Multiple Platform Method Research Articles

Shenling Baizhu San alleviates central fatigue through SIRT1-PGC-1α-Mediated mitochondrial biogenesis

Ethnopharmacological relevanceShenling Baizhu San (SLBZS) is a Traditional Chinese Medicine (TCM) formula composed of 10 medicinal herbs, historically used to strengthen the spleen, replenish qi, and alleviate fatigue-related symptoms. SLBZS originates from the 'Taiping Huimin Heji Ju Fang' of the Song Dynasty. Central fatigue (CF), a subtype of fatigue, is considered in TCM to be closely associated with spleen deficiency. However, there is currently a lack of research on SLBZS's therapeutic effects on CF and the pharmacological mechanisms underlying its potential benefits. Aim of the studyThis study aims to assess the effects of SLBZS on CF in rats induced by the Modified Multiple Platform Method (MMPM) and to elucidate the underlying mechanisms, focusing on mitochondrial biogenesis and SIRT1/PGC-1α pathway regulation. Materials and methodsCF was induced in male Wistar rats using MMPM, involving intermittent sleep deprivation over 21 days. SLBZS was administered at low(LSLBZS), medium(MSLBZS), and high doses(HSLBZS). Chemical components of SLBZS were identified and quantified using Liquid Chromatography-Tandem Mass Spectrometry(LC-MS/MS). Behavioral tests evaluated physical performance, emotional state, and cognitive function, while serum biochemical markers, mitochondrial morphology, and the protein and gene expression levels of the SIRT1/PGC-1α pathway were analyzed to explore underlying mechanisms. ResultsA total of 141 main compounds in SLBZS were identified, comprising various components such as flavonoids, phenylpropanoids, terpenoids, among others. SLBZS significantly improved physical performance, alleviated negative emotions, and enhanced cognitive function in CF rats. Biochemically, SLBZS increased serum ATP levels and reduced fatigue-related markers. Mitochondrial analysis demonstrated that SLBZS reversed mitochondrial degeneration, increased mitochondrial number, and increased mtDNA copy number in the hippocampus. Furthermore, SLBZS upregulated SIRT1/PGC-1α pathway expression at both the protein and gene levels in the hippocampus. Notably, the HSLBZS group demonstrated particularly pronounced effects. ConclusionSLBZS significantly alleviates CF symptoms enhances mitochondrial function via upregulating the SIRT1/PGC-1α pathway, positioning it as a promising alternative for CF management by addressing both its physiological and symptomatic aspects.

Proteomic Insights into the Effects of Jianweixiaoshi Tablets on Functional Dyspepsia with Spleen Deficiency in Rats.

Jianweixiaoshi tablets (JWXS) is widely used in traditional Chinese medicine for treating functional dyspepsia with spleen deficiency (SD-FD) in China. However, the molecular mechanisms underlying the therapeutic effects of JWXS remain incompletely understood. Functional dyspepsia was induced in rats with spleen deficiency by iodoacetamide in combination with the modified multiple platform method. The SD-FD rats were administered JWXS at both low and high doses, as well as domperidone. We conducted a comprehensive evaluation of the treatment effects of JWXS, including body weight, gastrointestinal motility, immune organ index, biochemical analysis, gastrointestinal hormones, and hematological studies. Quantitative proteomic analysis based on data-independent acquisition (DIA) was used to determine the changes in protein profiles of gastric and duodenal tissues in SD-FD rats and JWXS intervention rats. The results showed that JWXS effectively alleviated gastrointestinal motility disorders in SD-FD rats, as indicated by accelerated gastric emptying and intestinal propulsion, increased levels of gastrin, motilin, and ghrelin, and reduced levels of cholecystokinin-octapeptide, vasoactive intestinal peptide, and somatostatin. Additionally, JWXS increased the spleen and thymus index, increased %lymphocyte in blood, reduced white blood cell count and %neutrophil, and improved immune function. Through quantitative proteomic analysis of gastric tissues, we identified 333 differentially expressed proteins in the JWXS treatment group and the model group. Notably, the mechanism by which JWXS accelerated gastric emptying may be related to PLC-γ and SERCA2 in the calcium signaling pathway. Furthermore, JWXS treatment altered the expression of 732 proteins in rat duodenal samples. The differentially expressed proteins were enriched in immune-related functions and pathways, including antigen processing and presentation, as well as the intestinal immune network for IgA production. In conclusion, JWXS exhibits a multi-faceted impact on various pathways, demonstrating its efficacy in treating SD-FD. These findings provide a foundation for the clinical application of JWXS in managing SD-FD.

