糖原合成酶激酶3在阿尔茨海默病发病机制中的作用

大量研究报告显示,糖原合成酶激酶-3（glycogen synthase kinase3,GSK3）在阿尔茨海默病（Alzheimer＇s disease,AD）发病机制中起着重要的作用。AD患者存在GSK3活性的增强和水平的提高。细胞培养、无脊椎动物和哺乳类动物模型研究发现,GSK3活性增强导致tau的过度磷酸化、AB产生的增加、学习和记忆能力的缺损,同时伴有神经退行性变。GSK3抑制剂能防止AD转基因动物tau的过度磷酸化,使得GSK3抑制剂有望用于预防和治疗AD。     

轻度认知功能障碍患者淋巴细胞糖原合成酶激酶-3活性的变化

目的 探讨轻度认知功能障碍(MCI)患者淋巴细胞糖原合成酶激酶-3(GSK-3)活性的变化.方法 应用免疫印迹法检测不同病程(1年,2年,3年)MCI患者及正常老年人(正常对照组)外周血淋巴细胞GSK-3B Ser9位点的磷酸化水平及总GSK-3β水平.结果 1年,2年及3年MCI组淋巴细胞GSK-36 Ser9位点磷酸化水平比正常对照组显著降低(均P＜0.01);总GSK-3β水平与正常对照组相比差异无统计学意义;MCI组间淋巴细胞GSK-3β Set9位点磷酸化水平及总GSK-3β水平差异无统计学意义(均P＞0.05).结论 MCI患者淋巴细胞GSK-3的活性升高,对MCI的诊断可能有参考价值.     

Hypoxia activates glycogen synthase kinase-3 in mouse brain in vivo: protection by mood stabilizers and imipramine.

BACKGROUND: Glycogen synthase kinase-3 (GSK3), which is primarily regulated by an inhibitory phosphorylation of an N-terminal serine, has been implicated as contributing to mood disorders by the finding that it is inhibited by the mood stabilizer lithium. METHODS: This study tested if the antidepressant imipramine or the mood stabilizers lithium and sodium valproate regulated pathophysiological serine-dephosphorylation of GSK3 caused by hypoxia in mouse brain in vivo. RESULTS: Hypoxia caused rapid serine-dephosphorylation of both isoforms of GSK3, GSK3beta and GSK3alpha, in mouse cerebral cortex, hippocampus, and striatum. Pretreatment of mice with imipramine, sodium valproate, or lithium attenuated hypoxia-induced serine-dephosphorylation of GSK3beta and GSK3alpha in all three brain regions. CONCLUSIONS: These results demonstrate that imipramine and mood stabilizers are capable of blocking pathophysiologically induced serine-dephosphorylation of GSK3, supporting the hypothesis that stabilization of serine-phosphorylation of GSK3 contributes to their therapeutic effects.     

Glycogen synthase kinase-3beta regulates differentiation-induced apoptosis of human neural progenitor cells

Glycogen synthase kinase-3beta is a multifunctional key regulator enzyme in neural developmental processes and a main component of the canonical Wnt signaling pathway. It is already known that the Wnt-driven differentiation of neural progenitor cells is accompanied by an increase of apoptosis at which the pro-apoptotic function of GSK-3beta is still discussed. The aim of the present study was to investigate whether the phosphorylation level of GSK-3beta at serine 9 is the primary regulatory mechanism of differentiation-induced apoptosis.     

An alternatively spliced form of glycogen synthase kinase-3β is targeted to growing neurites and growth cones

The serine/threonine kinase glycogen synthase kinase-3β (GSK-3β) is expressed in two, alternatively spliced, isoforms: a short form (GSK-3β1) and a long form containing a 13 amino acid insert in the catalytic domain (GSK-3β2). We examined the expression of these isoforms in the rat using specific antibodies and found that GSK-3β2, in contrast to GSK-3β1, is only expressed in the nervous system. The highest levels of GSK-3β2 are found in the developing nervous system but expression persists into adulthood. In the adult central nervous system the highest expression of GSK-3β2 occurs in regions with a high proportion of white matter, suggesting that GSK-3β2 is expressed in axons. Consistent with this finding, sub-cellular fractionation of neonatal rat brain showed that GSK-3β2 is present in fractions enriched in neurites and growth cones. Furthermore, we found that when we separated neuronal cell bodies from neurites by culturing embryonic cortical neurons in neurite outgrowth inserts, GSK-3β2 was present in both compartments. Finally, a rabbit polyclonal antibody raised to the 13 amino acid insert of GSK-3β2 (anti-8A) that specifically recognises GSK-3β2, labels the cell body, including the nucleus, neurites and growth cones of embryonic neurons in culture. To compare functionally the two isoforms, we performed in vitro kinase assays. These showed that GSK-3β1 is more efficient at phosphorylating the microtubule-associated protein MAP1B than GSK-3β2, consistent with previous findings with the microtubule-associated protein tau. However, when co-expressed with MAP1B in COS-7 cells, both GSK-3β isoforms equally efficiently phosphorylated MAP1B and had a similar influence on the regulation of microtubule dynamics by MAP1B in these cells. We conclude that the alternatively spliced isoform of GSK-3β, GSK-3β2, is neuron-specific and has overlapping activities with GSK-3β1.     

