Endoplasmic Reticulum Stress-Activated Neuronal and Microglial Autophagy Contributes to Postoperative Cognitive Dysfunction in Neonatal rats.

Surgery and anesthesia in neonates may lead to cognitive impairment or abnormal behaviors. It has been shown that autophagy plays a critical role in neuropsychiatric disorders, while the role of autophagy in postoperative cognitive impairment in neonates is not known. Here, we determined this role and the involvement of endoplasmic reticulum (ER) stress in regulating brain cell autophagy after surgery. Seven-day old neonatal rats (P7) had right common carotid artery exposure under anesthesia with 3% sevoflurane for 2 h. Learning and memory were tested using Barnes maze (BM) and fear conditioning (FC) on P31-42 and P42-44, respectively. In another experiment, rat brains were harvested for biochemical studies. The ratio of microtubule-associated protein 1 light chain 3 (LC3) BII/I was increased and sequestosome 1 (P62/SQSTM1) levels were decreased in the brain 24 h after surgery and anesthesia in neonatal rats. Immunofluorescent staining of LC3B was co-localized with a neuronal or a microglial marker but was not co-localized with a marker for astrocytes in rats with surgery. These rats had a poorer performance in the BM and FC tests than control rats when they were adolescent. Pretreatment with an autophagy inhibitor, 3-methyladenine, attenuated the poor performance. Surgery and anesthesia increased the expression of 78 kDa glucose-regulated protein (BIP/GRP78), an indicator of ER stress, 6 h after surgery and anesthesia. The ER stress activator tunicamycin and inhibitor tauroursodeoxycholic acid increased the markers for autophagy in control rats and decreased the autophagy markers in rats with surgery, respectively. Our results suggest that surgery in neonatal rats induces ER stress that then activates neuronal and microglial autophagy, which contributes to learning and memory impairment later in life.

Appropriate exercise level attenuates gut dysbiosis and valeric acid increase to improve neuroplasticity and cognitive function after surgery in mice.

Postoperative cognitive dysfunction (POCD) affects the outcome of millions of patients each year. Aging is a risk factor for POCD. Here, we showed that surgery induced learning and memory dysfunction in adult mice. Transplantation of feces from surgery mice but not from control mice led to learning and memory impairment in non-surgery mice. Low intensity exercise improved learning and memory in surgery mice. Exercise attenuated surgery-induced neuroinflammation and decrease of gut microbiota diversity. These exercise effects were present in non-exercise mice receiving feces from exercise mice. Exercise reduced valeric acid, a gut microbiota product, in the blood. Valeric acid worsened neuroinflammation, learning and memory in exercise mice with surgery. The downstream effects of exercise included attenuating growth factor decrease, maintaining astrocytes in the A2 phenotypical form possibly via decreasing C3 signaling and improving neuroplasticity. Similar to these results from adult mice, exercise attenuated learning and memory impairment in old mice with surgery. Old mice receiving feces from old exercise mice had better learning and memory than those receiving control old mouse feces. Surgery increased blood valeric acid. Valeric acid blocked exercise effects on learning and memory in old surgery mice. Exercise stabilized gut microbiota, reduced neuroinflammation, attenuated growth factor decrease and preserved neuroplasticity in old mice with surgery. These results provide direct evidence that gut microbiota alteration contributes to POCD development. Valeric acid is a mediator for this effect and a potential target for brain health. Low intensity exercise stabilizes gut microbiota in the presence of insult, such as surgery.

Toll-like receptor 2 activation and up-regulation by high mobility group box-1 contribute to post-operative neuroinflammation and cognitive dysfunction in mice.