Male Gender Aggravates Insomnia-Associated Disturbances in IPGTT, Insulin Level and Insulin Resistance, with Less Improvement on Retrieval of Normal Sleep Compared to Aged Females

Abstract Background and Aim Prevalence of type 2 diabetes is growing worldwide in the last decades. Recent studies showed that sleeping hours had decreased in the last decades amongst both young and aged population. This study was conducted to investigate the effects of chronic insomnia and its relief on the intraperitoneal glucose tolerance test (IPGTT),insulin level and insulin resistance in aged male and female rats with insight in the underlying mechanisms. Methods The study was carried out on 66 local strain aged rats of both sexes. Range of male and female rat weights was 350-400 and 250-350 gm, respectively. Range of ages of male and female rats were 23-25 and 24-26 months, respectively. Rats were allocated into two study groups, males (n°= 32) and females (n°= 34). The male group was further subdivided into the following 4 subgroups: Male one month control group (MC1, n°=7); Male sleep -deprived group for one month (MSD, n°=10); Male two -month control group (MC2, n°=7); Male sleep deprived group for one month followed by one month reversion to regular sleep (MSDR, n°=8). The female group was further subdivided into 4 subgroups; Female one month control group (FC1, n°=9); Female sleep deprived group for one month (FSD, n°=7); Female two -month control group (FC2, n°=9); Female sleep deprived group for one month followed by one month reversion to regular sleep (FSDR, n°=9). The experimental method of sleep restriction was the modified multiple platform method (MMPM) which deprives the animal from rapid eye movement (REM) sleep, while the non-REM sleep unaffected without the need for monitoring the EEG. The study duration was 2 months, Rats were assessed for IPGTT, fasting serum insulin (FSI) and homeostasis model assessment of insulin resistance (HOMA-IR) as well as serum levels of corticosterone (Cs) and melatonin (Mt). Results After one month of insomnia, male rats (MSD) became prediabetic with fasting hyperglycemia (FBG 107 mg/dl) and significantly elevated blood glucose values in the IPGTT and area under the curve (AUC), FSI and HOMA-IR as well as Cs and significant decrease of Mt. compared to their matched controls (MC1). Reversion to normal sleep normalized the FBG of MSDR group and blood glucose levels at 30, 90, 120 minutes and area under the curve (AUC), with significant decrease of FSI and HOMA-IR, although not yet normalized compared to MC2 rat group. Similarly, Cs and Mt. levels showed partial improvement compared to MSD group, but not fully normalized compared to MC2 group. On the other hand, FSD rat group had no significant change in their FBG compared to FC1 group but had significant increase in their blood glucose levels at 60, 90, 120 min., AUC of IPGTT as well as FSI and HOMA-IR, Cs and significant decrease of Mt. compared to FC1 group. Reversion to normal sleep normalized IPGTT value of FSDR rats, although FSI and HOMA-IR, remained significantly high compared to FC2 rat group. Similarly, Cs and Mt. levels showed partial improvement compared to FSD group, but not fully normalized compared to FC2 group. When MSD and FSD rat groups were compared, MSD rats showed significantly high FBG and IPGTT values compared to FSD rat group with no significant difference in insulin and insulin resistance values, with significantly higher values of Cs in MSD compared to FSD rat groups, with no significant difference in Mt. levels. Reversion to normal sleep in MSDR and FSDR rats caused more improvement in all values of IPGTT as well as AUC, insulin level and insulin resistance in females (FSDR) compared to males (MSDR) with significantly higher values of Mt in FSDR compared to MSDR rat groups, although Cs. Conclusion chronic insomnia has deleterious effects on glucose utilization and insulin resistance in aged male compared to aged female rats with lack of full recovery with reversion to normal sleep in IPGTT parameters, insulin levels and insulin resistance in aged males compared to aged females. These changes can be attributed to the hormonal changes of Cs. and Mt. who showed significant changes in both insomniac male and female rats with better improvement in Mt. in recovered female rat group compared male recovered group. Assessment of sleep history in prediabetic aged males should be recommended as well as assessment of IPGTT and insulin resistance in insomniac aged patients particularly males.

Chronic sleep deprivation induces erectile dysfunction through increased oxidative stress, apoptosis, endothelial dysfunction, and corporal fibrosis in a rat model.

Sleep is foundational for nocturnal erections, facilitating nutrient exchange and waste removal, which has brought widespread attention to the relationship between sleep and erectile dysfunction (ED). However, there is currently a lack of basic research confirming whether chronic sleep deprivation (CSD) leads to erectile impairment and its underlying pathological mechanisms. The study sought to investigate whether CSD impairs erectile function in rats and the potential tissue damage it may cause in rats. The modified multiple platform method was employed to induce CSD in 14 rats, randomly divided into a platform control group and a CSD group. After 3 weeks, erectile function was evaluated by measuring intracavernosal pressure following cavernous nerve stimulation. Arterial blood samples were then analyzed for testosterone levels, and cavernous tissues were processed for advanced molecular biology assays, including Western blotting and immunofluorescence. After inducing CSD, rats exhibited a marked reduction in erectile function, yet their serum testosterone levels remained statistically unchanged when compared with the control group. More importantly, rats in the CSD group exhibited a significant increase in oxidative stress levels, accompanied by low expression of HO-1 and high expression of NOX1 and NOX4. Subsequently, elevated oxidative stress induced increased apoptosis in smooth muscle and endothelial cells, as evidenced by significant decreases in CD31 and α-smooth muscle actin expression in the CSD group, demonstrated through Western blotting and immunofluorescence assays. Endothelial cell apoptosis led to a significant decrease in endothelial nitric oxide synthase, resulting in lowered levels of nitric oxide and cyclic guanosine monophosphate, which severely impaired the erectile mechanism. Additionally, activation of the transforming growth factor β1 fibrotic pathway led to increased levels of tissue fibrosis, resulting in irreversible damage to the penile tissue in the CSD group. Our study lacks further exploration of the molecular mechanisms linking CSD and ED, representing a future research focus for potential targeted therapies. Our findings demonstrated that CSD significantly impairs erectile function in rats. CSD severely impairs erectile function in rats. When exposed to CSD, rats exhibit significantly elevated oxidative stress levels, which lead to increased tissue apoptosis, endothelial dysfunction, and ultimately irreversible fibrotic changes in the tissues. Further researches into the potential molecular mechanisms are needed to identify possible therapeutic targets for ED related to CSD.

