MLK3‐MKK3/6‐P38MAPK cascades following N‐methyl‐D‐aspartate receptor activation contributes to amyloid‐β peptide‐induced apoptosis in SH‐SY5Y cells

Amyloid‐β peptide (Aβ) has been implicated in the development of Alzheimer's disease (AD), but the underlying molecular mechanisms remain unclear. The present study explores the proapoptosis signaling evoked by N‐methyl‐D‐aspartate (NMDA) receptors in Aβ neurotoxicity. Oligomeric Aβ25–35 incubation resulted in significant apoptosis of neuronal SH‐SY5Y cells. Preadministration of the potent NMDA receptor antagonist MK801 promoted neuronal survival. Both NVP‐AAM077 and Ro25–6981, GluN2A‐ and GluN2B‐subunit‐selective NMDA receptor antagonists, respectively, showed effects similar to those of MK801, supporting a critical role of GluN2A‐ or GluN2B‐containing NMDA receptors in Aβ neurotoxicity. Exposure to oligomeric Aβ25–35 increased the phosphorylation (activation) of mixed‐lineage kinase 3 (MLK3), dual‐specific mitogen‐activated protein kinase kinase 3/6 (MKK3/6), and P38 mitogen‐activated protein kinase (P38MAPK) in SH‐SY5Y cells. Inhibition of P38MAPK activation by SB239063 had a neuroprotective effect. K252a attenuated the phosphorylation of MLK3, MKK3/6, and P38MAPK but also partially prevented SH‐SY5Y cells apoptosis. MK801, NVP‐AAM077, and Ro25–6981, abrogated the MLK3‐MKK3/6‐P38MAPK activation induced by oligomeric Aβ25–35. These results suggest that the activation of GluN2A‐ or GluN2B‐containing NMDA receptors is responsible for the activation of MLK3‐MKK3/6‐P38MAPK cascades, which contributes to Aβ‐mediated cell apoptosis. © 2014 Wiley Periodicals, Inc.     

Glutamate protects neuromuscular junctions from deleterious effects of β‐amyloid peptide and conversely: An in vitro study in a nerve–muscle coculture

Murine models of Alzheimer's disease with elevated levels of amyloid‐β (Aβ) peptide present motor axon defects and neuronal death. Aβ_1–42 accumulation is observed in motor neurons and spinal cords of sporadic and familial cases of amyotrophic lateral sclerosis (ALS). Motor neurons are highly susceptible to glutamate, which has a role in ALS neuronal degeneration. The current study investigates the link between Aβ and glutamate in this neurodegenerative process. Primary rat nerve and human muscle cocultures were intoxicated with glutamate or Aβ. Neuromuscular junction (NMJ) mean size and neurite length were evaluated. The role of N‐methyl‐D‐aspartate receptor (NMDAR) was investigated by using MK801. Glutamate and Aβ production were evaluated in culture supernatant. The current study shows that NMJs are highly sensitive to Aβ peptide, that the toxic pathway involves glutamate and NMDAR, and that glutamate and Aβ act in an interlinked manner. Some motor diseases (e.g., ALS), therefore, could be considered from a new point of view related to these balance disturbances. © 2014 Wiley Periodicals, Inc.     

Memantine protects rat cortical cultured neurons against β‐amyloid‐induced toxicity by attenuating tau phosphorylation

It has been suggested that accumulation of beta‐amyloid (Aβ) peptide triggers neurodegeneration, at least in part, via glutamate‐mediated excitotoxicity in Alzheimer’s disease (AD) brain. This is supported by observations that toxicity induced by Aβ peptide in cultured neurons and in adult rat brain is known to be mediated by activation of glutamatergic N‐methyl‐d ‐aspartate (NMDA) receptors. Additionally, recent clinical studies have shown that memantine, a noncompetitive NMDA receptor antagonist, can significantly improve cognitive functions in some AD patients. However, very little is currently known about the potential role of memantine against Aβ‐induced toxicity. In the present study, we have shown that Aβ_1–42‐induced toxicity in rat primary cortical cultured neurons is accompanied by increased extracellular and decreased intracellular glutamate levels. We subsequently demonstrated that Aβ toxicity is induced by increased phosphorylation of tau protein and activation of tau kinases, i.e. glycogen synthase kinase‐3β and extracellular signal‐related kinase 1/2. Additionally, Aβ treatment induced cleavage of caspase‐3 and decreased phosphorylation of cyclic AMP response element binding protein, which are critical in determining survival of neurons. Memantine treatment significantly protected cultured neurons against Aβ‐induced toxicity by attenuating tau‐phosphorylation and its associated signaling mechanisms. However, this drug did not alter either conformation or internalization of Aβ_1–42 and it was unable to attenuate Aβ‐induced potentiation of extracellular glutamate levels. These results, taken together, provide new insights into the possible neuroprotective action of memantine in AD pathology.     

