| Background and Objectives:Oxygen is one of the essential substances for aerobic metabolism,hypoxia is common in clinical diseases and daily life.Systemic hypoxia drives multi-organ damage such as cognitive decline,pulmonary hypertension,heart failure,liver injury and renal damage.Different organs have varying tolerance to hypoxia,partially due to both the systemic redistribution of the inadequate O2 supply in favor of vital organs and the distinctive intrinsic organ responses.However,the hypoxia sensitivity of various organs is unclear.The brain is a highly oxidative organ with 20%of the total systemic oxygen consumption and low glycolytic capacity,it contains a large number of enzymes that use O2 as a substrate and is one of the most oxygen-sensitive organs.The visual cortex is the main part that processes visual information which is anatomically complex,and there are functional connections between the internal layers as well as external connection pathways with other brain regions.In recent years,a large number of clinical studies have confirmed the appearance of visual impairment in hypoxia-related diseases.A previous series of studies by our group found that plateau hypoxia exposure causes changes in the insula and the primary visual cortex.However,it is not clear whether the primary visual cortex is the first to be damaged after hypoxia exposure and thus visual dysfunction occurs.In this study we investigate the effects of hypoxia on the structures of different brain regions,we clarified the primary visual cortex as a hypoxia-sensitive brain region,and explored the mechanism of hypoxia damage.Methods:The first part explored the hypoxia sensitivity of each organ.Adult male mice were exposed to 6%O2 for 6 hours.The establishment of the acute hypoxic model was verified by the changes of physiological and behavior indexes.In vivo and in vitro imaging of anti-HIF-1α-NMs-cy5.5 nanocomposites to investigate the real-time distribution of HIF-1α.Western blot and immunofluorescence were also applied to verify the quantification and localization of HIF-1α.To explore the changes in metabolic characteristics of hypoxia-sensitive organs,an NMR-based metabolomics technology was used to explore the metabolic alterations in the liver,kidney and brain after hypoxia exposure.The second part studied the hypoxia-sensitive brain regions,a chronic hypoxic model with 14%O2 exposure for two weeks was established,and MRI structural image scans were performed before and after hypoxia,respectively.Voxel-based morphometry was used to find brain regions with differences in gray matter volume after hypoxia.Immunofluorescence and Nissl staining were performed on the brain removed by execution immediately after MRI scanning.To clarify whether the primary visual cortex was the first to be damaged by hypoxia,two-photon imaging techniques were applied to calcium imaging and angiography.To investigate the mechanism of hypoxic visual cortex injury and the role of microglia,a visual stimulation and binocular visual deprivation model was established to perform two-photon calcium imaging on primary visual cortex.Csf1r heterozygous deletion mice were established for transmission electron microscopy scanning and two-photon angiography of primary visual cortex,and further immunofluorescence techniques were applied to compare the microglia in primary visual cortex after hypoxia.Results:In the first part,acute hypoxia exposure decreased blood oxygen concentration,increased respiratory rate and heart rate,decreased exploratory behavior,and deteriorated voluntary locomotion in mice.the expression level of HIF-1α was the highest in the liver,followed by kidney and brain.HIF-1α was detected in the hepatocytes of liver,distal convoluted tubules of kidney and neurons of cerebral cortex.Liver,kidney and brain 1H-NMR spectra identified 44,59 and 39 metabolites,respectively,and hypoxia did not cause changes in the type of metabolites in mice,but rather in the concentration.The liver,kidney and brain showed distinct metabolic profiles but an identical change in glutamate.The liver had more characteristic metabolites and more disturbed metabolic pathways related to glutaminolysis and glycolysis.The level of O-phosphocholine,GTP,NAD and aspartate were upregulated in hypoxic mice brain,which displayed significant positive correlations with the locomotor activity in control mice,but not in hypoxic mice with impaired locomotor activities.In the second part,gray matter volume in the primary visual cortex was significantly reduced after chronic hypoxic exposure.Neuronal density in layers 2/3,4,and 6 of primary visual cortex was significantly reduced and morphologically abnormal,with increased microvessels,significantly reduced tight junction proteins compared to primary motor cortex and primary somatosensory cortex.Neurons in primary visual cortex showed a strong response to hypoxic stimulation,enhanced neural activity and blood flow.When visual stimulation was performed,calcium signals within primary visual cortex neurons were enhanced,excited primary visual cortex neurons were more sensitive to hypoxia.The Ca2+wave in the primary visual cortex region of the visual deprivation model did not show significant changes after hypoxia.This suggests that stronger neural activity and high ATP demand in visual cortex made them more sensitive to hypoxia.Hypoxia did not cause changes in the number of microglia,but led to vascular leakage in the primary visual cortex,releasing fibrinogen which recruited and activated perivascular microglia.Further disruption of the blood-brain barrier in the primary visual cortex may due to phagocytosis and destruction of endothelial cells by activated microglia.Microglia deficiency delays the effect of hypoxia on cerebral blood flow in the primary visual cortex and the blood-brain barrier is not damaged by hypoxia.Conclusion:The liver,kidney and brain are the three main organs of the body that are strongly respond to acute hypoxia,and the liver exhibited the highest hypoxic sensitivity.The metabolic disorders appear to underlie the physiological function changes.The primary visual cortex is a hypoxia-sensitive brain region,and its response to acute hypoxia by enhanced neuronal activity and increased cerebral blood flow.Chronic hypoxia causes neuronal loss and cerebrovascular damage and mice exhibit visual defects,microglia activation is associated with hypoxic cerebrovascular injury.This project is the first to compare the hypoxia sensitivity of vital organs,and to explore the metabolic alterations in hypoxia-sensitive organs,which is a necessary step to the mechanisms of oxidative stress.The exploration of hypoxia-sensitive brain regions can predict the visual damage which caused by hypoxic,which has a certain guiding role in common altitude training and altitude travel,and provides a theoretical basis for relevant protective measures.It improving people’s understanding of the impairment of brain function in hypoxiarelated diseases. |
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