Dual-factor model of sleep and diet: a new approach to understanding central fatigue.

Numerous studies have recently examined the impact of dietary factors such as high-fat diets on fatigue. Our study aims to investigate whether high-fat diet (HFD) alone or combined with alternate-day fasting (ADF) can lead to the central fatigue symptoms and to investigate the potential integration of dietary and sleep variables in the development of central fatigue models. Seventy-five male Wistar rats were divided into five groups: control, HFD, HFD + ADF, modified multiple platform method (MMPM), and MMPM+HFD + ADF. Each group underwent a 21-day modeling period according to their respective protocol. Their behavioral characteristics, fatigue biochemical markers, hippocampal pathological changes, mitochondrial ultrastructure, and oxidative stress damage were analyzed. Our findings demonstrate that using only HFD did not cause central fatigue, but combining it with ADF did. This combination led to reduced exercise endurance, decreased locomotor activity, impaired learning and memory abilities, along with alterations in serum levels of alanine aminotransferase (ALT), creatine kinase (CK), and lactate (LAC), as well as hippocampal pathological damage and other central fatigue symptoms. Moreover, the MMPM+HFD + ADF method led to the most obvious central fatigue symptoms in rats, including a variety of behavioral changes, alterations in fatigue-related biochemical metabolic markers, prominent pathological changes in hippocampal tissue, severe damage to the ultrastructure of mitochondria in hippocampal regions, changes in neurotransmitters, and evident oxidative stress damage. Additionally, it was observed that rats subjected to HFD + ADF, MMPM, and MMPM+HFD + ADF modeling method exhibited significant brain oxidative stress damage. We have demonstrated the promotive role of dietary factors in the development of central fatigue and have successfully established a more stable and clinically relevant animal model of central fatigue by integrating dietary and sleep factors.

Abstract Tu063: Acute sleep deprivation induces cardiac remodeling via activation of AT1R/ERK/GSK-3β signaling

Background: Sleep deprivation (SD) has an adverse effect on cardiovascular system. However, whether acute SD contributes to the morphological and functional changes of the heart remains unclear. Approach and Results: The modified multiple platform method (MMPM) was used to establish the SD model in Sprague-Dawley rats. Acute SD significantly enhanced cardiac ejection fraction and fractional shortening in rats as observed by echocardiography. The elevated ratio of heart weight to body weight (HW/BW) and enlarged cross-section area of cardiomyocytes by WGA staining were detected in SD groups (3, 7,10 days). The upregulation of ANP and BNP was also detected by western blotting. Masson-trichrome staining showed significant fibrosis in the SD groups, which was confirmed by western blotting of the fibrosis markers α-SMA and collagen 1. Furthermore, SD increased the level of angiotensin II (Ang II) and angiotensin type 1 receptor (AT1R) in the heart tissue of SD groups accompanied by the activation of the ERK/GSK-3β pathway. Administration of valsartan and benazepril dramatically ameliorated SD-induced cardiac fibrosis and hypertrophy by downregulating Ang II/AT1R and phosphor-ERK/GSK-3β levels. Conclusion: Our results indicated that cardiac remodeling was induced by the acute and short duration of SD, in which the AT1R/ ERK/GSK3β signal pathway was involved. Our study demonstrates a novel complementary phenomenon induced by SD and provides detailed evidence for the risk of SD on the cardiovascular system.

Chronic Partial Sleep Deprivation Increased the Incidence of Atrial Fibrillation by Promoting Pulmonary Vein and Atrial Arrhythmogenesis in a Rodent Model.

Sleep deprivation (SD) is a recognized risk factor for atrial fibrillation (AF), yet the precise molecular and electrophysiological mechanisms behind SD-induced AF are unclear. This study explores the electrical and structural changes that contribute to AF in chronic partial SD. We induced chronic partial SD in Wistar rats using a modified multiple-platform method. Echocardiography demonstrated impaired systolic and diastolic function in the left ventricle (LV) of the SD rats. The SD rats exhibited an elevated heart rate and a higher low-frequency to high-frequency ratio in a heart-rate variability analysis. Rapid transesophageal atrial pacing led to a higher incidence of AF and longer mean AF durations in the SD rats. Conventional microelectrode recordings showed accelerated pulmonary vein (PV) spontaneous activity in SD rats, along with a heightened occurrence of delayed after-depolarizations in the PV and left atrium (LA) induced by tachypacing and isoproterenol. A Western blot analysis showed reduced expression of G protein-coupled receptor kinase 2 (GRK2) in the LA of the SD rats. Chronic partial SD impairs LV function, promotes AF genesis, and increases PV and LA arrhythmogenesis, potentially attributed to sympathetic overactivity and reduced GRK2 expression. Targeting GRK2 signaling may offer promising therapeutic avenues for managing chronic partial SD-induced AF. Future investigations are mandatory to investigate the dose-response relationship between SD and AF genesis.