Post-operative cognitive dysfunction (POCD) is common and is associated with poor clinical outcome. Toll-like receptor (TLR) 3 and 4 have been implied in the development of POCD. The role of TLR2, a major brain TLR, in POCD is not clear. High mobility group box-1 (HMGB1) is a delayed inflammatory mediator and may play a role in POCD. The interaction between HMGB1 and TLRs in the perioperative period is not known. We hypothesize that TLR2 contributes to the development of POCD and that HMGB1 regulates TLR2 for this effect. To test these hypotheses, 6- to 8-week old male mice were subjected to right carotid artery exposure under isoflurane anesthesia. CU-CPT22, a TLR1/TLR2 inhibitor, at 3 mg/kg was injected intraperitoneally 30 min before surgery and 1 day after surgery. Glycyrrhizin, a HMGB1 antagonist, at 200 mg/kg was injected intraperitoneally 30 min before surgery. Mice were subjected to Barnes maze and fear conditioning tests from 1 week after surgery. Hippocampus and cerebral cortex were harvested 6 hr or 12 hr after the surgery for Western blotting, ELISA, immunofluorescent staining, and chromatin immunoprecipitation. There were neuroinflammation and impairment of learning and memory in mice with surgery. Surgery increased the expression of TLR2 and TLR4 but not TLR9 in the brain of CD-1 male mice. CU-CPT22 attenuated surgery-induced neuroinflammation and cognitive impairment. Similarly, surgery induced neuroinflammation and cognitive dysfunction in C57BL/6J mice but not in TLR2-/- mice. TLR2 staining appeared in neurons and microglia. Surgery increased HMGB1 in the cell nuclei of the cerebral cortex and hippocampus. Glycyrrhizin ameliorated this increase and the increase of TLR2 in the hippocampus after surgery. Surgery also increased the amount of tlr2 DNA precipitated by an anti-HMGB1 antibody in the hippocampus. Our results suggest that TLR2 contributes to surgery-induced neuroinflammation and cognitive impairment. HMGB1 up-regulates TLR2 expression in the hippocampus after surgery to facilitate this contribution. Thus, TLR2 and HMGB1 are potential targets for reducing POCD.

Critical role of UQCRC1 in embryo survival, brain ischemic tolerance and normal cognition in mice.

Ubiquinol cytochrome c reductase core protein I (UQCRC1) is a component of the complex III in the respiratory chain. Its biological functions are unknown. Here, we showed that knockout of UQCRC1 led to embryonic lethality. Disrupting one UQCRC1 allele in mice (heterozygous mice) of both sexes did not affect their growth but reduced UQCRC1 mRNA and protein in the brain. These mice had decreased complex III formation, complex III activity and ATP content in the brain at baseline. They developed worsened neurological outcome after brain ischemia/hypoxia or focal brain ischemia compared with wild-type mice. The ischemic cerebral cortex of the heterozygous mice had decreased mitochondrial membrane potential and ATP content as well as increased free radicals. Also, the heterozygous mice performed poorly in the Barnes maze and novel object recognition tests. Finally, UQCRC1 was expressed abundantly in neurons and astrocytes. These results suggest a critical role of UQCRC1 in embryo survival. UQCRC1 may also be important by forming the complex III to maintain normal brain ischemic tolerance, learning and memory.

Sevoflurane promotes migration, invasion, and colony-forming ability of human glioblastoma cells possibly via increasing the expression of cell surface protein 44.

Surgical resection of primary solid tumor under anesthesia remains a common practice. It has been concerned whether general anesthetics, especially volatile anesthetics, may promote the growth, migration, and invasion of cancer cells. In this study, we examined the effects of sevoflurane on human glioblastoma cells and determined the role of cluster of differentiation (CD) 44, a cell surface protein involved in cell growth, migration, and invasion, in sevoflurane's effects. We showed that exposure to 1%-4% sevoflurane did not change the cell proliferation, but concentration-dependently increased the invasion of human glioblastoma U251 cells. Furthermore, 4% sevoflurane significantly increased the migration and colony-forming ability of U251 cells. Similar results were observed in human glioblastoma A172 cells. Exposure to sevoflurane concentration-dependently increased the activity of calpains, a group of cysteine proteinases, and CD44 protein in U251 and A172 cells. Knockdown of CD44 with siRNA abolished sevoflurane-induced increases in calpain activity, migration, invasion, and colony-forming ability of U251 cells. Inhalation of 4% sevoflurane significantly increased the tumor volume and invasion/migration distance of U87 cells from the tumor mass in the nude mice bearing human glioblastoma U87 xenograft in the brain. The aggravation by sevoflurane was attenuated by CD44 silencing. In conclusion, sevoflurane increases the migration, invasion, and colony-forming ability of human glioblastoma cells in vitro, and their tumor volume and invasion/migration in vivo. Sevoflurane enhances these cancer cell biology features via increasing the expression of CD44.

