Days Of Intermittent Hypoxia Research Articles

Abstract 3004: Intermittent Hypoxia Decreases Aortic Stiffness And Facilitates Abdominal Aortic Aneurysm

Introduction: Intermittent hypoxia (IH) is a phenomenon observed with obstructive sleep apnea (OSA), which is highly prevalent in patients with abdominal aortic aneurysm (AAA). However, it remains unknown if IH is a risk factor for the development of AAA. The goal of this study was to determine the effects of IH exposure on arterial stiffness, and to assess whether IH increases the susceptibility to angiotensin (Ang) II-induced AAA in mice. Methods: Male C57BL/6 mice (8-week-old) were subjected to IH (6.5%-21% O 2 , 90 s/cycle, 12 h/day) or room air (RA, 21% O 2 , 24 h/day) for short-term (6 weeks) or long-term (6 months) (n = 10/group). Short-term mice were examined for arterial stiffness by pulse wave velocity (PWV) and atomic force microscopy, while long-term mice were used for AAA studies. To induce AAA, mice were infused with Ang II (1 μg/kg/min) using osmotic minipumps during final 14 days of IH or RA exposure. Sham surgery was performed in control mice. Outcomes were assessed via transabdominal ultrasound imaging and histology. Results: Short-term IH exposure decreased the stiffness of the abdominal (0.88 vs. 2.32 m/s in RA; p=0.0006 ) and thoracic (2.0 vs. 3.5 kPa in RA; p=0.01 ) aorta. Ang II increased abdominal aortic diameter in RA mice compared to sham and long-term IH with Ang II further augmented the aortic diameter (1.56 vs. 1.34 mm in RA Ang II; p=0.03 ) and incidence of AAA (50 vs. 10% in RA Ang II; p=0.05 ). Histologically, long-term IH plus Ang II augmented medial elastin fragmentation (7.7 vs. 5.3 in RA Ang II; p=0.04 ) and aortic collagen content (3.1 vs. 1.3 in RA Ang II; p=0.06 ). Ang II tended to non-significantly increase PWV in both groups (RA sham 1.9 vs. RA Ang II 2.5 and IH Ang II 2.3 m/s). Notably, a negative correlation between maximum aortic diameter and PWV was observed in AAA-induced mice (R 2 =0.48, p=0.0009 ). Conclusions: IH favors AAA development in association with changes in aortic stiffness. These findings support a potential causal link between OSA and AAA pathogenesis. Clarification of the molecular mechanisms may reveal novel therapeutic targets for the management of AAA.

Early Increase in Blood-Brain Barrier Permeability in a Murine Model Exposed to Fifteen Days of Intermittent Hypoxia.

Obstructive sleep apnea (OSA) is characterized by intermittent repeated episodes of hypoxia-reoxygenation. OSA is associated with cerebrovascular consequences. An enhanced blood-brain barrier (BBB) permeability has been proposed as a marker of those disorders. We studied in mice the effects of 1 day and 15 days intermittent hypoxia (IH) exposure on BBB function. We focused on the dorsal part of the hippocampus and attempted to identify the molecular mechanisms by combining in vivo BBB permeability (Evans blue tests) and mRNA expression of several junction proteins (zona occludens (ZO-1,2,3), VE-cadherin, claudins (1,5,12), cingulin) and of aquaporins (1,4,9) on hippocampal brain tissues. After 15 days of IH exposure we observed an increase in BBB permeability, associated with increased mRNA expressions of claudins 1 and 12, aquaporins 1 and 9. IH seemed to increase early for claudin-1 mRNA expression as it doubled with 1 day of exposure and returned near to its base level after 15 days. Claudin-1 overexpression may represent an immediate response to IH exposure. Then, after 15 days of exposure, an increase in functional BBB permeability was associated with enhanced expression of aquaporin. These BBB alterations are possibly associated with a vasogenic oedema that may affect brain functions and accelerate neurodegenerative processes.

Caloric restriction prevents obesity- and intermittent hypoxia-induced cardiac remodeling in leptin-deficient ob/ob mice.