Sleep deprivation in adolescent mice impairs long-term memory till early adulthood via suppression of hippocampal astrocytes.

Sleep deficiency is a rampant issue in modern society, serving as a pathogenic element contributing to learning and memory impairment, with heightened sensitivity observed in children. Clinical observations suggest that learning disabilities associated with insufficient sleep during adolescence can persist through adulthood, but experimental evidence for this is lacking. In this study, we examined the impact of early-life sleep deprivation (SD) on both short-term and long-term memory, tracking the effects sequentially into adulthood. We employed a modified multiple-platform method mouse model to investigate these outcomes. SD induced over a 14-day period, beginning on postnatal day 28 (PND28) in mice, led to significant impairment in long-term memory (while short-term memory remained unaffected) at PND42. Notably, this dysfunction persisted into adulthood at PND85. The specific impairment observed in long-term memory was elucidated through histopathological alterations in hippocampal neurogenesis, as evidenced by bromodeoxyuridine (BrdU) signals, observed both at PND42 and PND85. Furthermore, the hippocampal region exhibited significantly diminished protein expressions of astrocytes, characterized by lowered levels of aquaporin 4 (AQP4), a representative molecule involved in brain clearance processes, and reduced protein expressions of brain-derived neurotrophic factors. In conclusion, we have presented experimental evidence indicating that sleep deficiency-related impairment of long-term memory in adolescence can endure into adulthood. The corresponding mechanisms may indicate that the modification of astrocyte-related molecules has led to changes in hippocampal neurogenesis.

Methamphetamine and REM sleep deprivation interact to affect behavioral performance in adult and adolescent rats

Drug addiction may result in sleep problems. Importantly, sleep deprivation (SD) is known as an important risk factor for relapse to drug abuse as SD mimics the effects of psychostimulants on dopamine system of the brain. Moreover, aging may affect sleep and drug addiction. This study, therefore, set out to assess the effects of methamphetamine (METH) and REM sleep deprivation (RSD) on locomotor activity, anxiety-like behavior and spatial memory in adult and adolescent rats. Adult and adolescent male Wistar rats received a neurotoxic METH regimen; four subcutaneous injections of 6 mg/kg, at 2 h intervals. Five days later, the animals underwent a 48-h RSD episode using the multiple platforms method. They were then examined using the open field (OF), elevated plus maze (EPM) and Y-maze tasks. We found that the METH and RSD paradigms showed synergistic effects to increase locomotion and risk-taking behavior in both adult and adolescent animals, while only adolescent rats revealed RSD-induced anxiety-like behavior. Moreover, adolescent animals revealed greater sensitization for vertical activity following METH plus RSD episode. In addition, METH and RSD paradigms revealed synergistic effects to impair spatial working memory, but neither METH nor RSD alone affected performance of animals in the Y-maze task. Our findings may indicate that there are important relationships between METH and RSD to induce hyperlocomotion, risk-taking behavior and spatial memory impairment, particularly in adolescent animals. Moreover, it seems that adolescent rats may be more susceptible to anxiety-like behavior and hyperlocomotion than adults.

Replicating human characteristics: A promising animal model of central fatigue

Central fatigue is a common pathological state characterized by psychological loss of drive, lack of appetite, drowsiness, and decreased psychic alertness. The mechanism underlying central fatigue is still unclear, and there is no widely accepted successful animal model that fully represents human characteristics. We aimed to construct a more clinically relevant and comprehensive animal model of central fatigue. In this study, we utilized the Modified Multiple Platform Method (MMPM) combined with alternate-day fasting (ADF) to create the animal model. The model group rats are placed on a stationary water environment platform for sleep deprivation at a fixed time each day, and they were subjected to ADF treatment. On non-fasting days, the rats were allowed unrestricted access to food. This process was sustained over a period of 21 days. We evaluated the model using behavioral assessments such as open field test, elevated plus maze test, tail suspension test, Morris water maze test, grip strength test, and forced swimming test, as well as serum biochemical laboratory indices. Additionally, we conducted pathological observations of the hippocampus and quadriceps muscle tissues, transmission electron microscope observation of mitochondrial ultrastructure, and assessment of mitochondrial energy metabolism and oxidative stress-related markers. The results revealed that the model rats displayed emotional anomalies resembling symptoms of depression and anxiety, decreased exploratory behavior, decline in learning and memory function, and signs of skeletal muscle fatigue, successfully replicating human features of negative emotions, cognitive decline, and physical fatigue. Pathological damage and mitochondrial ultrastructural alterations were observed in the hippocampus and quadriceps muscle tissues, accompanied by abnormal mitochondrial energy metabolism and oxidative stress in the form of decreased ATP and increased ROS levels. In conclusion, our ADF+MMPM model comprehensively replicated the features of human central fatigue and is a promising platform for preclinical research. Furthermore, the pivotal role of mitochondrial energy metabolism and oxidative stress damage in the occurrence of central fatigue in the hippocampus and skeletal muscle tissues was corroborated.

