Mycorrhizal feedbacks influence global forest structure and diversity.

One mechanism proposed to explain high species diversity in tropical systems is strong negative conspecific density dependence (CDD), which reduces recruitment of juveniles in proximity to conspecific adult plants. Although evidence shows that plant-specific soil pathogens can drive negative CDD, trees also form key mutualisms with mycorrhizal fungi, which may counteract these effects. Across 43 large-scale forest plots worldwide, we tested whether ectomycorrhizal tree species exhibit weaker negative CDD than arbuscular mycorrhizal tree species. We further tested for conmycorrhizal density dependence (CMDD) to test for benefit from shared mutualists. We found that the strength of CDD varies systematically with mycorrhizal type, with ectomycorrhizal tree species exhibiting higher sapling densities with increasing adult densities than arbuscular mycorrhizal tree species. Moreover, we found evidence of positive CMDD for tree species of both mycorrhizal types. Collectively, these findings indicate that mycorrhizal interactions likely play a foundational role in global forest diversity patterns and structure.

Pro-survival Phenotype of HIF-1α: Neuroprotection Through Inflammatory Mechanisms.

Hypoxia-inducible factor 1 (HIF-1) is a major player in the oxygen sensor system as well as a transcription factor. HIF-1 is also associated in the pathogenesis of many brain diseases including Alzheimer's disease (AD), epilepsy and stroke. HIF-1 regulates the expression of many genes such as those involved in glycolysis, erythropoiesis, angiogenesis and proliferation in hypoxic condition. Despite several studies, the mechanism through which HIF-1 confers neuroprotection remains unclear, one of them is modulating metabolic profiles and inflammatory pathways. Characterization of the neuroprotective role of HIF-1 may be through its stabilization and the regulation of target genes that aid in the early adaptation to the oxidative stressors. It is interesting to note that mounting data from recent years point to an additional crucial regulatory role for hypoxia-inducible factors (HIFs) in inflammation. HIFs in immune cells regulate the production of glycolytic energy as well as innate immunity, pro-inflammatory gene expression, and mediates activation of pro-survival pathways. The present review highlights the contribution of HIF-1 to neuroprotection where inflammation is the crucial factor in the pathogenesis contributing to neural death. The potential mechanisms that contribute to neuroprotection as a result of the downstream targets of HIF-1α are discussed. Such mechanisms include those mediated through IL-10, an anti-inflammatory molecule involved in activating pro-survival signaling mechanisms via AKT/ERK and JAK/STAT pathways.

Brainstem Damage Underlies Changes in Hypoxic Ventilatory Response Following Cardiac Arrest and Resuscitation in Rats.

People resuscitated after sudden cardiac arrest remain at high risk for mortality, with treatment for survivors varying from monitoring to life support. With respect to assessing survivability post cardiac arrest and resuscitation (CAR), we previously demonstrated the potential of the hypoxic ventilatory response (HVR) as a reliable indicator for discerning between survivors and non-survivors in the early stages of recovery following CAR in rats. Since HVR describes the increase in ventilation in response to hypoxia, we hypothesize that damage to cardiorespiratory regulatory centers in the brainstem underlie the loss of HVR observed post resuscitation in nonsurvivors. Wistar rats underwent cardiac arrest (12-min) and resuscitation. At 1 day post-resuscitation, rats were perfused transcardially and the brains were harvested and processed for immunohistostaining of caspase-3, a marker of apoptosis. Positive caspase-3 staining was observed in brainstem regions such as the rostral ventral lateral medulla (RVLM); Co-localization of caspase-3 and NeuN was observed in the RVLM as well, suggesting that apoptosis most likely occurs in neurons. Our results showed positive markers for neuronal apoptosis present in pathways of the brainstem involved in respiratory and cerebrovascular regulation, suggesting brain stem damage underlies changes in HVR following CAR.

Postnatal Exposure to Brief Hypoxia Alters Brain VEGF Expression and Capillary Density in Adult Mice.