Perioperative use of cefazolin ameliorates postoperative cognitive dysfunction but induces gut inflammation in mice.

BACKGROUND: Emerging evidence indicates that long-time use of multiple antibiotics can induce cognitive dysfunction via gut dysbiosis. Cefazolin is often used for 3 to 5 days to prevent perioperative infection. This study is to detect the impact of perioperative use of cefazolin on inflammatory responses and postoperative cognition. METHODS: The anti-inflammatory effect of cefazolin was determined in mouse C8-B4 microglial cells treated with lipopolysaccharide (LPS). Interleukin (IL)-6 and IL-1ß at 6 and 24 h after LPS treatment were detected. Six- to 8-week-old CD-1 mice were subjected to laparotomy. Cefazolin at 300 mg/kg was injected intraperitoneally 1 h before surgery and then once per day for 5 days after surgery. Their learning and memory were assessed by Barnes maze and fear conditioning tests which started 1 week after the surgery. The brain and colon were harvested 24 h and 6 days after surgery to determine inflammatory cytokines. The colon and its luminal contents were harvested 6 and 19 days after surgery for the determination of bacteria flora. Cefazolin concentrations in the serum and brain were measured 0.5, 1, and 2 h after cefazolin injection. RESULTS: IL-6 and IL-1ß levels were decreased by 250 µg/ml cefazolin in the LPS-stimulated C8-B4 cells. Laparotomy increased the time for mice to identify the target hole in the Barnes maze on day 1 and day 8 after training sessions and reduced context-related freezing behavior in fear conditioning test. Cefazolin attenuated these surgical effects but reduced context-related freezing behavior in mice without surgery. IL-6 in the hippocampus and cerebral cortex, IL-1ß in the cerebral cortex, and IL-6 and IL-1ß in the serum and colon were increased 24 h after laparotomy. Cefazolin attenuated these effects. Cefazolin treatment for 5 days in mice without surgery induced colon dysbiosis and increased IL-6 and IL-1ß in the colon and IL-1ß in the cerebral cortex. Colon dysbiosis disappeared in mice treated with cefazolin alone but persisted in mice with surgery and cefazolin 19 days after surgery. High cefazolin concentrations in the serum but not in the brain were detected after cefazolin injection. CONCLUSIONS: These results suggest that cefazolin has a direct anti-inflammatory effect and can attenuate surgery-induced postoperative memory and learning impairment in mice. Cefazolin alone may induce cognitive dysfunction possibly by transient gut dysbiosis in mice without surgery.

Early administration of pyrrolidine dithiocarbamate extends the therapeutic time window of tissue plasminogen activator in a male rat model of embolic stroke.