The Effect of Anti-fatigue Decoction on the Behaviors and Serological Indicators in a Central Fatigue Rat Model.

This study aimed to assess the effects of Anti-fatigue Decoction (AFD) against central fatigue by observing the behaviors and serological indicators of rats modeled by the modified multiple platform method (MMPM) after drug intervention. Grip strength measurements were used to evaluate the muscle strength of rats. The open field test was utilized to assess anxiety-like behavior, while the Morris water maze test was conducted to evaluate the memory function of the rats. Following the behavioral assessments, rat serum samples were collected to measure the concentrations of corticosterone (CORT) and lactic acid (LAC). The concentration of LAC was determined using the colorimetric method, while the concentration of CORT was measured using the enzyme-linked immunosorbent assay (ELISA) method. Compared to the blank control group, following MMPM modeling, rats exhibited significant reductions in grip strength and impaired ability to memory. The serum analysis revealed increased levels of LAC and CORT in the model group rats. AFD can noticeably reverse these adverse changes to a certain extent. These findings highlight the positive effects of AFD and coenzymeQ10 on physical and cognitive abilities and alterations in serum biomarker levels of central fatigue rats.

Spinal bestrophin-1 and anoctamin-1 channels have a pronociceptive role in the tactile allodynia induced by REM sleep deprivation in rats

Bestrophin-1 and anoctamin-1 are members of the calcium-activated chloride channels (CaCCs) family and are involved in inflammatory and neuropathic pain. However, their role in pain hypersensitivity induced by REM sleep deprivation (REMSD) has not been studied. This study aimed to determine if anoctamin-1 and bestrophin-1 are involved in the pain hypersensitivity induced by REMSD. We used the multiple-platform method to induce REMSD. REM sleep deprivation for 48 h induced tactile allodynia and a transient increase in corticosterone concentration at the beginning of the protocol (12 h) in female and male rats. REMSD enhanced c-Fos and α2δ-1 protein expression but did not change activating transcription factor 3 (ATF3) and KCC2 expression in dorsal root ganglia and dorsal spinal cord. Intrathecal injection of CaCCinh-A01, a non-selective bestrophin-1 blocker, and T16Ainh-A01, a specific anoctamin-1 blocker, reverted REMSD-induced tactile allodynia. However, T16Ainh-A01 had a higher antiallodynic effect in male than female rats. In addition, REMSD increased bestrophin-1 protein expression in DRG but not in DSC in male and female rats. In marked contrast, REMSD decreased anoctamin-1 protein expression in DSC but not in DRG, only in female rats. Bestrophin-1 and anoctamin-1 promote pain and maintain tactile allodynia induced by REM sleep deprivation in both male and female rats, but their expression patterns differ between the sexes.

Cognitive Enhancing Activities of Coenzyme Q10, Ramipril, and Vinpocetine Through Modulating Neuroinflammatory Response and Oxidative Damage Inflicted by Acute REM Sleep Deprivation in Rats

Sleep plays a vital role to help in normal biological functions that are required for normal cognitive functioning. This study was done to determine the cognition-modulating effects of coenzyme Q10 (CoQ10), ramipril, and vinpocetine on REM sleep-deprived acute insomniac rat models. A total of Forty-eight albino rats were divided into eight groups (Gr) (n=6).). Gr. 1 was REM control and Gr. 2 was REM sleep-deprived rats treated with water. Gr. 3 to Gr.8 - REM sleep-deprived rats were administered corn oil, donepezil, vinpocetine, coenzyme Q10+corn oil, ramipril, and (coenzyme Q10 + corn oil + ramipril) respectively. Except for the control Gr 1, REM sleep deprivation was induced in Gr. 2 and 8 daily for 7 days. All the rats were subjected to a Morris water maze (MWM) to test the navigation memory dysfunction after 7 days of acute insomnia. The rats were deprived of REM sleep by using a modified multiple platform method. The body weight of the animals was measured on day 1 and day 7. On day 1 and Day 2 acquisition trials, all groups of rats showed comparable latency time required to reach the hidden platform. However, on day 3 and Day 4, rats treated with coenzyme Q10, ramipril, and the combination (CoQ10+ Ramipril) showed a significant decrease in latency time (p<0.01). In the probe trial, sleep-deprived rats showed a significant decrease (p<0.001) in the percentage of time spent in the target quadrant as compared to the REM control. However, there was a significant increase in the percentage time spent in CoQ10, ramipril, the combination (CoQ10+ ramipril), donepezil, and vinpocetine as compared to sleep-deprived rats (p<0.05). At the end of day 7 of insomnia, when the body weight of rats was compared with day 1, there was a significant decrease in weight gain was seen with the sleep-deprived rats treated with corn oil, ramipril, vinpocetine, CoQ10, and (CoQ10+ramipril) (p<0.05). The present study shows that coenzyme Q10, ramipril, and their combination improve sleep deprivation induced cognition impairment.