Memantine lowers amyloid‐β peptide levels in neuronal cultures and in APP/PS1 transgenic mice

Memantine is a moderate‐affinity, uncompetitive N‐methyl‐D‐aspartate (NMDA) receptor antagonist that stabilizes cognitive, functional, and behavioral decline in patients with moderate to severe Alzheimer's disease (AD). In AD, the extracellular deposition of fibrillogenic amyloid‐β peptides (Aβ) occurs as a result of aberrant processing of the full‐length Aβ precursor protein (APP). Memantine protects neurons from the neurotoxic effects of Aβ and improves cognition in transgenic mice with high brain levels of Aβ. However, it is unknown how memantine protects cells against neurodegeneration and affects APP processing and Aβ production. We report the effects of memantine in three different systems. In human neuroblastoma cells, memantine, at therapeutically relevant concentrations (1–4 μM), decreased levels of secreted APP and Aβ_1–40. Levels of the potentially amylodogenic Aβ_1–42 were undetectable in these cells. In primary rat cortical neuronal cultures, memantine treatment lowered Aβ_1–42 secretion. At the concentrations used, memantine treatment was not toxic to neuroblastoma or primary cultures and increased cell viability and/or metabolic activity under certain conditions. In APP/presenilin‐1 (PS1) transgenic mice exhibiting high brain levels of Aβ_1–42, oral dosing of memantine (20 mg/kg/day for 8 days) produced a plasma drug concentration of 0.96 μM and significantly reduced the cortical levels of soluble Aβ_1–42. The ratio of Aβ_1–40/Aβ_1–42 increased in treated mice, suggesting effects on the γ‐secretase complex. Thus, memantine reduces the levels of Aβ peptides at therapeutic concentrations and may inhibit the accumulation of fibrillogenic Aβ in mammalian brains. Memantine's ability to preserve neuronal cells against neurodegeneration, to increase metabolic activity, and to lower Aβ level has therapeutic implications for neurodegenerative disorders. © 2009 Wiley‐Liss, Inc.     

A novel antibody targeting sequence 31–35 in amyloid β protein attenuates Alzheimer's disease‐related neuronal damage

Amyloid β protein (Aβ) plays a critical role in pathogenesis of Alzheimer's disease (AD). Our previous studies indicated that the sequence 31–35 in Aβ molecule is an effective active center responsible for Aβ neurotoxicity in vivo and in vitro. In the present study, we prepared a novel antibody specifically targeting the sequence 31–35 of amyloid β protein, and investigated the neuroprotection of the anti‐Aβ_31–35 antibody against Aβ_1–42‐induced impairments in neuronal viability, spatial memory, and hippocampal synaptic plasticity in rats. The results showed that the anti‐Aβ_31–35 antibody almost equally bound to both Aβ_31–35 and Aβ_1–42, and pretreatment with the antibody dose‐dependently prevented Aβ_1–42‐induced cytotoxicity on cultured primary cortical neurons. In behavioral study, intracerebroventricular (i.c.v.) injection of anti‐Aβ_31–35 antibody efficiently attenuated Aβ_1–42‐induced impairments in spatial learning and memory of rats. In vivo electrophysiological experiments further indicated that Aβ_1–42‐induced suppression of hippocampal synaptic plasticity was effectively reversed by the antibody. These results demonstrated that the sequence 31–35 of Aβ may be a new therapeutic target, and the anti‐Aβ_31–35 antibody could be a novel immunotheraputic approach for the treatment of AD. © 2016 Wiley Periodicals, Inc.     