Effect of lithium on the circadian rhythms of locomotor activity and glycogen synthase kinase-3 protein expression in the mouse suprachiasmatic nuclei

A circadian clock located in the suprachiasmatic nucleus (SCN) regulates the period of physiological and behavioural rhythms to approximately 24 h. Lithium can lengthen the period of circadian rhythms in most organisms although little is known about the underlying mechanism. In the present study, we examined Drosophila shaggy ortholog glycogen synthase kinase-3 (GSK-3) protein expression in the SCN after lithium treatment. When locomotor activity was assessed, we found an association between the effect of lithium and the period of circadian oscillation as well as the level of GSK-3 protein expression. The decreased expression of GSK-3 and increased expression of phosphorylated GSK-3 (pGSK-3) resulted in an antiphasic circadian rhythm between the two in the SCN of lithium-treated mice housed under both light-dark and constant dark conditions. The enzyme activity of GSK-3 in the SCN was low when the level of pGSK-3 protein was high, as examined by immunoblotting analysis. Thus, GSK-3 enzyme activity has a correlation with the expression of GSK-3 protein in the SCN. Although both GSK-3 and pGSK-3 proteins are also expressed in the arcuate nucleus, lithium did not affect their expression. Based on the association that we found between lengthened circadian period and GSK-3 protein and GSK-3 activity in the SCN, we suggest that GSK-3 plays a role in regulating the period of the mammalian circadian pacemaker.     

Lack of effect of acute,subchronic, or chronic stress on glycogen synthase kinase-3beta protein levels in rat frontal cortex

Glycogen synthase kinase (GSK)-3β is a conserved serine/threonine protein kinase highly abundant in brain tissue. A dominant mechanism by which cells react to stress involves GSK-3β. We studied the effect of stress on GSK-3β levels ex vivo. We have previously found reduced GSK-3β protein levels and GSK-3 activity in postmortem prefrontal cortex of schizophrenic patients. Since schizophrenic patients experience stress more severely than healthy people, we questioned whether their GSK-3β reduction is stress-related using a rat model. Rats were exposed to acute, subchronic, or chronic stress using brief cold restraint. No effect was found on frontal cortex GSK-3β protein levels. These results suggest that reduction in GSK-3β levels in schizophrenic patients is not affected by cold restraint stress and supports the possibility that the changes observed in postmortem brains may be related to the disease.     

Ketamine up-regulates a cluster of intronic miRNAs within the serotonin receptor 2C gene by inhibiting glycogen synthase kinase-3

Objectives: We examined mechanisms that contribute to the rapid antidepressant effect of ketamine in mice that is dependent on glycogen synthase kinase-3 (GSK3) inhibition.

Methods: We measured serotonergic (5HT)-2C-receptor (5HTR2C) cluster microRNA (miRNA) levels in mouse hippocampus after administering an antidepressant dose of ketamine (10?mg/kg) in wild-type and GSK3 knockin mice, after GSK3 inhibition with L803-mts, and in learned helpless mice.

Results: Ketamine up-regulated cluster miRNAs 448-3p, 764-5p, 1264-3p, 1298-5p and 1912-3p (2- to 11-fold). This up-regulation was abolished in GSK3 knockin mice that express mutant constitutively active GSK3. The GSK3 specific inhibitor L803-mts was antidepressant in the learned helplessness and novelty suppressed feeding depression-like behaviours and up-regulated the 5HTR2C miRNA cluster in mouse hippocampus. After administration of the learned helplessness paradigm mice were divided into cohorts that were resilient (non-depressed) or were susceptible (depressed) to learned helplessness. The resilient, but not depressed, mice displayed increased hippocampal levels of miRNAs 448-3p and 1264-3p. Administration of an antagonist to miRNA 448-3p diminished the antidepressant effect of ketamine in the learned helplessness paradigm, indicating that up-regulation of miRNA 448-3p provides an antidepressant action.