Perinatal hypoxia leads to changes in cerebral angiogenesis and persistent structural and functional changes in the adult brain. It may also result in greater vulnerability to subsequent challenges. We investigated the effect of postnatal day 2 (P2) hypoxic preconditioning on adult brain capillary density and brain vascular endothelial growth factor (VEGF) expression in mice. P2 mice were exposed to hypoxia (5% O2) in a normobaric chamber for 2 h then returned to normoxia while their littermates remained in normoxia (P2 control). After 2-6 months, they were euthanised and their brains were removed for capillary density determination. Another set of animals (P2 hypoxic mice and P2 controls) were euthanised at 2, 10, 23, and 60 days after birth and brain VEGF expression was assessed by western blot. Adult brain capillary density was significantly increased in the P2 hypoxic mice when compared to the P2 control mice. Additionally, VEGF expression appeared to be elevated in the P2-hypoxia mice when compared to the P2-control mice at all time points, and VEGF levels in P2-hypoxia mice declined with age similarly to P2-control mice. These data demonstrate that transient early-postnatal hypoxic stress leads to an increase in capillary density that persists in the adult, possibly due to increased VEGF expression. These results might be explained by epigenetic factors in the VEGF gene.

Impaired Cognitive Performance in Mice Exposed to Prolonged Hyperoxia.

Supplementation of oxygen at concentrations significantly above environmental level for prolonged periods may lead to hyperoxia and tissue toxicity. The mammalian brain undergoes structural and functional changes during adaptation to hypoxia and hyperoxia. In this study we investigated the effect of prolonged hyperoxic exposure on cognitive and motor performance in mice. Two-month-old male mice were placed in either hyperoxic (50% O2) or normoxic conditions for 3 weeks. Cognitive function was measured using the Y-maze test. High alteration rate between the three arms of the maze is indicative of sustained memory and cognitive function. Motor function was measured using the grip strength and rotarod tests. In the rotarod test high speed and long latency are indicative of coordination and resistance. After 3 weeks of exposure, hematocrit levels were significantly decreased in the hyperoxia group compared to normoxic control littermates (%, mean ± SD, 37.8 ± 1.3, n = 15 vs. 49.9 ± 5.1, n = 15, p < 0.05). In the Y-maze test, chronic hyperoxic exposure resulted in a statistically significant decrease in alteration rate compared to normoxic control (%, mean ± SD, 53.4 ± 9.9, n = 30 vs. 61.2 ± 9.5, n = 15, p < 0.05). The rotarod and grip strength tests did not show statistically significant changes between the two groups. Our data suggest that chronic hyperoxia may lead to decreased cognitive performance in adult mice, which may be secondary to structural and functional changes in the brain.

Chronic Ketosis Modulates HIF1α-Mediated Inflammatory Response in Rat Brain.

Hypoxia inducible factor alpha (HIF1α) is associated with neuroprotection conferred by diet-induced ketosis but the underlying mechanism remains unclear. In this study we use a ketogenic diet in rodents to induce a metabolic state of chronic ketosis, as measured by elevated blood ketone bodies. Chronic ketosis correlates with neuroprotection in both aged and following focal cerebral ischaemia and reperfusion (via middle cerebral artery occlusion, MCAO) in mouse and rat models. Ketone bodies are known to be used efficiently by the brain and metabolism of ketone bodies is associated with increased cytosolic succinate levels that inhibits prolyl hydroxylases allowing HIF1α to accumulate. Ketosis also regulates inflammatory pathways, and HIF1α is reported to be essential for gene expression of interleukin10 (IL10). Therefore we hypothesised that ketosis-stabilised HIF1α modulates the expression of inflammatory cytokines orchestrating neuroprotection. To test changes in cytokine levels in rodent brain, eight-week-old rats were fed either the standard chow diet (SD) or the ketogenic (KG) diet for 4 weeks before ischaemia experiments (MCAO) were performed and the brain tissues were collected. Consistent with our hypothesis, immunoblotting analysis shows IL10 levels were significantly higher in KG diet rat brain compared to SD, whereas the TNFα and IL6 levels were significantly lower in the brains of KG diet fed group.

Experimental and observational evidence of negative conspecific density dependence in temperate ectomycorrhizal trees.

