoxia blocked expression of a heat-shock-inducible lacZ transgene. Cyanide, a metabolic poison, did not immediately block gene expression or turnover, arguing against a passive response to energy limitation. In contrast, nitric oxide, a putative hypoxia signal, induced a reversible arrest of development, gene expression and turnover. Reciprocally, a nitric oxide scavenger allowed continued gene expression and turnover during hypoxia, but it reduced hypoxia tolerance. We suggest that hypoxia-induced stasis preserves the status quo of embryonic processes and promotes survival. Our data implicate nitric oxide as a mediator of this response and provide a system in which to investigate its action.

Topics: Animals; Body Patterning; Cyclic N-Oxides; Cycloheximide; Dactinomycin; Drosophila melanogaster; Drosophila Proteins; Electron Transport; Embryo, Nonmammalian; Enzyme Inhibitors; Free Radical Scavengers; Gene Expression Regulation, Developmental; Homeodomain Proteins; Hypoxia; Imidazoles; Nitric Oxide; Nitric Oxide Donors; Nucleic Acid Synthesis Inhibitors; Oxygen; Protein Synthesis Inhibitors; RNA Stability; S-Nitroso-N-Acetylpenicillamine; Sodium Cyanide; Transcription Factors; Transcription, Genetic; Transgenes

The relationship between nitric oxide (NO) and intracellular Ca2+ in hypoxic-ischemic brain damage is not known in detail. Here we used rat striatal slices perfused under low-oxygen and Ca2+-free conditions and cultured human astrocytoma cells incubated under similar conditions as models to study the dynamics of intracellular NO and Ca2+ in hypoxia-induced tissue damage. Exposure of rat striatal slices for 70 min to low oxygen tension elicited a delayed and sustained increase in the release of 45Ca2+. This was potentiated by the NO donors sodium nitroprusside (SNP) and spermine-NO and inhibited by N-omega-nitro-L-arginine methyl ester (L-NAME) or by the NO scavenger 2-phenyl-4,4,5,5 tetramethylimidazoline-1-oxyl-3-oxide (PTIO). A membrane-permeant form of heparin in combination with either ruthenium red (RR) or ryanodine (RY) also inhibited 45Ca2+ release. In human astrocytoma U-373 MG cells, hypoxia increased intracellular Ca2+ concentration ([Ca2+]i) by 67.2 +/- 13.1% compared to normoxic controls and this effect was inhibited by L-NAME, PTIO or heparin plus RR. In striatal tissue, hypoxia increased NO production and LDH release and both effects were antagonized by L-NAME. Although heparin plus RR or RY antagonized hypoxia-induced increase in LDH release they failed to counteract increased NO production. These data therefore indicate that NO contributes to hypoxic damage through increased intracellular Ca2+ mobilization from endoplasmic reticulum and suggest that the NO-Ca2+ signalling might be a potential therapeutic target in hypoxia-induced neuronal degeneration.

Topics: Animals; Anticoagulants; Astrocytoma; Calcium; Cell Line, Tumor; Corpus Striatum; Cyclic N-Oxides; Dose-Response Relationship, Drug; Drug Combinations; Drug Interactions; Enzyme Inhibitors; Free Radical Scavengers; Fura-2; Heparin; Humans; Hydro-Lyases; Hypoxia; Imidazoles; In Vitro Techniques; Intracellular Space; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Donors; Nitroprusside; Perfusion; Rats; Rats, Sprague-Dawley; Ruthenium; Ryanodine