Background: Intermittent hypoxia (IH), a key characteristic of obstructive sleep apnea, is independently associated with cardiometabolic impairment. While endogenous leptin levels may provide cardioprotective effects against hypoxia, leptin resistance is common among obese individuals presenting with obstructive sleep apnea. Methods: Here, we assessed left ventricle (LV) function using M-mode echocardiography in lean wild-type, calorically-restricted ob/ob, and obese ob/ob mice before and after 6 days of IH to determine how obesity and intermittent hypoxia interact to affect cardiac function independent of leptin signaling. Results: Calorically-restricting ob/ob mice for 4 weeks prior to IH exposure prevented weight gain (−2.1 ± 1.4 g) compared to free-fed ob/ob mice (8.7 ± 1.1 g). Free-fed ob/ob mice exhibited increased LV mass (0.713 ± 0.008 g) relative to wild-type mice (0.685 ± 0.004 g) and increased posterior wall thickness (0.089 ± 0.006 cm) relative to calorically-restricted ob/ob mice (0.072 ± 0.004 cm). Following 6 days of IH, free-fed ob/ob mice exhibited increases in cardiac output (44.81 ± 2.97 pre-IH vs. 57.14 ± 3.09 ml/min post-IH), LV diameter (0.400 ± 0.007 pre-IH vs. 0.428 ± 0.009 cm post-IH) and end diastolic volume (0.160 ± 0.007 pre-IH vs. 0.195 ± 0.012 ml post-IH) that were not detected in wild-type or calorically-restricted ob/ob mice. Conclusion: Caloric restriction can prevent obesity-induced LV hypertrophy and protect against acute IH-induced cardiac remodeling independent of leptin signaling. These findings may have clinical implications for obstructive sleep apnea.

0033 Dysregulated sleep and NREM sleep electroencephalogram delta power induced by intermittent hypoxia exposure are attenuated in NLRP3 knockout mice

Abstract Introduction The nucleotide-binding domain leucine rich family pyrin containing 3 (NLRP3) inflammasome protein complex activates caspase-1 to convert the pro-forms of IL-18 and IL-1 beta (IL-1β) into their active forms and is involved in homeostatic sleep and sleep responses to pathogenic stimuli. Intermittent hypoxia (IH) is a hallmark of sleep disordered breathing (SDB) and both IL-18 receptors are associated with SDB in humans. Thus, we hypothesized that NLRP3 inflammasomes are involved in SDB-related sleep disturbances. Methods Sleep architecture was assessed by polysomnography in NLRP3 knockout (KO) and wild-type (WT) mice (N = 8 per group). A gas exchange mixer delivered house air serving as a control or chronic IH that involved 90 second episodic oxygen reductions that consisted of ambient oxygen (21%) with brief hypoxic conditions (~10%) that lasted for 3 seconds for the first 10 h of the light period for 5 consecutive days. Gene expression and protein levels in the brain and lungs were assayed using real-time polymerase chain reaction and enzyme-linked immunosorbent assays, respectively. Results In WT mice, significant increased non-rapid-eye movement (NREM) sleep amounts and NREM sleep electroencephalogram delta power (0.5-4 Hz) were found after 1 day of IH compared to control conditions (p < 0.001). After 5 days of IH, WT mice showed a significant attenuation in NREM sleep amounts and NREM sleep delta power (p < 0.001) and increased wake bout frequency (p = 0.006) when compared to control conditions. However, the IH-induced NREM sleep and NREM sleep delta power enhancements and reductions were attenuated (21-35%) in NLRP3 KO mice compared to WT mice. In WT mice, NLRP3, IL-1β, IL-18 and caspase-1 gene expression, IL-1β and IL-18 protein levels, and caspase-1 activity were significantly increased in the somatosensory cortices, NTS, and lungs after both 1 and 5 days of IH when compared to control conditions (p < 0.05 for all), although NLRP3 KO mice did not exhibit significant differences in molecules downstream of NLRP3 inflammasome activation. Conclusion Our findings indicate that altered NLRP3 inflammasome activation contributes to dysregulated sleep occurring from IH and likely is involved in sleep disturbances in SDB. Support (If Any) VA Career Development Aware IBX002823, VA Merit BX004500, NIH/NIMH 1R21MH125242, NIH/NHLBI R03 HL154284, and NIH/NHLBI R35 HL135818. JTM received partial salary compensation and funding from Merck Investigator Sponsored Programs but has no conflict of interest with this work.

Intermittent Hypoxia (IH) Impairs Hippocampal Oxygen Consumption and Neurophysiological Responses to Metabolic Challenge