Tissue plasminogen activator (tPA) is used in fewer than 4% of patients after ischemic stroke because of its narrow therapeutic time window. We tested whether pyrrolidine dithiocarbamate (PDTC), a drug with multiple mechanisms to provide neuroprotection, can be used to extend the therapeutic time window of tPA. Three-month-old male Sprague-Dawley rats were subjected to embolic stroke in the area supplied by the right middle cerebral artery. tPA at 10 mg/kg was given intravenously 4 h after the onset of stroke. PDTC at 50 mg/kg was given via gastric gavage at 30 min or 4 h after the onset of stroke. Two days after the stroke, neurological outcome was evaluated and the right frontal cortex area 1 (Fr1), an ischemic penumbral region, was harvested for analysis. PDTC given at 30 min after the stroke reduced infarct volumes and improved neurological functions no matter whether the rats received tPA. PDTC also reduced tPA-increased hemorrhagic volumes. Consistent with these results, PDTC in the presence or absence of tPA treatment attenuated the increase of proinflammatory cytokines, oxidative stress and matrix metalloprotease 2 activity in the right Fr1. However, PDTC given at 4 h after the onset of stroke did not improve the neurological outcome of rats treated with or without tPA. Our results suggest that PDTC given at an early time point but not in a delayed phase provides neuroprotection against embolic stroke and may be used to extend the therapeutic time window of tPA.

Comparison of Iron-Binding Ability Between Thr70-NapA and Ser70-NapA of Helicobacter pylori.

BACKGROUND: The neutrophil-activating protein (NapA) of Helicobacter pylori (H. pylori), with DNA-binding and iron seizing properties, is a fundamental virulence factor involved in H. pylori-related diseases. Compared with Ser70-NapA strain, Thr70-NapA strain is more intimately correlated with iron-deficiency anemia. METHODS: To investigate whether two types of proteins differ in iron-binding ability, mutated Thr70-NapA and Ser70-NapA strains were established. Isothermal titration calorimetry (ITC) method was conducted to measure the binding between the NapA protein and Fe(2+) . The structural changes of NapA protein were also tested during iron interaction by fast protein liquid chromatography (FPLC) and circular dichroism (CD) methods. DNA-binding assay was performed for evaluate the affinity of both mutated and wild types of NapA with DNA. RESULTS: Mutated Thr70-NapA had higher iron-binding ability than wild Ser70-NapA. The structural stability of Thr70-NapA was disrupted and became more active along with the rising concentration of Fe(2+) , whereas no similar association was observed between Ser70-NapA and Fe(2+) level. When the iron/protein molar ratio ranged from 10 to 20, both Ser70-NapA and Thr70-NapA displayed weaker DNA-binding ability. CONCLUSIONS: Thr70-NapA has much stronger ability to sequester ferrous ion compared with Ser70-NapA in H. pylori. In addition, the DNA-binding property of NapA is dependent upon the Fe(2+) concentration.

Gut microbiota of old mice worsens neurological outcome after brain ischemia via increased valeric acid and IL-17 in the blood.

BACKGROUND: Aging is a significant risk factor for ischemic stroke and worsens its outcome. However, the mechanisms for this worsened neurological outcome with aging are not clearly defined. RESULTS: Old C57BL/6J male mice (18 to 20 months old) had a poorer neurological outcome and more severe inflammation after transient focal brain ischemia than 8-week-old C57BL/6J male mice (young mice). Young mice with transplantation of old mouse gut microbiota had a worse neurological outcome, poorer survival curve, and more severe inflammation than young mice receiving young mouse gut microbiota transplantation. Old mice and young mice transplanted with old mouse gut microbiota had an increased level of blood valeric acid. Valeric acid worsened neurological outcome and heightened inflammatory response including blood interleukin-17 levels after brain ischemia. The increase of interleukin-17 caused by valeric acid was inhibited by a free fatty acid receptor 2 antagonist. Neutralizing interleukin-17 in the blood by its antibody improved neurological outcome and attenuated inflammatory response in mice with brain ischemia and receiving valeric acid. Old mice transplanted with young mouse feces had less body weight loss and better survival curve after brain ischemia than old mice transplanted with old mouse feces or old mice without fecal transplantation. CONCLUSIONS: These results suggest that the gut microbiota-valeric acid-interleukin-17 pathway contributes to the aging-related changes in the outcome after focal brain ischemia and response to stimulus. Valeric acid may activate free fatty acid receptor 2 to increase interleukin-17.