Conspecific negative density dependence (CNDD) promotes tree species diversity by reducing recruitment near conspecific adults due to biotic feedbacks from herbivores, pathogens, or competitors. While this process is well-described in tropical forests, tests of temperate tree species range from strong positive to strong negative density dependence. To explain this, several studies have suggested that tree species traits may help predict the strength and direction of density dependence: for example, ectomycorrhizal-associated tree species typically exhibit either positive or weaker negative conspecific density dependence. More generally, the strength of density dependence may be predictably related to other species-specific ecological attributes such as shade tolerance, or the relative local abundance of a species. To test the strength of density dependence and whether it affects seedling community diversity in a temperate forest, we tracked the survival of seedlings of three ectomycorrhizal-associated species experimentally planted beneath conspecific and heterospecific adults on the Prospect Hill tract of the Harvard Forest, in Massachusetts, USA. Experimental seedling survival was always lower under conspecific adults, which increased seedling community diversity in one of six treatments. We compared these results to evidence of CNDD from observed sapling survival patterns of 28 species over approximately 8 years in an adjacent 35-ha forest plot. We tested whether species-specific estimates of CNDD were associated with mycorrhizal association, shade tolerance, and local abundance. We found evidence of significant, negative conspecific density dependence (CNDD) in 23 of 28 species, and positive conspecific density dependence in two species. Contrary to our expectations, ectomycorrhizal-associated species generally exhibited stronger (e.g., more negative) CNDD than arbuscular mycorrhizal-associated species. CNDD was also stronger in more shade-tolerant species but was not associated with local abundance. Conspecific adult trees often have a negative influence on seedling survival in temperate forests, particularly for tree species with certain traits. Here we found strong experimental and observational evidence that ectomycorrhizal-associating species consistently exhibit CNDD. Moreover, similarities in the relative strength of density dependence from experiments and observations of sapling mortality suggest a mechanistic link between negative effects of conspecific adults on seedling and sapling survival and local tree species distributions.

Hypoxic Preconditioning Averts Sporadic Alzheimer's Disease-Like Phenotype in Rats: A Focus on Mitochondria.

Aims: Brief episodes of sublethal hypoxia reprogram brain response to face possible subsequent lethal stimuli by triggering adaptive and prosurvival events-a phenomenon denominated hypoxic preconditioning (HP). To date, the potential therapeutic implications of HP to forestall sporadic Alzheimer's disease (sAD) pathology remain unexplored. Using a well-established protocol of HP and focusing on hippocampus as a first brain region affected in AD, this study was undertaken to investigate the potential protective effects of HP in a sAD rat model induced by the intracerebroventricular (icv) administration of streptozotocin (STZ) and to uncover the mitochondrial adaptations underlying this nonpharmacological strategy. Results: HP prevented the memory and learning deficits as well as tau pathology in the icvSTZ rat model. HP also attenuated icvSTZ-related reactive astrogliosis, as noted by increased glial fibrillary acidic protein immunoreactivity and myo-inositol levels. Notably, HP abrogated the icvSTZ-related impaired energy metabolism and oxidative damage. Particularly, HP averted increased lactate, glutamate, and succinate levels, and decreased mitochondrial respiratory chain function and mitochondrial DNA content. Concerning mitochondrial adaptations underlying HP-triggered tolerance to icvSTZ, preconditioned hippocampal mitochondria displayed an enhanced complex II-energized mitochondrial respiration, which resulted from a coordinated interaction between mitochondrial biogenesis and fusion-fission. Mitochondrial biogenesis was stimulated immediately after HP, whereas in a latter phase mitochondrial fusion-fission events are modulated favoring the generation of elongated mitochondria. Innovation and Conclusion: Overall, these results demonstrate for the first time that HP prevents the sAD-like phenotype, in part, by targeting mitochondria emerging as a preventive strategy in the context of AD. Antioxid. Redox Signal. 37, 739-757.

Tree species diversity increases with conspecific negative density dependence across an elevation gradient.

Elevational and latitudinal gradients in species diversity may be mediated by biotic interactions that cause density-dependent effects of conspecifics on survival or growth to differ from effects of heterospecifics (i.e. conspecific density dependence), but limited evidence exists to support this. We tested the hypothesis that conspecific density dependence varies with elevation using over 40 years of data on tree survival and growth from 23 old-growth temperate forest stands across a 1,000-m elevation gradient. We found that conspecific-density-dependent effects on survival of small-to-intermediate-sized focal trees were negative in lower elevation, higher diversity forest stands typically characterised by warmer temperatures and greater relative humidity. Conspecific-density-dependent effects on survival were less negative in higher elevation stands and ridges than in lower elevation stands and valley bottoms for small-to-intermediate-sized trees, but were neutral for larger trees across elevations. Conspecific-density-dependent effects on growth were negative across all tree size classes and elevations. These findings reveal fundamental differences in biotic interactions that may contribute to relationships between species diversity, elevation and climate.