IH is a consequence of several clinical conditions such as sleep apnea. Previous work has shown that IH impairs many neurophysiological functions such as hippocampal adult neurogenesis and NMDAr‐dependent synaptic plasticity. While IH produces hippocampal oxidative stress, IH‐dependent changes in hippocampal metabolism may also contribute to impaired neurophysiology. The objective of this study is to understand how IH impacts hippocampal physiology in response to metabolic challenge. We hypothesize that ten days of IH perturbs hippocampal synaptic transmission when metabolically challenged. In vitro recordings of field excitatory post‐synaptic potentials (fEPSPs) from hippocampal slices showed that glucose deprivation (i.e., 30mM Fructose aCSF with 95% O2) caused a complete loss of the fEPSP within 20min; whereas, the fEPSP in control tissue was reduced to 40% of baseline within the same period of time. This difference coincided with a reduction in both maximal and ATP‐linked O2 consumption rates (OCRs) in hippocampal tissue following IH. Despite the reductions in hippocampal OCRs, neither ATP/ADP ratio nor lactate concentrations were different between the two groups. These data indicate that while ATP/ADP levels following IH are sufficiently maintained under basal conditions, metabolic processes fail to support synaptic physiology during glucose deprivation. Our ongoing experiments will further delineate how IH impacts the contribution of glycolytic and oxidative metabolism in the hippocampus. These findings may better define how IH impacts hippocampal responses to metabolic stressors, which could contribute to impairing hippocampal physiology in conditions such as sleep apnea.

Intermittent Hypoxia causes targeted disruption to NMDA receptor dependent synaptic plasticity in area CA1 of the hippocampus

Changed NMDA receptor (NMDAr) physiology is implicated with cognitive deficit resulting from conditions ranging from normal aging to neurological disease. Using intermittent hypoxia (IH) to experimentally model untreated sleep apnea, a clinical condition whose comorbidities include neurocognitive impairment, we recently demonstrated that IH causes a pro-oxidant condition that contributes to deficits in spatial memory and in NMDAr-dependent long-term potentiation (LTP). However, the impact of IH on additional forms of synaptic plasticity remains ill-defined. Here we show that IH prevents the induction of NMDAr-dependent LTP and long-term depression (LTD) in hippocampal brain slices from mice exposed to ten days of IH (IH10) yet spares NMDAr-independent forms of synaptic plasticity. Deficits in synaptic plasticity were accompanied by a reduction in hippocampal GluN1 expression. Acute manipulation of redox state using the reducing agent, Dithiothreitol (DTT) stimulated the NMDAr-dependent fEPSP following IH10. However, acute use of either DTT or MnTMPyP did not restore NMDAr-dependent synaptic plasticity after IH10 or prevent the IH-dependent reduction in GluN1, the obligatory subunit of the NMDAr. In contrast, MnTMPyP during IH10 (10-MnTMPyP), prevented the suppressive effects of IH on both NMDAr-dependent synaptic plasticity and GluN1 expression. These findings indicate that while the IH-dependent pro-oxidant state causes reversible oxidative neuromodulation of NMDAr activity, acute manipulation of redox state is ineffective in rescuing two key effects of IH related to the NMDAr within the hippocampus. These IH-dependent changes associated with the NMDAr may be a primary avenue by which IH enhances the vulnerability to impaired learning and memory when sleep apnea is left untreated in normal aging and in disease.

Histone Deacetylase 5 Is an Early Epigenetic Regulator of Intermittent Hypoxia Induced Sympathetic Nerve Activation and Blood Pressure.

Intermittent hypoxia (IH) is a hallmark manifestation of obstructive sleep apnea (OSA). Long term IH (LT-IH) triggers epigenetic reprogramming of the redox state involving DNA hypermethylation in the carotid body chemo reflex pathway resulting in persistent sympathetic activation and hypertension. Present study examined whether IH also activates epigenetic mechanism(s) other than DNA methylation. Histone modification by lysine acetylation is another major epigenetic mechanism associated with gene regulation. Equilibrium between the activities of histone acetyltransferases (HATs) and histone deacetylases (HDACs) determine the level of lysine acetylation. Here we report that exposure of rat pheochromocytoma (PC)-12 cells to IH in vitro exhibited reduced HDAC enzyme activity due to proteasomal degradation of HDAC3 and HDAC5 proteins. Mechanistic investigations showed that IH-evoked decrease in HDAC activity increases lysine acetylation of α subunit of hypoxia inducible factor (HIF)-1α as well as Histone (H3) protein resulting in increased HIF-1 transcriptional activity. Trichostatin A (TSA), an inhibitor of HDACs, mimicked the effects of IH. Studies on rats treated with 10 days of IH or TSA showed reduced HDAC activity, HDAC5 protein, and increased HIF-1 dependent NADPH oxidase (NOX)-4 transcription in adrenal medullae (AM) resulting in elevated plasma catecholamines and blood pressure. Likewise, heme oxygenase (HO)-2 null mice, which exhibit IH because of high incidence of spontaneous apneas (apnea index 72 ± 1.2 apnea/h), also showed decreased HDAC activity and HDAC5 protein in the AM along with elevated circulating norepinephrine levels. These findings demonstrate that lysine acetylation of histone and non-histone proteins is an early epigenetic mechanism associated with sympathetic nerve activation and hypertension in rodent models of IH.