Inhibition of ERK/CREB signaling contributes to postoperative learning and memory dysfunction in neonatal rats.

Exposure to surgery with anesthesia early in life may lead to abnormal behavior, learning, and memory in humans. Pre-clinical studies have suggested a critical role of glial cell-derived neurotrophic factor (GDNF) in these effects. We hypothesize that the inhibition of extracellular signal-regulated kinase (ERK)-cAMP response element-binding protein (CREB) pathway contributes to GDNF decrease and the dysfunction of learning and memory. To address this hypothesis, 7-day-old Sprague-Dawley male and female rats were subjected to right carotid artery exposure (surgery) under sevoflurane anesthesia. Their learning and memory were tested by the Barnes maze, and novel object recognition tests started 23 days after the surgery. Blood and brain were harvested at various times after surgery for biochemical analyses. Rats with surgery and anesthesia performed poorly in the Barnes maze and novel object recognition tests compared with control rats. Rats with surgery had a decreased GDNF concentration in the brain and urine. The concentrations of urine GDNF were negatively correlated with the performance of rats in a delayed memory phase of the Barnes maze test. Surgery increased proinflammatory cytokines in the blood and brain. Intracerebroventricular injection of GDNF attenuated the increased inflammatory response in surgery rats. Surgery inhibited ERK and CREB. Inhibiting ERK reduced GDNF and induced poor performance in the Barnes maze and novel object recognition tests of rats without surgery. Surgery also increased brain-derived natriuretic peptide (BNP) in the brain. Intracerebroventricular injection of BNP inhibited ERK and CREB, reduced GDNF, and impaired learning and memory. Surgery, ERK inhibition, and BNP reduced the expression of synaptic proteins. Our results suggest that surgery increases BNP that inhibits ERK-CREB signaling to reduce GDNF, which leads to an unbalanced inflammatory response and a reduced synaptic protein expression for the development of postoperative cognitive dysfunction. KEY MESSAGES: Surgery increases BNP and decreases ERK/CREB signaling to reduce GDNF. The increase in BNP and decrease in ERK/CREB signaling contribute to postoperative cognitive dysfunction. GDNF reduction contributes to neuroinflammatory response after surgery. Urine GDNF concentrations are negatively corrected with poor spatial memory performance.

Whisker trimming induces anti-anxiety like status via activation of dorsomedial hypothalamus nucleus in mice.

Whiskers are highly developed tactile organs in mice. Here, we showed that mice with whisker trimming had a decreased anxiety behavior and activation of dorsomedial hypothalamus compared to control mice. Inhibition or damage of dorsomedial hypothalamus reversed the decrease of anxiety level induced by whisker trimming. These results expand the role of whiskers in regulating mouse behaviors to anxiety and suggest a novel function of dorsomedial hypothalamus. These findings indicate importance of normal sensory functions in modulating animal behavior.

Activation of the Lateral Habenula-Ventral Tegmental Area Neural Circuit Contributes to Postoperative Cognitive Dysfunction in Mice.

Postoperative cognitive dysfunction (POCD) is common and is associated with poor outcome. Neural circuit involvement in POCD is unknown. Lateral habenula (LHb) that regulates coping and depression-like behaviors after aversive stimuli is activated by surgery in the previous study. Here, surgery activated LHb and ventral tegmental area (VTA) are presented. VTA is known to receive projections from LHb and project to the prefrontal cortex and hippocampus. Direct chemogenetic inhibition of LHb or damaging LHb attenuates surgery-induced learning and memory impairment, N-methyl-d-aspartate (NMDA) receptor activation, endoplasmic reticulum stress, inflammatory responses and cell injury in the VTA, and activation of rostromedial tegmental nucleus, an intermediate station to connect LHb with VTA. LHb inhibition preserves dendritic spine density in the prefrontal cortex and hippocampus. Retrograde inhibition of LHb via its projections to VTA attenuated surgery-induced learning and memory dysfunction is observed. Retrograde activation of LHb induced learning and memory dysfunction is observed. Inhibition of NMDA receptors, dopamine synthesis, and endoplasmic reticulum stress in the VTA reduced surgery-induced learning and memory impairment, inflammatory responses, and cell injury are observed. These results suggest that surgery activates the LHb-VTA neural circuit, which contributes to POCD and neuropathological changes in the brain. These novel findings represent initial evidence for neural circuit involvement in surgery effects.