Effect of Sleep Deprivation on the Metabolism of Hippocampal Amino Acids and Monoamine Neurotransmitters in Mice and Their Behaviors

To investigate the effect of sleep deprivation on the metabolism of the hippocampal region in mice. The mice were randomly assigned to three groups, a control group, a 24-h sleep deprivation (SD) group, and a 48-h SD group. Each group had 10 mice. The sleep deprivation model was induced by the modified multiple platform method. The mice's anxiety-like behaviors were assessed with the open field test (OFT) and their depression-like behaviors were assessed with the sucrose preference test (SPT), the forced swimming test (FST), and tail suspension test (TST). High performance liquid chromatography (HPLC) was performed to determine the levels of 6 monoamine neurotransmitters, including 5-hydroxytryptamine (5-HT), norepinephrine (NE), dopamine (DA), gamma-aminobutyric acid (GABA), 5-dihydroxyphenylacetic acid (5-DOPAC), and homovanillic acid (HVA), and 4 amino acids, including glutamic acid (Glu), aspartic acid (Asp), serine (Ser), and taurine (Tau), in the hippocampal region. Immunofluorescence staining was performed to examine the expression of glial cells in the hippocampal region of the mice. The main indicators measured were the levels of monoamine neurotransmitters and amino acids. According to the results of the behavioral analysis, in comparison with the findings for the control group, the 24-h SD mice exhibited increased consumption of sucrose in SFT, significantly decreased total immobility time in FST and TST, and increased total distance covered in OFT, while the 48-h SD mice showed decreased consumption of sucrose in SFT, prolonged total immobility time in FST and TST, and decreased total distance covered in OFT. The results of the HPLC analysis of the monoamine neurotransmitter showed that 24-h SD mice had in their hippocampal region increased levels of DA (P<0.001) and NE (P<0.01) and decreased levels of GABA (P<0.05) in comparison with those of the control mice, while their 5-HT, 5-DOPAC, and HVA levels were not significantly different from those of the control mice. In comparison with those of the control mice, the 48-h SD mice had, in their hippocampal region, decreased levels of 5-HT and NE (all P<0.05), decreased DA (P<0.01), and increased level of GABA (P<0.01), while the levels of 5-DOPAC and HAV were not significantly different. The 48-h SD group showed a significant decrease in the levels of Tau and Glu in comparison with those of the 24-h SD group (all P<0.05). According to the results of immunofluorescence assay, there was no significant difference between the control group and the 24-h SD group in the cell count of glial fibrillary acidic protein (GFAP)-positive cells, while a decline in GFAP-positive cells in comparison with that of the control group was observed in the 48-h SD group. SD of 24 hours may induce anxiety-like behavioral changes in mice by activating their hippocampal glial cells, upregulating the levels of 5-HT, DA, and NE, and increasing the levels of Glu and Tau in the hippocampal region. SD of 48 hours may induce depression-like behavioral changes in mice by inhibiting the activation of glial cells in the hippocampal region and regulating in the opposite direction the levels of the above-mentioned monoamine neurotransmitters and amino acids in the hippocampal region.

SIRT6 Improves Hippocampal Neurogenesis Following Prolonged Sleep Deprivation Through Modulating Energy Metabolism in Developing rats.

Prolonged sleep deprivation is known to have detrimental effects on the hippocampus during development or in adulthood. Furthermore, it is well-established that sleep deprivation disrupts energy metabolism broadly. SIRT6 is a critical regulator of energy metabolism in both central and peripheral tissues. This study aims to investigate the role of SIRT6 in modulating hippocampal neurogenesis following sleep deprivation during development, and elucidate the underlying mechanism. Male Sprague-Dawley rats, aged three weeks, were subjected to 2 weeks of sleep deprivation using the modified multiple platform method. Metabolomic profiling was carried out using the liquid chromatography-electrospray ionization-tandem mass spectrometry (LC‒ESI‒MS/MS). To investigate the role of SIRT6 in energy metabolism, the rats were administered with either the SIRT6-specific inhibitor, OSS128167, or SIRT6-overexpressing adeno-associated virus (AAV). Hippocampal neurogenesis was assessed by immunostaining with markers for neural stem cells (SOX2), immature neurons [doublecortin (DCX)] and newborn cells (BrdU). Sparse labeling of adult neurons was used to determine the density of dendritic spines in the dentate gyrus (DG). The Y-maze and novel object recognition (NOR) tests were performed to evaluate the spatial and recognition memory. SIRT6 expression was examined using immunofluorescence and western blotting (WB). The inhibition of SIRT6 was confirmed by assessing the acetylation of histone 3 lysine 9 (aceH3K9), a well-known substrate of SIRT6, through WB. Sleep deprivation for a period of two weeks leads to inhibited hippocampal neurogenesis, reduced density of dendritic spines in the DG, and impaired memory, accompanied by decreased SIRT6 expression and disrupted energy metabolism. Similar to sleep deprivation, administration of OSS128167 significantly decreased energy metabolism, leading to reduced neurogenesis and memory dysfunction. Notably, the abnormal hippocampal energy metabolism, neurogenetic pathological changes and memory dysfunction caused by sleep deprivation were alleviated by SIRT6 overexpression in the DG. Our results suggest that SIRT6 plays a critical role in maintaining energy metabolism homeostasis in the hippocampus after sleep deprivation, promoting hippocampal neurogenesis and enhancing memory during development.