Metformin improves infarct size in mice exposed to chronic intermittent hypoxia, a major feature of obstructive sleep apnea

Chronic intermittent hypoxia (IH), the major feature of obstructive sleep apnea (OSA) increases myocardial infarct size. Recent data demonstrate that IH-induced cardiomyocyte susceptibility to infarction could be explained by mitochondrial function and/or dynamics alterations. Moreover, AMP-activated protein kinase (AMPK), a major protein involved in cardiomyocyte metabolism is activated by IH in striated skeletal muscle. However, we recently found that AMPK failed to be activated in myocardium from mice exposed to IH. We aimed at demonstrating whether AMPK activation can be beneficial for ischemia-reperfusion injury following chronic IH exposure. Mice were exposed to 21 days of IH (21-5% FiO 2 , 60 s-cycles, 8 h/day), or normoxia (N) and treated or not with metformin (300 mg.kg −1 .day −1 ). Infarct size was measured after in vivo ischemia (45 min) and reperfusion (90 min). Mitochondrial function was assessed using oxygraphy and spectrofluorimetry. Mitochondrial dynamics and AMPK activation were assessed by Western blot. An insulin tolerance test was also performed. Metformin abolished the IH-induced increase in infarct size whereas it did not improve systemic insulin sensitivity. Metformin activated AMPK and did not impact the decrease in mitochondrial respiration (O 2 maximal consumption in complex I and II and respiratory control ratio). However, AMPK activation by Metformin seems to reestablish the distinctive IH impact on mitochondrial dynamics (fission and mitophagy). Metformin treatment is protective against ischemia-reperfusion injury induced by IH only. This suggests that metformin could be an alternative treatment to the currently restrictive one for apneic patients with a high CV risk. Ongoing studies will aim to better characterize the mechanisms by which metformin specifically decreases infarct size under IH.

Hif1a hemizygosity in vGlut1+ Cells mitigates the cellular and neurophysiological of Intermittent Hypoxia (IH)‐Dependent changes in the hippocampus

Arias et al. (eNeuro, 2020) recently demonstrated that intermittent hypoxia (IH), a principal outcome of sleep apnea, increases hippocampal oxidative stress that in turn, contributes to IH‑mediated deficits in spatial memory and NMDAr-dependent LTP. These phenomena were mitigated in hemizygous HIF1a mice indicating that IH-dependent HIF1a signaling plays a central role in mediating neurophysiological changes associated with impaired spatial memory. However, it remains unclear where such signaling originates. As glutamatergic synaptic communication in the hippocampus and cortex is critical for learning and memory, we sought to determine whether hemizygosity of HIF1a among glutamatergic neurons could mediate the impact of IH on hippocampal synaptic plasticity and spatial memory. To address this, we generated Vglut1-HIF1a+/- mice using Cre/LoxP recombination technology. Vglut1‑HIF1a+/- mice were exposed to either room air (control) or ten days of IH (IH10). In the hippocampus of wild-type mice, IH10 decreased expression of the obligatory NMDAr subunit, GluN1. This correlated with deficient induction of NMDAr-dependent synaptic plasticity. IH10 also increased hippocampal oxidative stress and caused a two-fold increase in hippocampal expression of the pro-oxidant enzyme, NADPH oxidase 4 (NOX4). In contrast to the effects in wild type mice, IH10 neither increased oxidative stress nor increased NOX4 expression in the hippocampus of Vglut1-HIF1a+/- mice. IH10 did not suppress GluN1 expression or impair hippocampal NMDAr-dependent synaptic plasticity in Vglut1-HIF1a+/- mice. In conclusion, our findings show that hemizygosity of HIF1a in vGlut1+ cells mitigate IH-dependent changes to hippocampal synaptic plasticity and protein expression. Thus, HIF1a signaling in vGlut1+ glutamatergic neurons appear to be a significant contributor for perturbing NMDAr-dependent synaptic plasticity and the molecular profile of the hippocampus following IH.