Phosphoproteome analysis of the pathogenic bacterium Helicobacter pylori reveals over-representation of tyrosine phosphorylation and multiply phosphorylated proteins.

Increasing evidence shows that protein phosphorylation on serine (Ser), threonine (Thr) and tyrosine (Tyr) residues is a major regulatory post-translational modification in the bacteria. To reveal the phosphorylation state in the Gram-negative pathogenic bacterium Helicobacter pylori, we carried out a global and site-specific phosphoproteomic analysis based on TiO(2) -phosphopeptide enrichment and high-accuracy LC-MS/MS determination. Eighty-two phosphopeptides from 67 proteins were identified with 126 phosphorylation sites, among which 79 class I sites were determined to have a distribution of 42.8:38.7:18.5% for the Ser/Thr/Tyr phosphorylation, respectively. The H. pylori phosphoproteome is characterized by comparably big size, high ratio of Tyr phosphorylation, high abundance of multiple phosphorylation sites in individual phosphopeptides and over-representation of membrane proteins. An interaction network covering 28 phosphoproteins was constructed with a total of 163 proteins centering on the major H. pylori virulence factor VacA, indicating that protein phosphorylation in H. pylori may be delicately controlled to regulate many aspects of the metabolic pathways and bacterial virulence.

Role of Sox2 in Learning, Memory, and Postoperative Cognitive Dysfunction in Mice.

Postoperative cognitive dysfunction (POCD) is a significant clinical issue. Its neuropathogenesis has not been clearly identified and effective interventions for clinical use to reduce POCD have not been established. This study was designed to determine whether environmental enrichment (EE) or cognitive enrichment (CE) reduces POCD and whether sex-determining region Y-box-2 regulated by sirtuin 1, plays a role in the effect. Eighteen-month-old male mice were subjected to right-common-carotid-artery exposure under sevoflurane anesthesia. Some of them stayed in cages with EE or CE after the surgery. Learning and memory of mice were tested by a Barnes maze and fear conditioning, starting 2 weeks after the surgery. Sex-determining region Y-box-2 (Sox2) in the brain was silenced by small hairpin RNA (shRNA). Immunofluorescent staining was used to quantify Sox2-positive cells. Surgery reduced Sox2-positive cells in the hippocampus (64 ± 9 cells vs. 91 ± 9 cells in control group, n = 6, p < 0.001) and impaired learning and memory (time to identify target box one day after training sessions in the Barnes maze test: 132 ± 53 s vs. 79 ± 53 s in control group, n = 10, p = 0.040). EE or CE applied after surgery attenuated this reduction of Sox2 cells and POCD. Surgery reduced sirtuin 1 activity and CE attenuated this reduction. Resveratrol, a sirtuin 1 activator, attenuated POCD and surgery-induced decrease of Sox2-positive cells. Silencing shRNA reduced the Sox2-positive cells in the hippocampus and impaired learning and memory in mice without surgery. These results suggest a role of Sox2 in learning, memory, and POCD. EE and CE attenuated POCD via maintaining Sox2-positive cells in the hippocampus.

Norepinephrine inhibits migration and invasion of human glioblastoma cell cultures possibly via MMP-11 inhibition.

