5-Hydroxymethylfurfural reduces skeletal muscle superoxide production and modifies force production in rats exposed to hypobaric hypoxia.

Decreased blood-tissue oxygenation at high altitude (HA) increases mitochondrial oxidant production and reduces exercise capacity. 5-Hydroxymethylfurfural (5-HMF) is an antioxidant that increases hemoglobin's binding affinity for oxygen. For these reasons, we hypothesized that 5-HMF would improve muscle performance in rats exposed to a simulated HA of ~5500 m. A secondary objective was to measure mitochondrial activity and dynamic regulation of fission and fusion because they are linked processes impacted by HA. Fisher 344 rats received 5-HMF (40 mg/kg/day) or vehicle during exposure to sea level or HA for 72 h. Right ankle plantarflexor muscle function was measured pre- and post-exposure. Post-exposure measurements included arterial blood gas and complete blood count, flexor digitorum brevis myofiber superoxide production and mitochondrial membrane potential (ΔΨm), and mitochondrial dynamic regulation in the soleus muscle. HA reduced blood oxygenation, increased superoxide levels and lowered ΔΨm, responses that were accompanied by decreased peak isometric torque and force production at frequencies >75 Hz. 5-HMF increased isometric force production and lowered oxidant production at sea level. In HA exposed animals, 5-HMF prevented a decline in isometric force production at 75-125 Hz, prevented an increase in superoxide levels, further decreased ΔΨm, and increased mitochondrial fusion 2 protein expression. These results suggest that 5-HMF may prevent a decrease in hypoxic force production during submaximal isometric contractions by an antioxidant mechanism.

Comparison of treatment recompression tables for neurologic decompression illness in swine model.

BACKGROUND: Significant reductions in ambient pressure subject an individual to risk of decompression illness (DCI); with incidence up to 35 per 10,000 dives. In severe cases, the central nervous system is often compromised (>80%), making DCI among the most morbid of diving related injuries. While hyperbaric specialists suggest initiating recompression therapy with either a Treatment Table 6 (TT6) or 6A (TT6A), the optimal initial recompression treatment for severe DCI is unknown. METHODS: Swine were exposed to an insult dive breathing air at 7.06 ATA (715.35 kPa) for 24 min followed by rapid decompression at a rate of 1.82 ATA/min (184.41 kPa/min). Swine that developed neurologic DCI within 1 hour of surfacing were block randomized to one of four United States Navy Treatment Tables (USN TT): TT6, TT6A-air (21% oxygen, 79% nitrogen), TT6A-nitrox (50% oxygen, 50% nitrogen), and TT6A-heliox (50% oxygen, 50% helium). The primary outcome was the mean number of spinal cord lesions, which was analyzed following cord harvest 24 hours after successful recompression treatment. Secondary outcomes included spinal cord lesion incidence and gross neurologic outcomes based on a pre- and post- modified Tarlov assessment. We compared outcomes among these four groups and between the two treatment profiles (i.e. TT6 and TT6A). RESULTS: One-hundred and forty-one swine underwent the insult dive, with 61 swine meeting inclusion criteria (43%). We found no differences in baseline characteristics among the groups. We found no significant differences in functional neurologic outcomes (p = 0.77 and 0.33), spinal cord lesion incidence (p = 0.09 and 0.07), or spinal cord lesion area (p = 0.51 and 0.17) among the four treatment groups or between the two treatment profiles, respectively. While the trends were not statistically significant, animals treated with TT6 had the lowest rates of functional deficits and the fewest spinal cord lesions. Moreover, across all animals, functional neurologic deficit had strong correlation with lesion area pathology (Logistic Regression, p < 0.01, Somers' D = 0.74). CONCLUSIONS: TT6 performed as well as the other treatment tables and is the least resource intensive. TT6 is the most appropriate initial treatment for neurologic DCI in swine, among the tables that we compared.

Exogenous ketone salts inhibit superoxide production in the rat caudal solitary complex during exposure to normobaric and hyperbaric hyperoxia.

