Activation of the histaminergic H 3 receptor induces phosphorylation of the Akt/GSK-3β pathway in cultured cortical neurons and protects against neurotoxic insults (original) (raw)
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Molecular Pharmacology, 2010
Though dopamine (DA) regulates the serine/threonine kinase, Akt, and its downstream substrate, glycogen synthase kinase-3β (GSK-3β), the direct influence of dopaminergic receptors remains poorly characterised. Short-term incubation of CHO-expressed human (h)D 2L and hD 3 receptors with DA (maximal effect, 5-10 min) phosphorylated Akt (Thr308 and Ser473) and GSK-3β (Ser9), actions blocked by the selective D 2 and D 3 antagonists, L741,626 and S33084, respectively. Similar findings were acquired with the specific D 2 /D 3 receptor agonist, quinelorane, which also enhanced (10 minutes after administration) levels of p-Akt and p-GSK-3β in rat nucleus accumbens, an action blocked by the D 2 /D 3 receptor antagonist, raclopride. Akt and GSK-3β phosphorylation mediated via CHO-expressed hD 2L and hD 3 receptors was prevented by pertussis toxin, and by inhibitors of insulin-like growth factor-1 receptors as well as phosphatidylinositol 3-kinase and Src. Likewise, chelation of intracellular Ca 2+ and interference with an "atypical" phorbol ester-insensitive Protein Kinase C abolished recruitment of Akt and GSK-3β. Inactivation of Protein Kinase Cμ (PKC) blocked Akt and GSK-3β phosphorylation at hD 2L receptors. However, blockade of conventional PKC isoforms attenuated the actions of DA at hD 3 receptors only. Further, Phospholipase C (PLC), calmodulin and Akt inhibitors abolished DA-induced GSK-3β phosphorylation by hD 3 receptors, whereas phosphorylation by hD 2L receptors partially involved calmodulin, Akt and ERK1/2. In conclusion, at both hD 2L and hD 3 receptors, DA elicited a Gi/o-and Ca 2+ /calmodulin-dependent phosphorylation of Akt and GSK-3β via transactivation of insulin-like growth factor 1 receptor. However, significant differences were seen as regards the involvement of PLC, calmodulin and ERK1/2.
Phosphatidylinositol-3-kinase activation blocks amyloid beta-induced neurotoxicity
Toxicology, 2008
The phosphatidylinositol-3-kinase (PI3-K) pathway has been suggested to play a pivotal role in neuronal survival. Although PI3-K has been recently identified as a neuroprotectant, there are no reports regarding the effect of a direct PI3-K activator on A-induced neurotoxicity. We investigated whether direct PI3-K activation prevents A-induced neurotoxicity. To evaluate the effect of A on neuronal cells, we treated primary cultured cortical neurons with several doses of A for 72 h. To investigate the protective effect that PI3-K activation has on A-induced neurotoxicity, cells were simultaneously treated with several doses of a PI3-K activator for 72 h. An MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, trypan blue staining, and DAPI staining showed that A decreased neuronal cell viability in a concentration-dependent manner and also that PI3-K activation effectively prevented A-induced neuronal cell death. A significantly decreased survival signals, including phosphorylated Akt, glycogen synthase kinase-3, and heat shock transcription factor-1. A also increased death signals, such as phosphorylated tau (pThr231) and activated caspase-3. Treatment with a PI3-K activator restored the survival signals and inhibited the death signals. These results suggest that the neurotoxic effect of A can be partially prevented by PI3-K activation.