Assessment of mitochondrial and non‐mitochondrial O 2 consumption using in hippocampal tissue exposed to intermittent hypoxia

Intermittent Hypoxia (IH) is a well-recognized trait of untreated sleep apnea. We have recently demonstrated that mitigating oxidative stress induced by IH prevents impairments to hippocampal NMDAr-dependent LTP, hippocampal adult neurogenesis, spatial learning and memory. As these processes are also dependent on metabolic activity, the changes of metabolism caused by IH could also contribute to the impairment of synaptic plasticity and adult neurogenesis. The objective of this on-going study is to assess mitochondrial respiration in hippocampal tissue after ten days of IH (IH10). We assessed hippocampal mitochondrial respiration using Seahorse XF24 Extracellular Flux Analyzer technology. The tissue biopsies (Diameter: 1mm, thickness: 350 μm) were prepared from hippocampal brain slices originating from control mice or mice exposed to IH10. Preliminary experiments indicate that the basal O2 consumption rate (OCR) tended to decrease in IH10 (control: 202 ± 15 pmol/min, n=8 vs. IH: 126 ± 39 pmol/min n=4, P=0.05) and maximal OCR induced by FCCP was decreased in IH10 (control: 278 ± 29 pmol/min vs. IH10: 126 ± 39 pmol/min, P<0.05). These findings suggest IH impaired mitochondrial function in the hippocampus. ATP-linked OCR induced by oligomycin and proton leak were not altered by IH10. Interestingly, the proportion of non‑mitochondrial OCR in control was smaller than IH10 (control: 13 ± 1 % vs. IH10: 20 ± 5 %, P<0.05). Such a difference in non-mitochondrial OCR may reflect a state favoring enhanced reactive oxygen species production independent of mitochondrial origin. In conclusion, our study provides greater insights into the molecular basis by which untreated sleep apnea accelerates neurodegeneration in conditions such as Alzheimer's disease.

IH‐dependent HIF1a signaling upregulates reactive oxygen species signaling to suppress expression of the hippocampal NMDA receptor by downregulating the NR1 subunit.

When left untreated, the periodic cessation in breathing occurring with sleep apnea can cause oscillations in blood oxygenation leading to intermittent hypoxia (IH) and disrupted cognitive function. IH causes oxidative stress in the nervous system and impair spatial learning and memory. This study examines how IH influences synaptic physiology of principal neurons in the hippocampus. We hypothesize that IH‐dependent HIF1a signaling remodels the glutamatergic synapse by upregulating reactive oxygen species (ROS), weakening N‐methyl‐D‐aspartate receptor (NMDAR)‐dependent long‐term potentiation (LTP), and impairing cognitive performance on hippocampal based behavior. Wildtype mice (WT) and mice hemizygous for HIF1a (HIF1a+/−) were exposed to ten days of IH. While spatial memory in the Barnes maze was impaired in WT following IH (n=6), performance in the Barnes maze was unaffected by IH among HIF1a+/− (n=6). Electrophysiological recordings in brain slices from WT (n=6) and HIF1a+/−(n=6) exposed to IH also revealed that NMDAR‐dependent LTP was suppressed only in WT slices. While IH caused a down regulation of the NR1 subunit from WT, protein carbonylation and HIF1a regulated oxidase, NOX4, was increased by IH in the WT exposed to IH. No changes in NR1, protein carbonylation, or NOX4 were observed in HIF1a+/− exposed to IH. Furthermore, the administration of the superoxide anion scavenger MnTMPyP (5 mg/Kg i.p) during IH mitigated the downregulation of NR1 and suppression of NMDAR‐dependent LTP in WT. These results show that IH‐dependent HIF1a signaling causes targeted disruption to NMDAR‐dependent LTP through a ROS‐based mechanism that involves the downregulation of NR1 subunit. We provide new insight into the mechanistic basis by which sleep apnea may disrupt cognitive performance.Support or Funding InformationSupported by NIH Grant R01‐NS‐1074210100

Duration‐dependent effects of IH‐stimulated Hif1a signaling on adult hippocampal neurogenesis

Intermittent hypoxia (IH), is a hallmark consequence of sleep apnea, a common untreated disorder associated with impaired neurological health. We have recently demonstrated that thirty days of IH impairs long term potentiation and reduced the proportion of neurons generated during adult neurogenesis. Hypoxia Inducible Factor 1a (HIF1a) is a transcription factor important to stem cell development that is regulated by the state of oxygenation. The objective of this on‐going study is to characterize how the duration of IH exposure influences the generation of adult‐born neurons and to determine the role of IH‐dependent HIF1a signaling in the generation of adult‐born neurons in the hippocampus. We hypothesize that IH has duration dependent influence over neurons generated through adult neurogenesis and involves HIF1a dependent signaling. We used a cre‐loxP transgenic strategy to birth label a discrete cohort of stem cells that experience room air for 30 days (control), 10 days of IH +20 days of recovery in room air (IH10+20RA), or 30 days of IH (IH30) in wild‐type reporter mice (Ai27) and in Ai27‐HIF1a+/flox mice. In wild‐type mice, exposure to IH10+20RA led to higher proportion of birth‐labelled neurons compared to the control (+19% ± 5%, n=6) while the proportion of neurons following IH30 decreased (−40% ± 10%, n=4). In Ai27‐HIF1a+/− mice, the proportion of birth‐labeled neurons was similar between control and IH10+20RA groups. However, following IH30 the proportion of birth‐labeled neurons was markedly decreased when compared to control in Ai27‐Hif1a+/− and to the IH30 wild‐type group. Dendritic complexity, as determined by Sholl analysis, was similar among labeled neurons in both wild‐type and Ai27‐HIF1a+/− groups suggesting that neither IH nor IH‐dependent HIF1a signaling influences dendritic complexity. These findings indicate that exposure duration is factor impacting how IH affects the generation of adult‐born neurons through a HIF1a signaling shaping how IH influences, associated with sleep apnea affects the generation of adult‐born neurons.Support or Funding InformationThis work was supported by NIH R01 NS10742101 (AJG).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Hippocampal neural stem cells undergo fate switch following intermittent hypoxia