The relationship between gut microbiota and inflammatory response, learning and memory in mice by sleep deprivation.

Sleep deprivation has developed into a common phenomenon, which can lead to inflammatory responses and cognitive impairment, but the underlying mechanism is ambiguous. Emerging evidence shows that gut microbiota plays a crucial role in theoccurrence and development of inflammatory and psychiatric diseases, possibly through neuroinflammation and the brain-gut axis. The current study investigated the influence of sleep deprivation on gut microbiota composition, pro-inflammatory cytokines, learning and memory in mice. Further, it explored whether changes in gut microbiota increase pro-inflammatory cytokine and induce learning and memory impairment. Healthy 8-week-old male C57BL/6J mice were randomly divided into the regular control group (RC), environmental control group (EC), and sleep deprivation group (SD). The sleep deprivation model was established by the Modified Multiple Platform Method. The experimental mice were subjected to sleep deprivation for 6h/d (8:00 am∼14:00 pm) in a sleep deprivation chamber, and the duration of sleep deprivation was 8 weeks. Morris water maze test to assess learning and memory in mice. Enzyme-Linked Immunosorbent Assay determined the concentrations of inflammatory cytokines. The changes in gut microbiota in mice were analyzed by 16S rRNA sequencing. We found that SD mice had elevated latency of exploration to reach the hidden platform (p>0.05) and significantly decreased traversing times, swimming distance, and swimming time in the target zone when the hidden platform was removed (p<0.05). Sleep deprivation caused dysregulated expression in serum IL-1β, IL-6, and TNF-α in mice, and the difference was significant (all p<0.001). Tannerellaceae, Rhodospirillales, Alistipes, and Parabacteroides were significantly increased in SD mice. Correlation analysis showed IL-1β was positively correlated with the abundance of Muribaculaceae (r=0.497, p<0.05) and negatively correlated with the abundance of Lachnospiraceae (r=-0.583, p<0.05). The TNF-α was positively correlated with the abundances of Erysipelotrichaceae, Burkholderiaceae, and Tannerellaceae (r=0.492, r=0.646, r=0.726, all p<0.05). Sleep deprivation can increase pro-inflammatory cytokine responses and learning and memory impairment in mice and may be caused by the disorder of the microbiota. These findings of this study may open avenues for potential interventions that can relieve the detrimental consequences of sleep loss.

The simultaneous action of acute paradoxical sleep deprivation and hypothyroidism modulates synaptosomal ATPases and acetylcholinesterase activities in rat brain

BackgroundThyroid dysfunctions as well as sleep abnormalities are usually followed by neurological, psychiatric and/or behavioral disorders. On the other hand, changes in the brain adenosine triphosphatases (ATPases) and acetylcholinesterase (AChE) activities show significant importance in pathogenetic pathways in the evolution of numerous neuropsychiatric diseases. MethodsThis study aimed to evaluate the in vivo simultaneous effects of hypothyroidism and paradoxical sleep deprivation for 72 h on synaptosomalATPases and AChE activities of whole rat brains.In order to induce hypothyroidism, 6-n-propyl-2-thiouracil was administrated in drinking water during 21 days. The modified multiple platform method was used to induce paradoxical sleep deprivation. The AChE and ATPases activities were measured using spectrophotometric methods. ResultsHypothyroidism significantly increased the activity of Na+/K+-ATPase compared to other groups, while at the same time significantly decreased AChE activity compared to the CT and SD groups. Paradoxical sleep deprivation significantly increased AChE activity compared to other groups. The simultaneous effect of hypothyroidism and sleep deprivation reduced the activity of all three enzymes (for Na+/K+-ATPase between HT/SD and HT group p < 0.0001, SD group p < 0.001,CT group p = 0.013; for ecto-ATPases between HT/SD and HT group p = 0.0034, SD group p = 0.0001, CT group p = 0.0007; for AChE between HT/SD and HT groupp < 0.05, SD group p < 0.0001, CT group p < 0.0001). ConclusionsThe effect of simultaneous existence of hypothyroidism and paradoxical sleep deprivation reduces the activity of the Na+/K+-ATPase, ecto-ATPases, and AChE, what is different from individual effect of hypothyroidism and paradoxical sleep deprivation itself. This knowledge could help in the choice of appropriate therapy in such condition.