PURPOSE: Growing evidence has shown that the stress hormones affect tumor progression. Patients with surgery to remove tumor often have increased norepinephrine during the perioperative period. However, the effect of norepinephrine on the progression of glioblastoma has not yet studied. Therefore, the present study aimed at investigating the effects of norepinephrine on the migration and invasion of the human glioblastoma U87 and U251 cell lines and the mechanism for the effects. METHODS: The U87 and U251 cells were treated with 0, 0.1, 1, 5, 10 or 50 µM norepinephrine. A scratch wound healing assay and a transwell invasion assay were used to investigate cell migration and invasion, respectively. The Human Tumor Metastasis RT2 Profiler PCR Array was used to detect the expression of 84 genes known to be involved in metastasis. RESULTS: Following norepinephrine treatment, the ability of the U87 and U251 cells to migrate and invade was significantly decreased. Human Tumor Metastasis RT2 Profiler PCR Array assay showed that matrix metallopeptidase-11 (MMP-11) was decreased following norepinephrine treatment. The ß-adrenergic receptor blocker (AR) propranolol blunted the suppressive effect of norepinephrine on the migration and invasion of U251 cells but did not have such an effect on the invasion of U87 cells. MMP-11 silencing inhibited the migration and invasion of U87 and U251 cells. The Cancer Genome Atlas data showed that patients with higher expression of MMP-11 in the glioblastoma tissues had poorer prognosis. CONCLUSION: Our results indicate that norepinephrine inhibits the migration and invasion of human glioblastoma cells. This effect may be mediated by the decrease of MMP-11. ß-AR may be a regulatory factor for this effect in U251 cells.

Paraventricular thalamic nucleus plays a critical role in consolation and anxious behaviors of familiar observers exposed to surgery mice.

Background: Consolation behaviors toward the sick are common in humans. Anxiety in the relatives of the sick is also common. Anxiety can cause detrimental effects on multiple systems. However, our understanding on the neural mechanisms of these behaviors is limited because of the lack of small animal models. Methods: Five of 6- to 8-week-old CD-1 male mice were housed in a cage. Among them, 2 mice had right common artery exposure (surgery) and the rest were without surgery. Allo-grooming and performance in light and dark box and elevated plus maze tests of the mice were determined. Results: Mice without surgery had increased allo-grooming toward mice with surgery but decreased allo-grooming toward non-surgery intruders. This increased allo-grooming toward surgery mice was higher in familiar observers of surgery mice than that of mice that were not cage-mates of surgery mice before the surgery. Familiar observers developed anxious behavior after being with surgery mice. Surgery mice with familiar observers had less anxious behavior than surgery mice without interacting with familiar observers. Multiple brain regions including paraventricular thalamic nucleus (PVT) were activated in familiar observers. The activated cells in PVT contained orexin receptors. Injuring the neurons with ibotenic acid, antagonizing orexin signaling with an anti-orexin antibody or inhibiting neurons by chemogenetic approach in PVT abolished the consolation and anxious behaviors of familiar observers. Conclusions: Mice show consolation behavior toward the sick. This behavior attenuates the anxious behavior of surgery mice. The orexin signaling in the PVT neurons play a critical role in the consolation of familiar observers toward surgery mice and their anxious behavior. Considering that about 50 million patients have surgery annually in the United States, our study represents the initial attempt to understand neural mechanisms for consolation and anxiety of a large number of people.

Surgery Trauma Severity but not Anesthesia Length Contributes to Postoperative Cognitive Dysfunction in Mice.