The use of hyperbaric oxygen (HBO2) in hyperbaric and undersea medicine is limited by the risk of seizures [i.e., central nervous system (CNS) oxygen toxicity, CNS-OT] resulting from increased production of reactive oxygen species (ROS) in the CNS. Importantly, ketone supplementation has been shown to delay onset of CNS-OT in rats by ∼600% in comparison with control groups (D'Agostino DP, Pilla R, Held HE, Landon CS, Puchowicz M, Brunengraber H, Ari C, Arnold P, Dean JB. Am J Physiol Regu Integr Comp Physiol 304: R829-R836, 2013). We have tested the hypothesis that ketone body supplementation inhibits ROS production during exposure to hyperoxygenation in rat brainstem cells. We measured the rate of cellular superoxide ([Formula: see text]) production in the caudal solitary complex (cSC) in rat brain slices using a fluorogenic dye, dihydroethidium (DHE), during exposure to control O2 (0.4 ATA) followed by 1-2 h of normobaric oxygen (NBO2) (0.95 ATA) and HBO2 (1.95, and 4.95 ATA) hyperoxia, with and without a 50:50 mixture of ketone salts (KS) dl-ß-hydroxybutyrate + acetoacetate. All levels of hyperoxia tested stimulated [Formula: see text] production similarly in cSC cells and coexposure to 5 mM KS during hyperoxia significantly blunted the rate of increase in DHE fluorescence intensity during exposure to hyperoxia. Not all cells tested produced [Formula: see text] at the same rate during exposure to control O2 and hyperoxygenation; cells that increased [Formula: see text] production by >25% during hyperoxia in comparison with baseline were inhibited by KS, whereas cells that did not reach that threshold during hyperoxia were unaffected by KS. These findings support the hypothesis that ketone supplementation decreases the steady-state concentrations of superoxide produced during exposure to NBO2 and HBO2 hyperoxia.NEW & NOTEWORTHY Exposure of rat medullary tissue slices to levels of O2 that mimic those that cause seizures in rats stimulates cellular superoxide ([Formula: see text]) production to varying degrees. Cellular [Formula: see text] generation in the caudal solitary complex is variable during exposure to control O2 and hyperoxia and significantly decreases during ketone supplementation. Our findings support the theory that ketone supplementation delays onset of central nervous system oxygen toxicity in mammals, in part, by decreasing [Formula: see text] production in O2-sensitive neurons.

Cardiovascular Parameters in a Swine Model of Normobaric Hypoxia Treated With 5-Hydroxymethyl-2-Furfural (5-HMF).

INTRODUCTION: The consequences of low partial pressure of O2 include low arterial O2 saturations (SaO2), low blood O2 content (CaO2), elevated mean pulmonary artery pressure (PAP), and decreased O2 consumption VO2. 5-hydroxymethyl-2-furfural (5-HMF) binds to the N-terminal valine of hemoglobin (HgB) and increases its affinity to O2. We used an instrumented, sedated swine model to study the effect of 5-HMF on cardiovascular parameters during exposure to acute normobaric hypoxia (NH). METHODS: Twenty-three sedated and instrumented swine were randomly assigned to one of three treatment groups and received equal volume of normal saline (VEH), 20 mg/kg 5-HMF (5-HMF-20) or 40 mg/kg 5-HMF (5-HMF-40). Animals then breathed 10% FiO2 for 120 min. Parameters recorded were Cardiac Output (CO), Mean Arterial Blood Pressure (MAP), Heart Rate (HR), Mean Pulmonary Artery Pressure (PAP), SaO2 and saturation of mixed venous blood (SvO2). The P50 was measured at fixed time intervals prior to and during NH. RESULTS: 5-HMF decreased P50. In the first 30 min of NH, treatment with 5-HMF-20 and 5-HMF-40 resulted in a (1) significantly smaller decrement in SaO2 and SvO2, (2) significantly lower HR and CO, and (3) smaller increase in PAP compared to VEH. In the 120 min of NH there was a trend toward improved mortality with 5-HMF treatment. CONCLUSION: 5-HMF treatment decreased P50, improved SaO2, and mitigated increases in PAP in this swine model of NH.

CNS function and dysfunction during exposure to hyperbaric oxygen in operational and clinical settings.

Hyperbaric oxygen (HBO2) is breathed during hyperbaric oxygen therapy and during certain undersea pursuits in diving and submarine operations. What limits exposure to HBO2 in these situations is the acute onset of central nervous system oxygen toxicity (CNS-OT) following a latent period of safe oxygen breathing. CNS-OT presents as various non-convulsive signs and symptoms, many of which appear to be of brainstem origin involving cranial nerve nuclei and autonomic and cardiorespiratory centers, which ultimately spread to higher cortical centers and terminate as generalized tonic-clonic seizures. The initial safe latent period makes the use of HBO2 practical in hyperbaric and undersea medicine; however, the latent period is highly variable between individuals and within the same individual on different days, making it difficult to predict onset of toxic indications. Consequently, currently accepted guidelines for safe HBO2 exposure are highly conservative. This review examines the disorder of CNS-OT and summarizes current ideas on its underlying pathophysiology, including specific areas of the CNS and fundamental neural and redox signaling mechanisms that are thought to be involved in seizure genesis and propagation. In addition, conditions that accelerate the onset of seizures are discussed, as are current mitigation strategies under investigation for neuroprotection against redox stress while breathing HBO2 that extend the latent period, thus enabling safer and longer exposures for diving and medical therapies.

Normobaric hyperoxia stimulates superoxide and nitric oxide production in the caudal solitary complex of rat brain slices.