Arbuscular mycorrhizal trees influence the latitudinal beta-diversity gradient of tree communities in forests worldwide.

Arbuscular mycorrhizal (AM) and ectomycorrhizal (EcM) associations are critical for host-tree performance. However, how mycorrhizal associations correlate with the latitudinal tree beta-diversity remains untested. Using a global dataset of 45 forest plots representing 2,804,270 trees across 3840 species, we test how AM and EcM trees contribute to total beta-diversity and its components (turnover and nestedness) of all trees. We find AM rather than EcM trees predominantly contribute to decreasing total beta-diversity and turnover and increasing nestedness with increasing latitude, probably because wide distributions of EcM trees do not generate strong compositional differences among localities. Environmental variables, especially temperature and precipitation, are strongly correlated with beta-diversity patterns for both AM trees and all trees rather than EcM trees. Results support our hypotheses that latitudinal beta-diversity patterns and environmental effects on these patterns are highly dependent on mycorrhizal types. Our findings highlight the importance of AM-dominated forests for conserving global forest biodiversity.

Chronic Ketosis Modulates HIF1α-Mediated Inflammatory Response in Rat Brain.

Hypoxia inducible factor alpha (HIF1α) is associated with neuroprotection conferred by diet-induced ketosis, but the underlying mechanism remains unclear. In this study, we use a ketogenic diet in rodents to induce a metabolic state of chronic ketosis, as measured by elevated blood ketone bodies. Chronic ketosis correlates with neuroprotection in both aged and following focal cerebral ischemia and reperfusion (via middle cerebral artery occlusion, MCAO) in mouse and rat models. Ketone bodies are known to be used efficiently by the brain, and metabolism of ketone bodies is associated with increased cytosolic succinate levels that inhibits prolyl hydroxylases allowing HIF1α to accumulate. Ketosis also regulates inflammatory pathways, and HIF1α is reported to be essential for gene expression of interleukin 10 (IL10). Therefore, we hypothesized that ketosis-stabilized HIF1α modulates the expression of inflammatory cytokines orchestrating neuroprotection. To test changes in cytokine levels in rodent brain, 8-week-rats were fed either the standard chow diet (SD) or the KG diet for 4 weeks before ischemia experiments (MCAO) were performed and the brain tissues were collected. Consistent with our hypothesis, immunoblotting analysis shows IL10 levels were significantly higher in KG diet rat brain compared to SD, whereas the TNFα and IL6 levels were significantly lower in the brains of KG diet-fed group.

Altered Behavioral Performance in the Neuron-Specific HIF-1- and HIF-2-Deficient Mice Following Chronic Hypoxic Exposure.

Hypoxia-inducible factors (HIFs) are transcriptional regulators that mediate in mice for HIF-1 and HIF-2. The objective of this study was to investigate the effect of neuronal deletion of HIF-1 and HIF-2 in hypoxic adaptation by using the neuron-specific knockout (KO) mice. The floxed control and KO mice were used. Hypoxic mice were kept in a hypobaric chamber at a pressure of 300 torr (0.4 ATM, which was equivalent to 8% oxygen under normobaric condition) for 3 weeks. The littermate, normoxic control mice were housed in the same room next to the chamber to match ambient conditions. Body weights were monitored throughout the 3-week course. Cognitive function was measured using a Y-maze test; motor functions were measured using the rotarod test and the grip strength test. The hematocrit increased significantly at the end of 3-week hypoxic exposure in both control and KO mice. In the Y-maze test, the alternation rate (indicative of sustained cognition) trended lower in the KO mice compared to the controls following hypoxia (%, 51.3 ± 13.1, n = 6 vs. 63.2 ± 12.0, n = 8). In the rotarod test, the latency (seconds) in the KO mice was significantly lower compared to the controls (50.4 ± 5.7 vs. 77.1 ± 5.0, n = 3 each before hypoxia and 66.4 ± 3.4, n = 6 vs. 98.1 ± 15.4 after hypoxia, n = 3). The grip strength in the KO mice was similar compared to the control mice before hypoxia, but the strength of KO mice trended higher after hypoxic exposure. Our data suggest that deficiency of neuronal HIF-1 and HIF-2 may result in changes in behavioral performance and other adaptative responses to hypoxia.