The Role of Akt-GSK-3β Signaling and Synaptic Strength in Phencyclidine-Induced Neurodegeneration
Neuropsychopharmacology, 2008
N-methyl-D-aspartate (NMDA) receptor antagonists such as phencyclidine (PCP) can induce positive and negative symptoms of schizophrenia in humans and related effects in rodents. PCP treatment of developing rats induces apoptotic neurodegeneration and behavioral deficits later in life that mimic some symptoms of schizophrenia. The precise mechanism of PCP-induced neural degeneration is unknown. This study used selective antagonists, siRNA, and Western analysis to investigate the role of the Akt-glycogen synthase kinase-3b (GSK-3b) pathway in PCP-induced neuronal apoptosis in both neuronal culture and postnatal day 7 rats. PCP administration in vivo and in vitro reduced the phosphorylation of Akt Ser427 and GSK-3b Ser9 , decreasing Akt activity and increasing GSK-3b activity. The alteration of Akt-GSK-3b signaling parallels the temporal profile of caspase-3 activation by PCP. Reducing GSK-3b activity by application of selective inhibitors or depletion of GSK-3b by siRNA attenuates caspase-3 activity and blocks PCP-induced neurotoxicity. Moreover, increasing synaptic strength by either activation of L-type calcium channels with BAY K8644 or potentiation of synaptic NMDA receptors with either a low concentration of NMDA or bicuculline plus 4-aminopyridine completely blocks PCP-induced cell death by increasing Akt phosphorylation. These neuroprotective effects are associated with activation of phosphoinositide-3-kinase-Akt signaling, and to a lesser extent, the MAPK signaling pathway. Overall, these data suggest that PCP-induced hypofunction of synaptic NMDA receptors impairs the Akt-GSK-3b cascade, which is necessary for neuronal survival during development, and that interference with this cascade by PCP or natural factors may contribute to neural pathologies, perhaps including schizophrenia.
Neuropharmacology, 2011
Histamine H 3 receptor antagonists enhance cognition in preclinical models and have been proposed as novel therapeutics for cognitive disorders, in particular Alzheimer's disease (AD). Increased neurotransmitter (e.g. acetylcholine and histamine) release associated with this pharmacology may lead to activation of postsynaptic signaling pathways relevant to cognition and neuroprotection, such as increased phosphorylation of CREB, a transcription factor germane to cognitive function, and the inhibitory residue (Ser-9) of GSK3b, a primary tau kinase associated with AD pathology. In the present studies, acute administration of the H 3-antagonist ABT-239 (0.01e1.0 mg/kg i.p.) increased cortical CREB and S 9-GSK3b phosphorylation in CD1 mice. Donepezil, while increasing CREB phosphorylation, did not increase pS 9-GSK3b expression in contrast to ABT-239. Continuous (2-wk) s.c. infusion of ABT-239 (0.7 mg/kg/day) normalized reduced cortical CREB and hippocampal S 9-GSK3b phosphorylation observed in Tg2576 (APP) AD-transgenic mice. In addition, ABT-239 infusion reversed tau hyperphosphorylation in the spinal cord and hippocampus of TAPP (tau  APP) AD-transgenic mice. Interestingly, ABT-239 produced signaling changes (pS 9-GSK3b) in a7 nicotinic acetylcholine receptor (nAChR) knockout mice. In contrast to wild type, these mice do not exhibit a7 nAChR agonist induced phosphorylation, thus suggesting that H 3-antagonist-mediated signaling is not dependent on AChstimulated a7 nAChR activation. In summary, results of these studies suggest that ABT-239 leads to biochemical signaling that promotes cognitive performance as well as attenuation of tau hyperphosphorylation, raising the intriguing possibility that H 3 antagonists have potential for both symptomatic and disease modifying benefit in the treatment of AD.