Conclusions: These findings identify a new outcome of GSK3 inhibition by ketamine that may contribute to antidepressant effects.     

Response to lithium augmentation in depression is associated with the glycogen synthase kinase 3-beta -50T/C single nucleotide polymorphism.

BACKGROUND: Glycogen synthase kinase 3-beta (GSK3B) is a serine/threonine kinase which is directly inhibited by lithium. A -50T/C single nucleotide polymorphism (SNP) localized within the promoter region of the GSK3B gene has previously been shown to be associated with response to lithium prophylaxis in bipolar disorder. This study investigates the association of the GSK3B -50T/C SNP and response to lithium augmentation in acutely depressed antidepressant nonresponders. METHODS: Eighty-one patients who had not responded to at least one adequate trial of antidepressant monotherapy underwent a standardized trial of lithium augmentation for up to 8 weeks. We genotyped for the GSK3B -50T/C SNP using polymerase chain reaction and restriction fragment length polymorphism methods and investigated the association with remission. RESULTS: The allele frequencies in our sample were CC 14.8%, CT 48.2% and TT 37% (no deviation from the Hardy-Weinberg equilibrium). Carriers of the C-allele of the -50T/C SNP showed a significantly better response to lithium augmentation (hazard ratio: 2.70, p = .007), with a mean remission rate of 56.25% after 4 weeks compared to 31% in patients with the TT-genotype (chi(2) = 4.1; p = .04). CONCLUSIONS: Our results support the finding of recent studies demonstrating a superior response of C-allele carriers with bipolar disorder to lithium prophylaxis.     

Distribution of tau protein kinase I/glycogen synthase kinase-3β, phosphatases 2A and 2B, and phosphorylated tau in the developing rat brain

When trying to elucidate the role played by tau protein kinase I/glycogen synthase kinase-3β (TPKI/GSK-3β) in tau phosphorylation, it is important to consider the balance that exists between the various kinases and phosphatases that are involved in vivo. We studied developmental changes in the expressions of TPKI/GSK-3β and phosphatases 2A and 2B in rat brains using immunoblot analysis. The expression of the kinase peaked postnatally at days 8–11 and returned then to low level after 5 weeks. Phosphatase 2A showed a similar pattern, increasing postnatally until day 14 and decreasing thereafter. On the other hand, phosphatase 2B was undetectable at the juvenile stage, but later its presence increased rapidly to peak at 5 weeks after birth, after which it was maintained at high levels throughout the adult stage. Immunohistochemical studies using the PAP method revealed details of the distribution of TPKI/GSK-3β. At postnatal days 3–21 both gray and white matter were immunoreactive. Later, after 5 weeks, the immunoreactivity became more restricted to the gray matter. The staining of tau phosphorylated at Ser 199, Ser 396, and Ser 413 followed mostly the pattern of the kinase distribution throughout all stages of development. These data, therefore, confirm that TPKI/GSK-3β is expressed primarily in neurons and especially in neurites until postnatal day 21, whereafter the distribution is concentrated mostly in the cell soma and the proximal neurite region.     

碳酸锂联合奥氮平对双相躁狂患者外周血单核细胞糖原合成酶激酶3活性的影响

目的:探讨碳酸锂联合奥氮平治疗对双相躁狂发作患者外周血单核细胞(PBMCs)糖原合成酶激酶3(GSK3)活性的影响. 方法:21例双相Ⅰ型躁狂发作患者给予碳酸锂合并奥氮平治疗8周,于治疗前、治疗4周和8周采集患者外周静脉血20ml,应用免疫印迹法检测PBMCs的GSK3α和GSK3β丝氨酸磷酸化水平以及总GSK3水平...     

氯化锂可恢复脆性X综合征小鼠记忆行为和糖原合成酶激酶3β的活性

In the present study,Fmr1 knockout mice (KO mice) were used as the model for fragile X syndrome.The results of step-through and step-down tests demonstrated that Fmr1 KO mice had shorter latencies and more error counts,indicating a learning and memory disorder.After treatment with 30,60,90,120,or 200 mg/kg lithium chloride,the learning and memory abilities of the Fmr1 KO mice were significantly ameliorated,in particular,the 200 mg/kg lithium chloride treatment had the most significant effect.Western blot analysis showed that lithium chloride significantly enhanced the expression of phosphorylated glycogen synthase kinase 3 beta,an inactive form of glycogen synthase kinase 3 beta,in the cerebral cortex and hippocampus of the Fmr1 KO mice.These results indicated that lithium chloride improved learning and memory in the Fmr1 KO mice,possibly by inhibiting glycogen synthase kinase 3 beta activity.     