Effect of 3-Day and 21-Day Hypoxic Preconditioning on Recovery Following Cerebral Ischemia in Rats.

We have previously reported that in a rat model of chronic hypoxia, HIF-1α and its target genes have significantly accumulated by 3 days of exposure, whereas no significant increase in capillary density has occurred; there is a significant increase in capillary density at 21 days of chronic hypoxic exposure. In this study we hypothesize that by utilizing 3 days and 21 days of hypoxic preconditioning, we would distinguish between the relative neuroprotective contributions of the accumulation of HIF-1α and its target genes and angiogenic adaptation in a rat middle cerebral artery occlusion (MCAO) model. Rats were randomly assigned to either hypoxic precondition groups (3-day and 21-day hypoxia) or normoxic control group. Hypoxic animals were kept in a hypobaric chamber at a constant pressure of 0.5 atmosphere (380 mmHg, equivalent to 10% normobaric oxygen at sea level) for either 3 or 21 days. Normoxic controls were housed in the same room next to the hypobaric chamber. Erythropoietin (EPO) was measured at 3 and 21 days of hypoxia using Western blotting analysis. Infarct volumes were measured following 24 hours of permanent MCAO. We found that EPO is upregulated at 3 days of hypoxia and returns to baseline by 21 days of hypoxia. The infarct volumes following 24-hour MCAO were significantly reduced with 3-day hypoxic preconditioning when compared to normoxic controls (%, 31.8 ± 5, n = 9 vs. 50.1 ± 10.9, n = 7). No significant differences in infarct volume were seen between the normoxic controls and 21-day hypoxic preconditioned rats. We have shown that a 3-day hypoxic preconditioning, but not 21-day hypoxic preconditioning, provides significant neuroprotection against focal ischemia in rats, supporting a larger role for the accumulations of HIF-1α and upregulation of its target genes in the neuroprotection against focal ischemia.

Environmental Enrichment Improved Cognitive Performance in Mice under Normoxia and Hypoxia.

The mammalian brain modulates its microvascular network to accommodate tissue energy demand in a process referred to as angioplasticity. There is an aging effect on cognitive function and adaptive responses to hypoxia. Hypoxia-induced angiogenesis is delayed in the aging mouse brain. Additionally, it has been shown that environmental enrichment provides an environment that fosters increased physical activity and sensory stimulation for mice as compared to standard housing; this stimulation increases neuronal activity and consequently brain oxygen demand. In this study, we investigated the effect of environmental enrichment and chronic hypoxia on cognitive performance in the young (2-4 months old) and the aged mice (17-21 months old). Mice were placed in a non-enriched or an enriched environment for 4 weeks under normoxia followed by 3 weeks of hypobaric hypoxia (~0.4 atm, equivalent to 8% normobaric oxygen at sea level). Cognitive function was evaluated using the Y-maze and the novel object recognition tests in the enriched or non-enriched mice under normoxic or hypoxic conditions. In Y-maze, a high alternation rate is indicative of sustained cognition as the animals must remember which arm was entered last, so as not to re-enter it. Novel object recognition is based on the natural tendency of rodents to investigate a novel object instead of a familiar one; a higher novel object exploration rate is indicative of better cognitive function. The young mice showed a significantly higher alternation rate (%, 63 ± 7 vs. 48 ± 10, n = 8 and 10, respectively) in the Y-Maze test as compared to the aged mice. Under normoxia, the enriched mice showed an improved alternation rate (%, 63 ± 10, n = 10) in Y-Maze test and a higher novel object exploration rate (%, 68 ± 10 vs. 52 ± 10) compared to the non-enriched controls. Similar results were observed for both young and aged mice following hypoxic exposure. Our data suggests that environmental enrichment improved the cognitive performance in the young and aged mice under normoxic and hypoxic conditions.