Adult neurogenesis is a phenomenon widely recognized as a process supporting neurophysiology of the hippocampus. We recently reported that 30 days of intermittent hypoxia (IH), a principal consequence for sleep apnea, stimulates the expansion of hippocampal neural stem cells (NSCs), yet suppresses the generation of adult‐born neurons. This study investigates how IH impacts NSCs by using a shorter duration of IH. We hypothesize that 10 days of IH (IH10) redirects the course of development of NSCs. We examined how NSCs in the neurogenic niche of the dentate gyrus are affected by IH10. Immunohistochemical analysis revealed that the number of NSCs (Sox2+ cells) was not increased following IH10, yet appears to increase in quiescent NSCs (i.e., Sox2+ cells that co‐express the astrocytic marker, S100b, and do not develop into adult‐born neurons). Fewer NSCs were observed transitioning into intermediate neuronal progenitors (INPs, SOX2+/TBR2+ cells) and a reduction in the total number of INPs (TBR2+ cells) was also noted. These experiments suggest that the impact of IH on cells involved with adult neurogenesis is cell‐type specific. Our findings provide further support that untreated sleep apnea may impact the fate of pluripotent cellular constituents in the central nervous system.Support or Funding InformationThis work was supported by NIH R01 NS10742101 (AJG)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Angiotensin type 1a receptors in the median preoptic nucleus support intermittent hypoxia-induced hypertension.

Chronic intermittent hypoxia (CIH) is a model of the hypoxemia from sleep apnea that causes a sustained increase in blood pressure. Inhibition of the central renin-angiotensin system or FosB in the median preoptic nucleus (MnPO) prevents the sustained hypertensive response to CIH. We tested the hypothesis that angiotensin type 1a (AT1a) receptors in the MnPO, which are upregulated by CIH, contribute to this hypertension. In preliminary experiments, retrograde tract tracing studies showed AT1a receptor expression in MnPO neurons projecting to the paraventricular nucleus. Adult male rats were exposed to 7 days of intermittent hypoxia (cycling between 21% and 10% O2 every 6 min, 8 h/day during light phase). Seven days of CIH was associated with a FosB-dependent increase in AT1a receptor mRNA without changes in the permeability of the blood-brain barrier in the MnPO. Separate groups of rats were injected in the MnPO with an adeno-associated virus containing short hairpin (sh)RNA against AT1a receptors to test their role in intermittent hypoxia hypertension. Injections of shRNA against AT1a in MnPO blocked the increase in mRNA associated with CIH, prevented the sustained component of the hypertension during normoxia, and reduced circulating advanced oxidation protein products, an indicator of oxidative stress. Rats injected with shRNA against AT1a and exposed to CIH had less FosB staining in MnPO and the rostral ventrolateral medulla after intermittent hypoxia than rats injected with the control vector that were exposed to CIH. Our results indicate AT1a receptors in the MnPO contribute to the sustained blood pressure increase to intermittent hypoxia.

Chronic Intermittent Hypoxia Alters Rat Ophthalmic Artery Reactivity Through Oxidative Stress, Endothelin and Endothelium-Derived Hyperpolarizing Pathways.

