Amelioration of Hippocampal Insulin Resistance Reduces Tau Hyperphosphorylation and Cognitive Decline Induced by Isoflurane in Mice.

General anesthetics can induce cognitive impairments and increase the risk of Alzheimer's disease (AD). However, the underlying mechanisms are still unknown. Our previous studies shown that long-term isoflurane exposure induced peripheral and central insulin resistance (IR) in adult mice and aggravated IR in type 2 diabetes mellitus (T2DM) mice. Clinical and preclinical studies revealed an association between impaired insulin signaling and tau pathology in AD and other tauopathies. We investigated if alleviation of hippocampal IR by the antidiabetic agent metformin could reduce tau hyperphosphorylation and cognitive decline induced by isoflurane in mice. The effects of prolonged (6 h) isoflurane anesthesia on hippocampal IR, hippocampal tau hyperphosphorylation, and hippocampus-dependent cognitive function were evaluated in wild type (WT) adult mice and the high-fat diet plus streptozotocin (HFD/STZ) mouse model of T2DM. Here we shown that isoflurane and HFD/STZ dramatically and synergistically induced hippocampal IR and fear memory impairment. Metformin pretreatment strongly ameliorated hippocampal IR and cognitive dysfunction caused by isoflurane in WT mice, but was less effective in T2DM mice. Isoflurane also induced hippocampal tau hyperphosphorylation and metformin reversed this effect. In addition, isoflurane significantly increased blood glucose levels in both adult and T2DM mice, and metformin reversed this effect as well. Administration of 25% glucose to metformin-pretreated mice induced hyperglycemia, but surprisingly did not reverse the benefits of metformin on hippocampal insulin signaling and fear memory following isoflurane anesthesia. Our findings show hippocampal IR and tau hyperphosphorylation contribute to acute isoflurane-induced cognitive dysfunction. Brief metformin treatment can mitigate these effects through a mechanism independent of glycemic control. Future studies are needed to investigate whether long-term metformin treatment can also prevent T2DM-induced hippocampal IR and cognitive decline.

Protective effects of grape seed procyanidin on isoflurane-induced cognitive impairment in mice.

Context: Oxidative imbalance-induced cognitive impairment is among the most urgent clinical concerns. Isoflurane has been demonstrated to impair cognitive function via an increase in oxidative stress. GSP has strong antioxidant capacities, suggesting potential cognitive benefits.Objective: This study investigates whether GSP pre-treatment can alleviate isoflurane-induced cognitive dysfunction in mice.Materials and methods: C57BL/6J mice were pre-treated with either GSP 25-100 mg/kg/d for seven days or GSP 100-400 mg/kg as a single dose before the 6 h isoflurane anaesthesia. Cognitive functioning was examined using the fear conditioning tests. The levels of SOD, p-NR2B and p-CREB in the hippocampus were also analysed.Results: Pre-treatment with either a dose of GSP 50 mg/kg/d for seven days or a single dose of GSP 200 mg/kg significantly increased the % freezing time in contextual tests on the 1st (72.18 ± 12.39% vs. 37.60 ± 8.93%; 78.27 ± 8.46% vs. 52.72 ± 2.64%), 3rd (93.80 ± 7.62% vs. 52.94 ± 14.10%; 87.65 ± 10.86% vs. 52.89 ± 1.73%) and 7th (91.36 ± 5.31% vs. 64.09 ± 14.46%; 93.78 ± 3.92% vs. 79.17 ± 1.79%) day after anaesthesia. In the hippocampus of mice exposed to isoflurane, GSP 200 mg/kg increased the total SOD activity on the 1st and 3rd day and reversed the decreased activity of the NR2B/CREB pathway.Discussion and conclusions: These findings suggest that GSP improves isoflurane-induced cognitive dysfunction by protecting against perturbing antioxidant enzyme activities and NR2B/CREB pathway. Therefore, GSP may possess a potential prophylactic role in isoflurane-induced and other oxidative stress-related cognitive decline.

Sevoflurane plays a reduced role in cognitive impairment compared with isoflurane: limited effect on fear memory retention.

Isoflurane and sevoflurane are both inhalation anesthetics, but in clinical application, sevoflurane has been considered to be less suitable for long-term anesthesia because of its catabolic compounds and potential nephrotoxicity. Nevertheless, recent studies have shown that these two inhalation anesthetics are similar in hepatorenal toxicity, cost, and long-term anesthetic effect. Moreover, sevoflurane possibly has less cognitive impact on young mice. In this study, C57BL/6 mice aged 8-10 weeks were exposed to 1.2% isoflurane or 2.4% sevoflurane for 6 hours. Cognitive function and memory were examined in young mice using the novel object recognition, contextual fear conditioning, and cued-fear extinction tests. Western blot assay was performed to detect expression levels of D1 dopamine receptor, catechol-O-methyltransferase, phospho-glycogen synthase kinase-3ß, and total glycogen synthase kinase-3ß in the hippocampus. Our results show that impaired performance was not detected in mice exposed to sevoflurane during the novel object recognition test. Contextual memory impairment in the fear conditioning test was shorter in the sevoflurane group than the isoflurane group. Long-term sevoflurane exposure did not affect memory consolidation, while isoflurane led to memory consolidation and reduced retention. Downregulation of hippocampal D1 dopamine receptors and phosphorylated glycogen synthase kinase-3ß/total glycogen synthase kinase-3ß and upregulation of catechol-O-methyltransferase may be associated with differing memory performance after exposure to isoflurane or sevoflurane. These results confirm that sevoflurane has less effect on cognitive impairment than isoflurane, which may be related to expression of D1 dopamine receptors and catechol-O-methyltransferase and phosphorylation of glycogen synthase kinase-3ß in the hippocampus. This study was approved by the Institutional Animal Care and Use Committee, Nanjing University, China on November 20, 2017 (approval No. 20171102).