Neurovascular and cortical responses to hyperoxia: enhanced cognition and electroencephalographic activity despite reduced perfusion.

KEY POINTS: Extreme aviation is accompanied by ever-present risks of hypobaric hypoxia and decompression sickness. Neuroprotection against those hazards is conferred through fractional inspired oxygen ( FI,O2 ) concentrations of 60-100% (hyperoxia). Hyperoxia reduces global cerebral perfusion (gCBF), increases reactive oxygen species within the brain and leads to cell death within the hippocampus. However, an understanding of hyperoxia's effect on cortical activity and concomitant levels of cognitive performance is lacking. This limits our understanding of whether hyperoxia could lower the brain's threshold of tolerance to physiological stressors inherent to extreme aviation, such as high gravitational forces. This study aimed to quantify the impact of hyperoxia upon global cerebral perfusion (gCBF), cognitive performance and cortical electroencephalography (EEG). Hyperoxia evoked a rapid reduction in gCBF, yet cognitive performance and vigilance were enhanced. EEG measurements revealed enhanced alpha power, suggesting less desynchrony, within the cortical temporal regions. Collectively, this work suggests hyperoxia-induced brain hypoperfusion is accompanied by enhanced cognitive processing and cortical arousal. ABSTRACT: Extreme aviators continually inspire hyperoxic gas to mitigate risk of hypoxia and decompression injury. This neuroprotection carries a physiological cost: reduced cerebral perfusion (CBF). As reduced CBF may increase vulnerability to ever-present physiological challenges during extreme aviation, we defined the magnitude and duration of hyperoxia-induced changes in CBF, cortical electrical activity and cognition in 30 healthy males and females. Magnetic resonance imaging with pulsed arterial spin labelling provided serial measurements of global CBF (gCBF), first during exposure to 21% inspired oxygen ( FI,O2 ) followed by a 30-min exposure to 100% FI,O2 . High-density EEG facilitated characterization of cortical activity during assessment of cognitive performance, also measured during exposure to 21% and 100% FI,O2 . Acid-base physiology was measured with arterial blood gases. We found that exposure to 100% FI,O2 reduced gCBF to 63% of baseline values across all participants. Cognitive performance testing at 21% FI,O2 was accompanied by increased theta and beta power with decreased alpha power across multiple cortical areas. During cognitive testing at 100% FI,O2 , alpha activity was less desynchronized within the temporal regions than at 21% FI,O2 . The collective hyperoxia-induced changes in gCBF, cognitive performance and EEG were similar across observed partial pressures of arterial oxygen ( PaO2 ), which ranged between 276-548 mmHg, and partial pressures of arterial carbon dioxide ( PaCO2 ), which ranged between 34-50 mmHg. Sex did not influence gCBF response to 100% FI,O2 . Our findings suggest hyperoxia-induced reductions in gCBF evoke enhanced levels of cortical arousal and cognitive processing, similar to those occurring during a perceived threat.

Local species diversity, ß-diversity and climate influence the regional stability of bird biomass across North America.

Biodiversity often stabilizes aggregate ecosystem properties (e.g. biomass) at small spatial scales. However, the importance of species diversity within communities and variation in species composition among communities (ß-diversity) for stability at larger scales remains unclear. Using a continental-scale analysis of 1657 North American breeding-bird communities spanning 20-years and 35 ecoregions, we show local species diversity and ß-diversity influence two components of regional stability: local stability (stability of bird biomass within sites) and spatial asynchrony (asynchronous fluctuations in biomass among sites). We found spatial asynchrony explained three times more variation in regional stability of bird biomass than did local stability. This result contrasts with studies at smaller spatial scales-typically plant metacommunities under 1 ha-that find local stability to be more important than spatial asynchrony. Moreover, spatial asynchrony of bird biomass increased with bird ß-diversity and climate heterogeneity (temperature and precipitation), while local stability increased with species diversity. Our study reveals new insights into the scale-dependent processes regulating ecosystem stability, providing evidence that both local biodiversity loss and homogenization can destabilize ecosystem processes at biogeographic scales.

Increased cerebral vascularization and decreased water exchange across the blood-brain barrier in aquaporin-4 knockout mice.