Obstructive sleep apnea recently has been associated with a higher frequency of ischemic optic neuropathies. Intermittent hypoxia (IH) has been proposed as a major component of obstructive sleep apnea cardiovascular consequences. However, there currently are no pathophysiologic data regarding the effect of IH on the ocular vascular system. Thus, we assessed the impact of chronic IH exposure on the morphology and vascular reactivity of the rat ophthalmic artery (OA). Rats were exposed to 14 days of IH or normoxia (NX). Ophthalmic artery reactivity was studied using wire myography in rats treated or not with tempol (1 mM/day). Expression of endothelin-1 (ET-1) and its receptors, and of the three nitric oxide synthase (NOS) isoform genes was quantified using quantitative polymerase chain reaction (qPCR) in the retina and optic nerve. Structural alterations (optical and electron microscopy) and superoxide anion production were studied in OA sections. Superoxide ion expression in the OA wall was increased by 23% after IH exposure. Ophthalmic artery contractile response to 3.10-8 M ET-1 was increased by 18.6% and nitric oxide-mediated relaxation was significantly delayed in IH compared to NX rats. In the absence of nitric oxide, cytochrome P450 blockade increased relaxation to acetylcholine in IH rats and delayed it in NX rats. Tempol treatment abolished the IH-induced changes in OA reactivity. These results strongly suggest that chronic IH induces oxidative stress in the rat OA, associated with endothelial dysfunction through alterations of nitric oxide and endothelium-derived hyperpolarising factors (EDHF) pathways.

Chronic Intermittent Hypoxia Causes Cellular Hypoxia in the Hippocampus

The hippocampus is a brain structure frequently affected in individuals suffering from sleep apnea. Although hypoxia is a well‐recognized environmental change that negatively impacts hippocampal neurophysiology, it is unclear whether brief periods of hypoxia experienced during sleep apnea are sufficient for producing cellular hypoxia. We conducted immunohistochemical studies in tissue harvested from mice (P30–P50) exposed to 0 or 10 days of intermittent hypoxia (IH) to address this issue. Staining for protein adducts of reductively activated 2‐nitroimidazoles was used as marker for cellular hypoxia. Our ongoing experiments indicate that IH causes an increase in the number of hypoxic cells restricted primarily to the granular cell layer and subgranular zone of the dentate gyrus. While these findings suggest that cellular hypoxia caused by IH exposure may not be widespread throughout the hippocampus, we propose that increased cellular hypoxia in the dentate gyrus may contribute to changes in adult neurogenesis and synaptic plasticity in this area critical to cognition.Support or Funding InformationFaculty Diversity Career Advancement GrantThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Chronic intermittent hypoxia promotes myocardial ischemia-related ventricular arrhythmias and sudden cardiac death

We investigated the effects of intermittent hypoxia (IH), such as that encountered in severe obstructive sleep apnea (OSA) patients, on the development and severity of myocardial ischemia-related ventricular arrhythmias. Rats were exposed to 14 days of IH (30 s at 5%O2 and 30 s at 21%O2, 8 h·day−1) or normoxia (N, similar air-air cycles) and submitted to a 30-min coronary ligature. Arterial blood pressure (BP) and ECG were recorded for power spectral analysis, ECG interval measurement and arrhythmia quantification. Left ventricular monophasic action potential duration (APD) and expression of L-type calcium (LTCC) and transient receptor potential (TRPC) channels were assessed in adjacent epicardial and endocardial sites. Chronic IH enhanced the incidence of ischemic arrhythmias, in particular ventricular fibrillation (66.7% vs. 33.3% in N rats, p < 0.05). IH also increased BP and plasma norepinephine levels along with increased low-frequency (LF), decreased high-frequency (HF) and increased LF/HF ratio of heart rate and BP variability. IH prolonged QTc and Tpeak-to-Tend intervals, increased the ventricular APD gradient and upregulated endocardial but not epicardial LTCC, TRPC1 and TRPC6 (p < 0.05). Chronic IH, is a major risk factor for sudden cardiac death upon myocardial ischemia through sympathoactivation and alterations in ventricular repolarization, transmural APD gradient and endocardial calcium channel expression.

Transcription factor ΔFosB acts within the nucleus of the solitary tract to increase mean arterial pressure during exposures to intermittent hypoxia.