The Akt/GSK-3? axis as a new signaling pathway of the histamine H 3 receptor
Journal of Neurochemistry, 2007
Drugs targeting the histamine H3 receptor (H3R) are suggested to be beneficial for the treatment of neurodegenerative diseases, such as Alzheimer’s and Parkinson’s disease. The H3R activates Gi/o-proteins to inhibit adenylyl cyclase activity and modulates phospholipase A2 and MAPK activity. Herein we show that, in transfected SK-N-MC cells, the H3R modulates the activity of the Akt/Glycogen synthase kinase 3β (GSK-3β) axis both in a constitutive and agonist-dependent fashion. H3R stimulation with the H3R agonist immepip induces the phosphorylation of both Ser473 and Thr308 on Akt, a serine/threonine kinase that is important for neuronal development and function. The H3R-mediated activation of Akt can be inhibited by the H3R inverse agonist thioperamide, and by Wortmannin, LY294002 and PTX, suggesting the observed Akt activation occurs via a Gi/o-mediated activation of phosphoinositide-3-kinase. H3R activation also results in the phosphorylation of Ser9 on GSK-3β, which acts downstream of Akt and has a prominent role in brain function. In addition, we show the H3R-mediated phosphorylation of Akt at Ser473 to occur in primary rat cortical neurons and in rat brain slices. The discovery of this signaling property of the H3R adds new understanding to the roles of histamine and the H3R in brain function and pathology.
The Role of PI3K/Akt and ERK in Neurodegenerative Disorders
Neurotoxicity Research, 2019
Disruption of Akt and Erk-mediated signal transduction significantly contributes in the pathogenesis of various neurodegenerative diseases (NDs), such as Parkinson's disease, Alzheimer's diseases, Huntington's disease, and many others. These regulatory proteins serve as the regulator of cell survival, motility, transcription, metabolism, and progression of the cell cycle. Therefore, targeting Akt and Erk pathway has been proposed as a reasonable approach to suppress ND progression. This review has emphasized on involvement of Akt/Erk cascade in the neurodegeneration. Akt has been reported to regulate neuronal toxicity through its various substrates like FOXos, GSK3β, and caspase-9 etc. Akt is also involved with PI3K in signaling pathway to mediate neuronal survival. ERK is another kinase which also regulates proliferation, differentiation, and survival of the neural cell. There has also been much progress in developing a therapeutic molecule targeting Akt and Erk signaling. Therefore, improved understanding of the molecular mechanism behind the regulatory aspect of Akt and Erk networks can make strong impact on exploration of the neurodegenerative disease pathogenesis.
Journal of Pharmacology and Experimental Therapeutics, 2007
6-[(3-Cyclobutyl-2,3,4,5-tetrahydro-1H-3-benzazepin-7-yl)oxy]-N-methyl-3-pyridinecarboxamide hydrochloride (GSK189254) is a novel histamine H 3 receptor antagonist with high affinity for human (pK i ϭ 9.59 -9.90) and rat (pK i ϭ 8.51-9.17) H 3 receptors. GSK189254 is Ͼ10,000-fold selective for human H 3 receptors versus other targets tested, and it exhibited potent functional antagonism (pA 2 ϭ 9.06 versus agonist-induced changes in cAMP) and inverse agonism [pIC 50 ϭ 8.20 versus basal guanosine 5Ј-O-(3-[ 35 S]thio)triphosphate binding] at the human recombinant H 3 receptor. In vitro autoradiography demonstrated specific [ 3 H]GSK189254 binding in rat and human brain areas, including cortex and hippocampus. In addition, dense H 3 binding was detected in medial temporal cortex samples from severe cases of Alzheimer's disease, suggesting for the first time that H 3 receptors are preserved in late-stage disease. After oral administration, GSK189254 inhibited cortical ex vivo R-(Ϫ)-␣-methyl[imidazole-2,5(n)-3 H]histamine dihydrochloride ([ 3 H]R-Article, publication date, and citation information can be found at
Role of glycogen synthase kinase-3 in ketamine-induced developmental neuroapoptosis in rats
British Journal of Anaesthesia, 2013