GLYX-13 pretreatment ameliorates long-term isoflurane exposure-induced cognitive impairment in mice.

Accumulating evidence indicates that inhalation anesthetics induce or increase the risk of cognitive impairment. GLYX-13 (rapastinel) acts on the glycine site of N-methyl-D-aspartate receptors (NMDARs) and has been shown to enhance hippocampus-dependent learning and memory function. However, the mechanisms by which GLYX-13 affects learning and memory function are still unclear. In this study, we investigated these mechanisms in a mouse model of long-term anesthesia exposure. Mice were intravenously administered 1 mg/kg GLYX-13 at 2 hours before isoflurane exposure (1.5% for 6 hours). Cognitive function was assessed using the contextual fear conditioning test and the novel object recognition test. The mRNA expression and phosphorylated protein levels of NMDAR pathway components, N-methyl-D-aspartate receptor subunit 2B(NR2B)-Ca2+/calmodulin dependent protein kinase II (CaMKII)-cyclic adenosine monophosphate response element binding protein (CREB), in the hippocampus were evaluated by quantitative RT-PCR and western blot assay. Pretreatment with GLYX-13 ameliorated isoflurane exposure-induced cognitive impairment and restored NR2B, CaMKII and CREB mRNA and phosphorylated protein levels. Intracerebroventricular injection of KN93, a selective CaMKII inhibitor, significantly diminished the effect of GLYX-13 on cognitive function and NR2B, CaMKII and CREB levels in the hippocampus. Taken together, our findings suggest that GLYX-13 pretreatment alleviates isoflurane-induced cognitive dysfunction by protecting against perturbation of the NR2B/CaMKII/CREB signaling pathway in the hippocampus. Therefore, GLYX-13 may have therapeutic potential for the treatment of anesthesia-induced cognitive dysfunction. This study was approved by the Experimental Animal Ethics Committee of Drum Tower Hospital affiliated to the Medical College of Nanjing University, China (approval No. 20171102) on November 20, 2017.

Photoacoustic treatment mitigates cognitive dysfunction in a model of sleep-wake rhythm disturbance.

Sleep-wake rhythm disturbances, which are characterized by abnormal sleep timing or duration, are associated with cognitive dysfunction. Photoacoustic treatments including light and sound stimulation have been found to be effective in modulating sleep patterns and improving cognitive behavior in abnormal sleep-wake pattern experiments. In this study, we examined whether light and sound interventions could reduce sleep-wake pattern disturbances and memory deficits in a sleep rhythm disturbance model. We established a model of sleep rhythm disturbance in C57BL/6J mice via a sleep deprivation method involving manual cage tapping, cage jostling, and nest disturbance. We used a Mini Mitter radio transmitter device to monitor motor activity in the mice and fear conditioning tests to assess cognitive function. Our results indicated that an intervention in which the mice were exposed to blue light (40-Hz flickering frequency) for 1 hour during their subjective daytime significantly improved the 24-hour-acrophase shift and reduced the degree of memory deficit induced by sleep deprivation. However, interventions in which the mice were exposed to a 40-Hz blue light at offset time or subjective night time points, as well as 2 Hz-blue light at 3 intervention time points (subjective day time, subjective night time, and offset time points), had no positive effects on circadian rhythm shift or memory deficits. Additionally, a 2000-Hz sound intervention during subjective day time attenuated the 24-hour-acrophase shift and memory decline, while 440-Hz and 4000-Hz sounds had no effect on circadian rhythms. Overall, these results demonstrate that photoacoustic treatment effectively corrected abnormal sleep-wake patterns and cognitive dysfunction associated with sleep-deprivation-induced disturbances in sleep-wake rhythm. All animal experiments were approved by the Experimental Animal Ethics Committee of Drum Tower Hospital Affiliated to the Medical College of Nanjing University, China (approval No. 20171102) on November 20, 2017.

Green tea polyphenols improve isoflurane-induced cognitive impairment via modulating oxidative stress.