Lithium chloride ameliorates learning and memory ability and inhibits glycogen synthase kinase-3 beta activity in a mouse model of fragile X syndrome

In the present study,Fmr1 knockout mice (KO mice) were used as the model for fragile X syndrome.The results of step-through and step-down tests demonstrated that Fmr1 KO mice had shorter latencies and more error counts,indicating a learning and memory disorder.After treatment with 30,60,90,120,or 200 mg/kg lithium chloride,the learning and memory abilities of the Fmr1 KO mice were significantly ameliorated,in particular,the 200 mg/kg lithium chloride treatment had the most significant effect.Western blot an...     

Variation of circadian activity rhythm according to body mass index in children

Background/objectivesThis study aimed to examine the variations of circadian activity rhythm of children according to objective body mass index (BMI) values, using a novel statistical framework (ie, Functional Linear Modeling, FLM), separately for school- and weekend days.MethodsA total of 107 participants (60 females; mean age: 10.25 ± 0.48 years) wore an actigraph for seven days during a regular school-week. While valid actigraphic data during school days were available for each of these children, this number decreased to 53 (31 females; mean age: 10.28 ± 0.51 years) during weekend days.ResultsExamining the school days, significantly higher motor activity in participants with higher BMI was observed from around 4:00 a.m. to 6:00 a.m., with a peak about 5:00 a.m. On the contrary, applying the FLM to the weekend days actigraphic data, no significantly different variation of circadian activity rhythm was observed, according to BMI.ConclusionsIn this specific sample of children, during school days, higher BMI is associated with higher activity level in a specific time window in the second half of nocturnal sleep. The lack of significant findings during weekend days could be explained because of higher variability of get-up time and/or the reduced sample size. Future longitudinal studies could explore if the higher motor activity in that specific time window qualifies as a predictive marker of the development of overweight and obesity. If so, early preventive strategies directed towards those at higher risk could be effectively implemented.     

Rapid antidepressive-like activity of specific glycogen synthase kinase-3 inhibitor and its effect on beta-catenin in mouse hippocampus.

BACKGROUND: Inhibition of glycogen synthase kinase-3 (GSK-3) is thought to be a key feature in the therapeutic mechanism of several mood stabilizers; however, the role of GSK-3 in depressive behavior has not been determined. In these studies, we evaluated the antidepressive effect of L803-mts, a novel GSK-3 peptide inhibitor, in an animal model of depression, the mouse forced swimming test (FST). METHODS: Animals were intracerebroventricularly injected with L803-mts or with respective control peptide (cp) 1 hour, 3 hours, or 12 hours before their subjection to FST. RESULTS: Animals administered L803-mts showed reduced duration of immobility at all three time points tested, as compared with cp-treated animals. Expression levels of beta-catenin, the endogenous substrate of GSK-3, increased in the hippocampus of L803-mts-treated animals by 20%-50%, as compared with cp-treated animals. CONCLUSIONS: Our studies show, for the first time, that in-vivo inhibition of GSK-3 produces antidepressive-like behavior and suggest the potential of GSK-3 inhibitors as antidepressants.     

Defective insulin signaling pathway and increased glycogen synthase kinase-3 activity in the brain of diabetic mice: parallels with Alzheimer's disease and correction by insulin

We have evaluated the effect of peripheral insulin deficiency on brain insulin pathway activity in a mouse model of type 1 diabetes, the parallels with Alzheimer's disease (AD), and the effect of treatment with insulin. Nine weeks of insulin-deficient diabetes significantly impaired the learning capacity of mice, significantly reduced insulin-degrading enzyme protein expression, and significantly reduced phosphorylation of the insulin-receptor and AKT. Phosphorylation of glycogen synthase kinase-3 (GSK3) was also significantly decreased, indicating increased GSK3 activity. This evidence of reduced insulin signaling was associated with a concomitant increase in tau phosphorylation and amyloid beta protein levels. Changes in phosphorylation levels of insulin receptor, GSK3, and tau were not observed in the brain of db/db mice, a model of type 2 diabetes, after a similar duration (8 weeks) of diabetes. Treatment with insulin from onset of diabetes partially restored the phosphorylation of insulin receptor and of GSK3, partially reduced the level of phosphorylated tau in the brain, and partially improved learning ability in insulin-deficient diabetic mice. Our data indicate that mice with systemic insulin deficiency display evidence of reduced insulin signaling pathway activity in the brain that is associated with biochemical and behavioral features of AD and that it can be corrected by insulin treatment.     