† Ketamine increased neuroapoptosis in neonatal rat pups, which correlated with decreased GSK-3b phosphorylation. † This suggests involvement of this cell survival signalling pathway in anaesthetic neurotoxicity. † Co-administration of lithium mitigated ketamine-induced neurotoxicity and reduction in GSK-3b phosphorylation, providing a potential approach to neuroprotection. Background. Ketamine-induced neuroapoptosis has been attributed to diverse stressrelated mechanisms. Glycogen synthase kinase-3b (GSK-3b) is a multifunctional kinase that is active in neuronal development and linked to neurodegenerative disorders. We hypothesized that ketamine would enhance GSK-3b-induced neuroapopotosis, and that lithium, an inhibitor of GSK-3b, would attenuate this response in vivo. Methods. Protein levels of cleaved caspase-3, protein kinase B (AKT), GSK-3b, and cyclin D1 were measured in post-natal day 7 rat pups after 1.5, 3, 4.5, and 6 h exposure to ketamine. A cohort of rat pups was randomized to a 6 h exposure to ketamine with and without lithium. Neuroapoptosis was measured by cleaved caspase-3 and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling staining by immunohistochemistry. Protein levels of cleaved caspase-3 and-9 and the total and phosphorylated forms of AKT, GSK-3b, and cyclin D1 (cell cycle protein) were also measured. Results. Ketamine produced a duration-dependent increase in cleaved caspase-3 and cyclin D1, which corresponded to decreases in phosphorylated AKT and GSK-3b. Co-administration of lithium with ketamine attenuated this response. Conclusions. Ketamine-induced neuroapoptosis is associated with a temporal decrease in GSK-3b phosphorylation, and simultaneous administration of lithium mitigated this response. These findings suggest that GSK-3b is activated during this ketamine-induced neuroapoptosis.
Glycogen synthase kinase 3 (GSK3 ) mediates 6-hydroxydopamine-induced neuronal death
Faseb Journal, 2004
The causes of sporadic Parkinson's disease (PD) are poorly understood. 6-Hydroxydopamine (6-OHDA), a PD mimetic, is widely used to model this neurodegenerative disorder in vitro and in vivo; however, the underlying mechanisms remain incompletely elucidated. We demonstrate here that 6-OHDA evoked endoplasmic reticulum (ER) stress, which was characterized by an up-regulation in the expression of GRP78 and GADD153 (Chop), cleavage of procaspase-12, and phosphorylation of eukaryotic initiation factor-2 α in a human dopaminergic neuronal cell line (SH-SY5Y) and cultured rat cerebellar granule neurons (CGNs). Glycogen synthase kinase-3 β (GSK3β) responds to ER stress, and its activity is regulated by phosphorylation. 6-OHDA significantly inhibited phosphorylation of GSK3β at Ser9, whereas it induced hyperphosphorylation of Tyr216 with little effect on GSK3β expression in SH-SY5Y cells and PC12 cells (a rat dopamine cell line), as well as CGNs. Furthermore, 6-OHDA decreased the expression of cyclin D1, a substrate of GSK3β, and dephosphorylated Akt, the upstream signaling component of GSK3β. Protein phosphatase 2A (PP2A), an ER stress-responsive phosphatase, was involved in 6-OHDA-induced GSK3β dephosphorylation (Ser9). Blocking GSK3β activity by selective inhibitors (lithium, TDZD-8, and L803-mts) prevented 6-OHDA-induced cleavage of caspase-3 and poly(ADP-ribose) polymerase (PARP), DNA fragmentations and cell death. With a tetracycline (Tet)-controlled TrkB inducible system, we demonstrated that activation of TrkB in SH-SY5Y cells alleviated 6-OHDA-induced GSK3β dephosphorylation (Ser9) and ameliorated 6-OHDA neurotoxicity. TrkB activation also protected CGNs against 6-OHDA-induced damage. Although antioxidants also offered neuroprotection, they had little effect on 6-OHDA-induced GSK3β activation. These results suggest that GSK3β is a critical intermediate in pro-apoptotic signaling cascades that are associated with neurodegenerative diseases, thus providing a potential target site amenable to pharmacological intervention.