BACKGROUND: Perioperative, modifiable factors contributing to perioperative neurocognitive disorders (PND) have not been clearly defined. OBJECTIVE: To determine the contribution of anesthesia lengths and the degrees of surgical trauma to PND and neuroinflammation, a critical process for PND. METHODS: Three-month-old C57BL/6J mice were subjected to 2 h or 6 h isoflurane anesthesia plus a 5 min or 15 min left common carotid artery exposure (surgery) in a factorial design (two factors: anesthesia with two levels and surgery with three levels). Their learning and memory were tested by Barnes maze and novel object recognition paradigms. Blood, spleen, and hippocampus were harvested for measuring interleukin (IL)-6 and IL-1ß. Eighteen-month-old C57BL/6J mice (old mice) were subjected to 6 h isoflurane anesthesia or 2 h isoflurane anesthesia plus 15 min surgery and then had learning and memory tested. RESULTS: Three-month-old mice with 15 min surgery (long surgery) under 2 h or 6 h anesthesia performed poorly in the learning and memory tests compared with controls. Anesthesia alone or anesthesia plus 5 min surgery did not affect mouse performance in these tests. Similarly, only mice with long surgery but not mice with other experimental conditions had increased IL-6 and IL-1ß in the blood, spleen, and hippocampus and decreased spleen weights. Splenocytes were found in the hippocampus after surgery. Similarly, old mice with long surgery but not the mice with isoflurane anesthesia alone had poor performance in the Barnes maze and novel object recognition tests. CONCLUSION: Surgical trauma, but not anesthesia, contributes to the development of PND and neuroinflammation. Splenocytes may modulate these processes.

Age-Related Upregulation of Carboxyl Terminal Modulator Protein Contributes to the Decreased Brain Ischemic Tolerance in Older Rats.

Stroke remains one of the leading causes of death worldwide. The underlying neuropathology for stroke is ischemic brain injury. Carboxyl terminal modulator protein (CTMP), an endogenous inhibitor of the prosurvival Akt, may increase brain ischemic injury in young animals. Aging decreases brain ischemic tolerance. We hypothesize that CTMP is increased with aging and that this increase contributes to the decreased brain ischemic tolerance. To address these hypotheses, we determined the expression of CTMP and its downstream proteins in the brain of various ages of rats (Fischer 344 and Sprague-Dawley rats). The role of CTMP in ischemic brain injury was investigated by RNA interference. Here, we showed that CTMP in the brain was increased with aging in rats. The phosphorylated/activated Akt was decreased with aging. Six- and 20-month-old rats had poorer neurological outcome than did 2-month-old rats after brain ischemia. The neurological outcome of 2-month-old rats was worsened by LY294002, an Akt inhibitor. The poor neurological outcome in 6-month-old rats was improved by silencing CTMP. CTMP was increased in ischemic penumbral brain tissues. Silencing this increase activated Akt. These results suggest that CTMP increase with aging contributes to the aging-dependent decrease of brain ischemic tolerance.

Critical role of matrix metallopeptidase 9 in postoperative cognitive dysfunction and age-dependent cognitive decline.

BACKGROUND: Postoperative cognitive dysfunction (POCD) is a significant clinical syndrome. Neuroinflammation is an important pathological process for POCD. However, it is not clear how systemic inflammation induced by surgery on peripheral tissues or organs is transmitted into the brain. We determined whether matrix metallopeptidase 9 (MMP9), a protein that can increase blood-brain barrier permeability, is critical in this transmission. The role of MMP9 in age-dependent cognitive decline was also determined. METHODS: Two-month old male C57BL/6J wild-type mice and MMP9-/- mice were randomly assigned to control or surgery groups. The surgery was right carotid artery exposure under isoflurane anesthesia. Cognitive function was tested from one week after the surgery by Barnes maze and fear conditioning. Cognitive function of 2-month old C57BL/6J mice was compared with that of 18-month old mice. RESULTS: Surgery increased the expression of interleukin 1ß, interleukin 6 and ionized calcium binding adapter molecule 1, inflammation indicators, in the brain of the wild-type mice. Blood-brain barrier permeability was increased by surgery. Surgery also impaired the learning and memory of these mice. These surgical effects were absent in the MMP9-/- mice. Eighteen-month old wild-type mice had poorer performance in Barnes maze and fear conditioning tests and lower MMP9 protein expression and activity than did the 2-month old mice. CONCLUSION: MMP9 is critical for transmission of systemic inflammation into the brain for POCD. MMP9 may also play a role in age-dependent cognitive decline.