Effects of prebiotics on intestinal physiology, neuropsychological function, and exercise capacity of mice with sleep deprivation

People suffered from insufficient or disrupted sleep due to night shifts, work pressure, and irregular lifestyles. Sleep deprivation caused by inadequate quantity or quality of sleep has been associated with not only increased risk of metabolic diseases, gut dysbiosis, and emotional disorders but also decreased work and exercise performance. In this study, we used the modified multiple platform method (MMPM) to induce pathological and psychological characteristics of sleep deprivation with C57BL/6J male mice, and investigated whether supplementing a prebiotics mixture of short-chain galactooligosaccharides (scGOS) and long-chain fructooligosaccharides (lcFOS) (9:1 ratio) could improve the impacts of sleep deprivation on intestinal physiology, neuropsychological function, inflammation, circadian rhythm, and exercise capacity. Results showed that sleep deprivation caused intestinal inflammation (increased TNFA and IL1B) and decreased intestinal permeability with a significant decrease in the tight junction genes (OCLN, CLDN1, TJP1, and TJP2) of intestine and brain. The prebiotics significantly increased the content of metabolite short-chain fatty acids (acetate and butyrate) while recovering the expression of indicated tight junction genes. In hypothalamus and hippocampus, clock (BMAL1 and CLOCK) and tight junction (OCLN and TJP2) genes were improved by prebiotics, and corticotropin-releasing hormone receptor genes, CRF1 and CRF2, were also significantly regulated for mitigation of depression and anxiety caused by sleep deprivation. Also, prebiotics brought significant benefits on blood sugar homeostasis and improvement of exercise performance. Functional prebiotics could improve physiological modulation, neuropsychological behaviors, and exercise performance caused by sleep deprivation, possibly through regulation of inflammation and circadian rhythm for health maintenance. However, the microbiota affected by prebiotics and sleep deprivation should warrant further investigation.

Effects of electroacupuncture on the ventral tegmental area- nucleus accumbens dopamine pathway in rats with chronic sleep deprivation.

Insomnia is a well-recognized clinical sleep disorder in the adult population. It has been established that acupuncture has a clinical effects in the treatment of insomnia; however, research on the underlying neural circuits involved in these effects is limited. The modified multiple platform method (MMPM) was used to establish a rat model of chronic sleep deprivation (CSD). Forty rats were randomly divided into a control (Con) group, (untreated) CSD group, electroacupuncture-treated CSD group (CSD + EA) and estazolam-treated CSD group (CSD + Estazolam group) with n = 10 per group. In the CSD + EA group, EA was delivered at Yintang and unilateral HT7 (left and right treated every other day) with continuous waves (2 Hz frequency) for 30 min/day over 7 consecutive days. In the CSD + Estazolam groups, estazolam was administered by oral gavage (0.1 mg/kg) for 7 consecutive days. The open field test (OFT) was used to observe behavioral changes. Immunofluorescence assays and enzyme-linked immunosorbent assay (ELISA) were used to observe the effects of EA on the ventral tegmental area (VTA)-nucleus accumbens (NAc) dopamine (DA) pathway. We also assessed the effects of EA on the expression of dopamine D1 receptor (D1R) and dopamine D2 receptor (D2R) in the NAc, which are the downstream targets of the VTA-NAc DA pathway. After CSD was established by MMPM, rats exhibited increased autonomous activity and increased excitability of the VTA-NAc DA pathway, with increased VTA and NAc DA content, increased D1R expression and decreased D2R expression in the NAc. EA appeared to reduce the autonomous ability of CSD rats, leading to lower DA content in the VTA and NAc, reduced expression of D1R in the NAc and increased expression of D2R. Most importantly, EA produced effects similar to estazolam with respect to the general condition of rats with CSD and regulation of the VTA-NAc DA pathway. The therapeutic effect of EA in chronic insomnia may be mediated by reduced excitability of the VTA-NAc DA pathway, with lower DA content in the VTA and NAc, downregulated expression of D1R in the NAc and increased expression of D2R.

Attenuation Effect of Recovery Sleep for Impaired Reproductive Function in Male Rats by Sleep Deprivation.

The aim of the present study was to test the hypothesis that recovery sleep could counteract the detrimental effects of sleep deprivation (SD) on male rats' fertility. Twenty-two rats were housed in groups of six per cage with unrestricted access to food and water in a room. The modified multiple platform method was used to induce SD in rats over a 96-hour period. We examined the effect of SD on semen quality, reproductive hormones, and testicular histology in adult male rats. Then, we investigated the effect of 7 days recovery sleep on impaired reproductive function induced by SD. After the acclimation period, 22 rats were randomly separated into three experimental groups (SD, recovery sleep, and the control groups). Ninety-six hours of SD resulted in a significant decrease in sperm motility (24.33±10.93 vs. 48.20±8.55, p<0.001) and the number of morphologically normal sperm (9.68±2.77 vs. 26.21±14.60, p<0.01) in rats, accompanied by a decrease in testosterone levels (1.53±0.55 vs. 4.44±0.56, p<0.001) and destruction of testicular tissue structure compared with control group. After 7 days of recovery sleep, semen quality, especially sperm motility, was improved and testosterone levels were significantly higher compared to post-SD (3.70±0.53 vs. 1.53±0.55, p<0.05), but remained low compared to the control group. In conclusion, 96 hours of SD deteriorated the parameters of sperm motility and the number of morphologically normal sperm in rats, probably due to the decrease in serum testosterone levels and the disruption of testicular tissue structure when compared to the control group. After 7 days of recovery sleep, semen parameter, especially sperm motility and testosterone levels did not return to baseline levels compared to the control group.