Central CO2-chemosensitive neurons in the caudal solitary complex (cSC) are stimulated not only by hypercapnic acidosis, but by hyperoxia as well. While a cellular mechanism for the CO2 response has yet to be isolated, previous data show that a redox-sensitive mechanism underlies neuronal excitability to hyperoxia. However, it remains unknown how changes in Po2 affect the production of reactive oxygen and nitrogen species (RONS) in the cSC that can lead to increased cellular excitability and, with larger doses, to cellular dysfunction and death. To this end, we used fluorescence microscopy in real time to determine how normobaric hyperoxia increases the production of key RONS in the cSC. Because neurons in the region are CO2 sensitive, we also examined the potential effects of CO2 narcosis, used during euthanasia before brain slice harvesting, on RONS production. Our findings show that normobaric hyperoxia (0.4 → 0.95 atmospheres absolute O2) increases the fluorescence rates of fluorogenic dyes specific to both superoxide and nitric oxide. Interestingly, different results were seen for superoxide fluorescence when CO2 narcosis was used during euthanasia, suggesting long-lasting changes in superoxide production and/or antioxidant activity subsequent to CO2 narcosis before brain slicing. Further research needs to distinguish whether the increased levels of RONS reported here are merely increases in oxidative and nitrosative signaling or, alternatively, evidence of redox and nitrosative stress.

Acute hypercapnic hyperoxia stimulates reactive species production in the caudal solitary complex of rat brain slices but does not induce oxidative stress.

Central CO2 chemoreceptive neurons in the caudal solitary complex (cSC) are stimulated by hyperoxia via a free radical mechanism. Hyperoxia has been shown to increase superoxide and nitric oxide in the cSC, but it remains unknown how changes in Pco2 during hyperoxia affect the production of O2-dependent reactive oxygen and nitrogen species (RONS) downstream that can lead to increased levels of oxidative and nitrosative stress, cellular excitability, and, potentially, dysfunction. We used real-time fluorescence microscopy in rat brain slices to determine how hyperoxia and hypercapnic acidosis (HA) modulate one another in the production of key RONS, as well as colorimetric assays to measure levels of oxidized and nitrated lipids and proteins. We also examined the effects of CO2 narcosis and hypoxia before euthanasia and brain slice harvesting, as these neurons are CO2 sensitive and hypothesized to employ CO2/H+ mechanisms that exacerbate RONS production and potentially oxidative stress. Our findings show that hyperoxia ± HA increases the production of peroxynitrite and its derivatives, whereas increases in Fenton chemistry are most prominent during hyperoxia + HA. Using CO2 narcosis before euthanasia modulates cellular sensitivity to HA postmortem and enhances the magnitude of the peroxynitrite pathway, but blunts the activity of Fenton chemistry. Overall, hyperoxia and HA do not result in increased production of markers of oxidative and nitrosative stress as expected. We postulate this is due to antioxidant and proteosomal removal of damaged lipids and proteins to maintain cell viability and avoid death during protracted hyperoxia.

Female rats are more susceptible to central nervous system oxygen toxicity than male rats.

Abstract Tonic-clonic seizures typify central nervous system oxygen toxicity (CNS-OT) in humans and animals exposed to high levels of oxygen, as are encountered during scuba diving. We previously demonstrated that high doses of pseudoephedrine (PSE) decrease the latency to seizure (LS) for CNS-OT in young male rats. This study investigated whether female rats respond similarly to PSE and hyperbaric oxygen (HBO). We implanted 60 virgin stock (VS) and 54 former breeder (FB) female rats with radio-telemetry devices that measured brain electrical activity. One week later, rats were gavaged with saline or PSE in saline (40, 80, 120, 160, or 320 mg/kg) before diving to five atmospheres absolute in 100% oxygen. The time between reaching maximum pressure and exhibiting seizure was LS. Vaginal smears identified estrus cycle phase. PSE did not decrease LS for VS or FB, primarily because they exhibited low LS for all conditions tested. VS had shorter LS than males at 0, 40, and 80 mg/kg (-42, -49, and -57%, respectively). FB also had shorter LS than males at 0, 40, and 80 mg/kg (-60, -86, and -73%, respectively). FB were older than VS (286 ± 10 days vs. 128 ± 5 days) and weighed more than VS (299 ± 2.7 g vs. 272 ± 2.1 g). Males tested were younger (88 ± 2 days), heavier (340 ± 4.5 g), and gained more weight postoperatively (7.2 ± 1.6 g) than either VS (-0.4 ± 1.5 g) or FB (-1.6 ± 1.5 g); however, LS correlated poorly with age, body mass, change in body mass, and estrus cycle phase. We hypothesize that differences in sex hormones underlie females' higher susceptibility to CNS-OT than males.