Anesthetic exposure induces learning and memory impairment and the mechanisms remain unknown. Green tea polyphenols(GTP) have been reported to be neuroprotective. The present study was performed to examine the therapeutic potential of GTP on isoflurane-induced cognitive deficits. Six-week-old male C57BL/6J mice were treated with 1.6% isoflurane for 6 hours. Multiple-dose of GTP at 25 mg/kg for 7 consecutive days and single-dose at 75 mg/kg on the 7th day were respectively administered intraperitoneally to model mice before anesthesia. Fear conditioning test and novel objection recognition were conducted to assess cognition of mice. Superoxide dismutase (SOD) was evaluated using assay kits. Protein expression levels of right hippocampus p-CaMKII, p-CREB and BDNF were examined by Western blot. Our results indicated that 6 h isoflurane anesthesia induced cognitive impairment in early 3 days. Meanwhile, the hippocampus SOD declined in step. The expression levels of p-CaMKII, p-CREB and BDNF were also downregulated. GTP 25mg/kg per day significantly attenuated cognitive dysfunction on Day 3 following isoflurane anesthesia. Moreover, GTP 25mg/kg per day effectively mitigated isodlurane-induced declines of SOD, as well as the p-CaMKII, p-CREB and BDNF levels. However, single-dose at 75 mg/kg of GTP had no significant effects. This study indicated that GTP attenuate isoflurane-induced cognition impairment and this positive effects may be related to its antioxidant properties.

Misaligned Feeding May Aggravate Pain by Disruption of Sleep-Awake Rhythm.

BACKGROUND: Increasing evidence suggests that patients with eating disorders are more likely to develop chronic pain. A misaligned diet has been reported to disrupt the sleep-awake rhythms. Combined with our previous investigation on circadian pain, we aimed to investigate the role of misaligned diet in the pain sensitivity and the underlying mechanisms. METHODS: Two-month-old C57BL/6J male mice were administered chronic constriction injury (CCI) surgery to establish neuropathic pain models. CCI mice were randomized to scheduled food access throughout the whole day (CCI-free), during the daytime (CCI-misaligned), and at night (CCI-aligned), respectively. The paw withdrawal mechanical threshold, indicating pain behavior, was measured by Von Frey. The gross motor activity pattern indicating the sleep-awake rhythm was monitored by Mini-Mitter. Melatonin (Mel) was administered to ameliorate the sleep-awake rhythm (CCI-free + Mel and CCI-misaligned + Mel). The expressions of circadian pain-related proteins were detected by quantitative polymerase chain reaction and western blot. The primary outcome is the pain threshold and the secondary outcome is the sleep-awake rhythm. RESULTS: Misaligned diet during the peri-CCI surgery period significantly decreased the paw withdrawal mechanical threshold compared with the CCI-free mice (day 14: 0.40 ± 0.09 vs 0.64 ± 0.15; P = .03;) and altered the sleep-awake rhythm. Mel pretreatment alleviated the increased pain (day 14, CCI-misaligned + Mel versus CCI-misaligned: day 14: 0.60 ± 0.13 vs 0.35 ± 0.12; P = .022) and the disrupted sleep-awake rhythm caused by misaligned feeding. The mRNA levels of N-methyl-D-aspartate receptor subtype 2B (NR2B), Ca/calmodulin-dependent protein kinase II (CaMKII), and cyclic adenosine monophosphate-response element binding protein (CREB) in the spinal dorsal horn increased in CCI-misaligned mice compared with the CCI-free mice. The phosphor-NR2B, phosphor-CaMKII, and phosphor-CREB also increased in CCI-misaligned mice compared with the CCI-free mice. However, the expressions of NR2B, CaMKII, and CREB were decreased in CCI-misaligned + Mel mice compared to CCI-misaligned mice at both transcriptional and translational levels. CONCLUSIONS: Misaligned diet might aggravate pain sensitivity through the disruption of the sleep-awake cycle, which could be recovered by Mel. NR2B-CaMKII-CREB may participate in the disruption of sleep-awake rhythm-mediated pain aggravation.

[Preparation and characterization of monoclonal antibody against gp120 V1/V2 domain of HIV-1 subtype CRF01_AE].

Objective To express the V1/V2 domain of HIV-1 subtype CRF01_AE gp120 in eukaryotic cells, and then prepare its monoclonal antibody (mAb) and identify its antigen reactivity. Methods Eukaryotic expression vector of pTriEx-3-V1/V2CNE55 was constructed and transiently transfected into HEK293F cell line. The V1/V2-His chimera protein was purified and injected into BALB/c mice. After the fusion between spleen cells of immunized BALB/c mice and myeloma cells SP 2/0, ELISA was used for screening the positive hybridoma cell clones against the V1/V2 recombinant protein. The specificity, titer and type of its mAb were characterized. Results We obtained a stable hybridoma cell line which secreted anti-HIV-1 AE subtype gp120 V1/V2 domain mAb. The ascite titer of the mAb was 1:81 000, and the type of the mAb was IgG1/κ. Western blotting showed that the mAb could recognize recombinant HIV-1 gp120 of different HIV-1 subtypes. Conclusion The study prepared successfully the anti-HIV-1 V1/V2 domain mAb.