ΔFosB is a member of the activator protein-1 family of transcription factors. ΔFosB has low constitutive expression in the central nervous system and is induced after exposure of rodents to intermittent hypoxia (IH), a model of the arterial hypoxemia that accompanies sleep apnea. We hypothesized ΔFosB in the nucleus of the solitary tract (NTS) contributes to increased mean arterial pressure (MAP) during IH. The NTS of 11 male Sprague-Dawley rats was injected (3 sites, 100 nl/site) with a dominant negative construct against ΔFosB (ΔJunD) in an adeno-associated viral vector (AAV)-green fluorescent protein (GFP) reporter. The NTS of 10 rats was injected with AAV-GFP as sham controls. Two weeks after NTS injections, rats were exposed to IH for 8 h/day for 7 days, and MAP was recorded using telemetry. In the sham group, 7 days of IH increased MAP from 99.8 ± 1.1 to 107.3 ± 0.5 mmHg in the day and from 104.4 ± 1.1 to 109.8 ± 0.6 mmHg in the night. In the group that received ΔJunD, IH increased MAP during the day from 95.9 ± 1.7 to 101.3 ± 0.4 mmHg and from 100.9 ± 1.7 to 102.8 ± 0.5 mmHg during the night (both IH-induced changes in MAP were significantly lower than sham, P < 0.05). After injection of the dominant negative construct in the NTS, IH-induced ΔFosB immunoreactivity was decreased in the paraventricular nucleus ( P < 0.05); however, no change was observed in the rostral ventrolateral medulla. These data indicate that ΔFosB within the NTS contributes to the increase in MAP induced by IH exposure. NEW & NOTEWORTHY The results of this study provides new insights into the molecular mechanisms that mediate neuronal adaptations during exposures to intermittent hypoxia, a model of the hypoxemias that occur during sleep apnea. These adaptations are noteworthy as they contribute to the persistent increase in blood pressure induced by exposures to intermittent hypoxia.

Epigenetic programming of autonomic functions in an experimental model of apnea of prematurity

Intermittent hypoxia (IH) is a hallmark manifestation of recurrent apneas, which is a major clinical problem in infants born preterm. Carotid body (CB) chemoreflex and catecholamine (CA) secretion from adrenal medullary chromaffin cells (AMCs) are two major mechanisms contributing to the maintenance of cardiorespiratory homeostasis during hypoxia. The purpose of this article is to highlight recent studies showing how neonates experiencing IH affect the CB and AMC function and their consequences in adult life. To simulate apneas, rat pups were treated with IH consisting of alternating cycles of hypoxia (1.5% O2) for 15 s and room air for 5 min, 8 h/day from ages P0–P10. Rats treated neonatal IH displayed augmented CB response to hypoxia and augmented CA secretion from AMC. Rats treated for 10 days of IH in the neonatal period were allowed to grow into adulthood. Remarkably, the effects of neonatal IH on CB and AMC persisted in the adulthood. Moreover, adult rats that were exposed to IH in neonatal period exhibited hypertension, increased incidence apnea. Analysis of the underlying molecular mechanisms revealed re-programming of the redox state by epigenetic mechanisms involving suppression of transcription of antioxidant enzyme genes by DNA hypermethylation. DNA hypomethylating agents might offer a novel therapeutic intervention to prevent early onset of cardiorespiratory morbidities caused by neonatal IH.

Exercise Attenuates Intermittent Hypoxia-Induced Cardiac Fibrosis Associated with Sodium-Hydrogen Exchanger-1 in Rats.

Purpose: To investigate the role of sodium–hydrogen exchanger-1 (NHE-1) and exercise training on intermittent hypoxia-induced cardiac fibrosis in obstructive sleep apnea (OSA), using an animal model mimicking the intermittent hypoxia of OSA.Methods: Eight-week-old male Sprague–Dawley rats were randomly assigned to control (CON), intermittent hypoxia (IH), exercise (EXE), or IH combined with exercise (IHEXE) groups. These groups were randomly assigned to subgroups receiving either a vehicle or the NHE-1 inhibitor cariporide. The EXE and IHEXE rats underwent exercise training on an animal treadmill for 10 weeks (5 days/week, 60 min/day, 24–30 m/min, 2–10% grade). The IH and IHEXE rats were exposed to 14 days of IH (30 s of hypoxia—nadir of 2–6% O2—followed by 45 s of normoxia) for 8 h/day. At the end of 10 weeks, rats were sacrificed and then hearts were removed to determine the myocardial levels of fibrosis index, oxidative stress, antioxidant capacity, and NHE-1 activation.Results: Compared to the CON rats, IH induced higher cardiac fibrosis, lower myocardial catalase, and superoxidative dismutase activities, higher myocardial lipid and protein peroxidation and higher NHE-1 activation (p < 0.05 for each), which were all abolished by cariporide. Compared to the IH rats, lower cardiac fibrosis, higher myocardial antioxidant capacity, lower myocardial lipid, and protein peroxidation and lower NHE-1 activation were found in the IHEXE rats (p < 0.05 for each).Conclusion: IH-induced cardiac fibrosis was associated with NHE-1 hyperactivity. However, exercise training and cariporide exerted an inhibitory effect to prevent myocardial NHE-1 hyperactivity, which contributed to reduced IH-induced cardiac fibrosis. Therefore, NHE-1 plays a critical role in the effect of exercise on IH-induced increased cardiac fibrosis.