Dietary N-3 PUFA deficiency affects sleep-wake activity in basal condition and in response to an inflammatory challenge in mice

Essential polyunsaturated fatty acids (PUFA) from the n-3 and n-6 series constitute the building blocks of brain cell membranes where they regulate most aspects of cell physiology. They are either biosynthesized from their dietary precursors or can be directly sourced from the diet. An overall increase in the dietary n-6/n-3 PUFA ratio, as observed in the Western diet, leads to reduced n-3 PUFAs in tissues that include the brain. Some clinical studies have shown a positive correlation between dietary n-3 PUFA intake and sleep quantity, yet evidence is still sparse. We here used a preclinical model of dietary n-3 PUFA deficiency to assess the precise relationship between dietary PUFA intake and sleep/wake activity. Using electroencephalography (EEG)/electromyography (EMG) recordings on n-3 PUFA deficient or sufficient mice, we showed that dietary PUFA deficiency affects the architecture of sleep-wake activity and the oscillatory activity of cortical neurons during sleep. In a second part of the study, and since PUFAs are a potent modulator of inflammation, we assessed the effect of dietary n-3 PUFA deficiency on the sleep response to an inflammatory stimulus known to modulate sleep/wake activity. We injected mice with the endotoxin lipopolysaccharide (LPS) and quantified the sleep response across the following 12 h. Our results revealed that n-3 PUFA deficiency affects the sleep response in basal condition and after a peripheral immune challenge. More studies are now required aimed at deciphering the molecular mechanisms underlying the intimate relationship between n-3 PUFAs and sleep/wake activity.     

Alteration in kinase activity but not in protein levels of protein kinase B and glycogen synthase kinase-3beta in ventral prefrontal cortex of depressed suicide victims.

BACKGROUND: Past studies in the neurobiology of suicide have reported alterations in serotonin and downstream effectors, such as Akt/protein kinase B. In this study, we aimed to examine possible abnormality in the Akt/glycogen synthase kinase-3beta (GSK-3beta) axis of depressed suicide victims' brains. METHODS: Twenty suicide victims and 20 drug-free non-suicide subjects were included for a postmortem study. The ventral prefrontal cortex area (BA'11) was used, and antemortem diagnoses of major depression disorder (MDD) (DSM-IV) were made from Institution's records. The protein levels of GSK-3alpha/beta and Akt-1 were assayed with the Western blot method, and the kinase activity of Akt and GSK-3alpha/beta were determined by phosphorylation of specific substrates. RESULTS: There was no change either in GSK-3alpha/beta and Akt-1 protein levels or in lithium-inhibitable total GSK-3alpha/beta enzyme activity of the ventral prefrontal cortex. The enzyme activity of Akt decreased significantly [analysis of variance (ANOVA): F(3,36)=5.372; p= .003], whereas GSK-3beta activity increased significantly [ANOVA: F(3,36)=8.567; p= .002] in depressed suicide victims and non-suicide subjects but not in non-depressed suicide victims. CONCLUSIONS: This study indicated that the activity rather than the protein levels of Akt and GSK-3beta was altered. The alteration was associated with MDD rather than with suicide per se.     

Age-independent and age-related deficits in visuospatial learning, sleep-wake states, thermoregulation and motor activity in PDAPP mice

Recent studies demonstrated that mice overexpressing the human mutant beta-amyloid precursor protein (hbetaAPP; PDAPP mice) show age-independent and age-related deficits in spatial learning. We used behavioral and electrophysiological techniques to determine in young and aged PDAPP mice whether deficits in spatial learning also involve alterations in sleep-wake states, thermoregulation and motor activity. Consistent with earlier studies, young PDAPP mice exhibited selective age-independent deficits using spatial, but not random and serial strategies in the circular maze. Aged PDAPP mice exhibited deficits using all search strategies. The core body temperature (Tb) in young and aged PDAPP mice was significantly lower than in age-matched non-transgenic (non-Tg) littermates. During the dark period, the motor activity (LMA) was significantly increased in young PDAPP mice, but not in aged PDAPP mice. During the light period, young PDAPP mice showed a reduction in the generation of rapid-eye-movement (REM) sleep. In contrast, aged PDAPP mice exhibited a reduction in the amount of time spent in W and an increase in SWS during the light period. Aged PDAPP mice also showed an increase in the amount of time spent in W and a reduction in REM sleep during the dark period. Our findings support previous reports indicating deficits in spatial learning in young and aged PDAPP mice. These data also suggest that PDAPP mice exhibit age-independent and age-related deficits in neural mechanisms regulating visuospatial learning, the total amount and the circadian distribution of sleep-wake states, thermoregulation and motor activity.