Aquaporin-4 (AQP4) plays an important role in regulating water exchange across the blood-brain barrier (BBB) and brain-cerebrospinal fluid interface. Studies on AQP-4 knockout mice (AQP4-KO) have reported considerable protection from brain edema induced by acute water intoxication and ischemic stroke, identifying AQP4 as a potential target for therapeutic interventions. However, the long-term effects of chronic AQP4 suppression are yet to be elucidated. In the current study, we evaluated the physiological and structural changes in adult AQP4-KO mice using magnetic resonance imaging (MRI) and immunohistochemical analysis. Water exchange across BBB was assessed by tracking an intravenous bolus injection of oxygen-17 (17O) water (H217O) using 17O-MRI. Cerebral blood flow (CBF) was quantified using arterial spin-labeling (ASL) MRI. Capillary density was determined by immunohistochemical staining for glucose transporter-1 (GLUT1). Compared to wildtype control mice, AQP4-KO mice showed a significant reduction in peak and steady-state H217O uptake despite unaltered CBF. Interestingly, a 22% increase in cortical capillary density was observed in AQP4-KO mice. These results suggest that increased cerebral vascularization may be an adaptive response to chronic reduction in water exchange across BBB in AQP4-KO mice.

Functionalized Phenylbenzamides Inhibit Aquaporin-4 Reducing Cerebral Edema and Improving Outcome in Two Models of CNS Injury.

Cerebral edema in ischemic stroke can lead to increased intracranial pressure, reduced cerebral blood flow and neuronal death. Unfortunately, current therapies for cerebral edema are either ineffective or highly invasive. During the development of cytotoxic and subsequent ionic cerebral edema water enters the brain by moving across an intact blood brain barrier and through aquaporin-4 (AQP4) at astrocyte endfeet. Using AQP4-expressing cells, we screened small molecule libraries for inhibitors that reduce AQP4-mediated water permeability. Additional functional assays were used to validate AQP4 inhibition and identified a promising structural series for medicinal chemistry. These efforts improved potency and revealed a compound we designated AER-270, N-[3,5-bis (trifluoromethyl)phenyl]-5-chloro-2-hydroxybenzamide. AER-270 and a prodrug with enhanced solubility, AER-271 2-{[3,5-Bis(trifluoromethyl) phenyl]carbamoyl}-4-chlorophenyl dihydrogen phosphate, improved neurological outcome and reduced swelling in two models of CNS injury complicated by cerebral edema: water intoxication and ischemic stroke modeled by middle cerebral artery occlusion.

Cerebral Angioplasticity: The Anatomical Contribution to Ensuring Appropriate Oxygen Transport to Brain.

In order to maintain proper function, mammalian brain requires a significant fraction of the energy provided through whole body oxygen consumption and oxidative phosphorylation. This has been fairly well known for a long time. More recently there has been an increased appreciation that, while whole brain blood flow remains fairly constant, there are large regional changes in local blood flow to account for spatial and temporal heterogeneity of neuronal activity. This latter phenomenon requires an extensive regulatory system for local oxygen delivery that involves arteriolar and capillary control mechanisms. The ISOTT has been a major contributor to the study of oxygen supply and demand through studies of the mechanisms of vascular dilation and constriction in response to energy expenditure and availability of substrate and oxygen. Nevertheless, it has become clear in the past few decades that in addition to acute, physiological responses to energy demand and oxygen/substrate availability, there are regulatory mechanisms that are continuously operating to control the capillary distribution over a time course of weeks. This process of "angioplasticity" results in the gradual acclimatization of the brain capillary bed to prolonged changes in oxygen/substrate availability and/or neuronal activity patterns. Angioplasticity is primarily regulated through the hypoxia inducible transcription factor, acting as a detector of the balance between oxygen delivery and energy demand at the level of the cell redox state, controlling vascular endothelial growth factor production which helps determine capillary density in consort with the cyclooxygenase-2/angiopoietin-2 pathway that controls endothelial cell junction mechanical stability. We can conclude that the structure-function of brain capillaries is regulated during prolonged challenges to energy supply-demand balance within the physiological range. We can conclude that over the physiological range of ambient oxygen, brain capillary density is proportional to fraction inspired oxygen. The primary mechanisms for regulation of brain capillary density are HIF-1/VEGF and COX-2/PGE2/ang-2 pathways of angiogenesis and angiolysis.