β‐Amyloid induced effects on cholinergic,serotonergic, and dopaminergic neurons is differentially counteracted by anti‐inflammatory drugs

β‐Amyloid (Aβ) is a small peptide that plays a potent role in synaptic plasticity as well as forms amyloid plaques in Alzheimer's disease (AD). Recent studies suggest that Aβ deposition is deleterious not only in AD, but also in Parkinson's disease (PD) and depression. This Aβ effect is associated with inflammatory processes. However, further evaluation is needed to understand how Aβ and inflammation interact and contribute to the regulation of the cholinergic, serotonergic, and dopaminergic neuronal populations. The aim of the present study was to investigate the effects of Aβ_1–42 on cholinergic neurons of the nucleus basalis of Meynert (which degenerate in AD), on serotonergic neurons of the dorsal raphe nucleus (which play a role in depression), and on dopaminergic neurons of the ventral mesencephalon (which degenerate in PD) in rat organotypic brain slices. Furthermore, we investigated whether anti‐inflammatory drugs (celecoxib, citalopram, cyclooxygenase‐2 inhibitor, ibuprofen, indomethacin, piclamilast) modulate or counteract Aβ‐induced effects. Two‐week‐old organotypic brain slices of the nucleus basalis of Meynert, dorsal raphe nucleus, and ventral mesencephalon were incubated with 50 ng/ml Aβ_1–42 with or without anti‐inflammatory agents for 3 days. Our results reveal that Aβ significantly decreased the number of choline acetyltransferase‐positive cholinergic, tryptophan hydroxylase‐positive serotonergic, and tyrosine hydroxylase‐positive dopaminergic neurons and that anti‐inflammatory drugs partially counteracted the Aβ‐induced neuronal decline. This decline was not due to apoptotic processes (as evaluated by TUNEL, propidium iodide, caspase), oxidative stress (as measured by nitrite, catalase, or superoxide dismutase‐2), or inflammation, but was most likely caused by a downregulation of these key enzymes. © 2012 Wiley Periodicals, Inc.     

β‐amyloid and ATP‐induced diffusional trapping of astrocyte and neuronal metabotropic glutamate type‐5 receptors

β‐Amyloid (Aβ) oligomers initiate synaptotoxicity following their interaction with the plasma membrane. Several proteins including metabotropic glutamate type 5 receptors (mGluR5s) contribute to this process. We observed an overexpression of mGluR5s in reactive astrocytes surrounding Aβ plaques in brain sections from an Alzheimer's disease mouse model. In a simplified cell culture system, using immunocytochemistry and single molecule imaging, we demonstrated a rapid binding of Aβ oligomers on the plasma membrane of astrocytes. The resulting aggregates of Aβ oligomers led to the diffusional trapping and clustering of mGluR5s. Further, Aβ oligomers induced an increase in ATP release following activation of astroglial mGluR5s by its agonist. ATP slowed mGluR5s diffusion in astrocytes as well as in neurons co‐cultured with astrocytes. This effect, which is purinergic receptor‐dependent, was not observed in pure neuronal cultures. Thus, Aβ oligomer‐ and mGluR5‐dependent ATP release by astrocytes may contribute to the overall deleterious effect of mGluR5s in Alzheimer's disease. GLIA 2013;61:1673–1686     

Microglial Activation Results in Inhibition of TGF-β-Regulated Gene Expression

Chronic inflammation mediated by persistent microglial activation is associated with the pathogenesis of neurodegenerative diseases. The mechanisms underlying chronic microglial activation are poorly understood. We have previously shown that anti-inflammatory TGF-β signaling is inhibited in LPS-treated microglia. In this study, we assessed whether different disease-related microglial activators could downregulate TGF-β induction of gene expression. We examined the effects of amyloid β (Aβ) (1–42)- or heat-killed Listeria monocytogenes (HKLM) on the TGF-β-regulated gene expression in primary rat microglia. We found that Aβ (1–42) oligomers and HKLM, in addition to LPS, suppressed TGF-β-mediated induction of gene expression in part through reducing expression of TβR1 mRNA encoding the TGF-β receptor 1 in primary microglia. Aβ (1–42) and LPS also prevented induction of TGF-β-induced genes in primary microglia. Additionally, Aβ (1–42) rescued primary microglia from TGF-β-mediated cell death without increasing cell proliferation. Blockage of NFκB signaling, but not the ERK or IRF3 pathways, inhibited Aβ (1–42)- and LPS-mediated reduction of TβR1 mRNA. Finally, LPS and Aβ (1–42) induced transient upregulation of mRNAs encoding SnoN and Bambi, inhibitors of TGF-β signaling. Our data indicate that one mechanism through which activators may prolong microglial stimulation is through direct inhibition of anti-inflammatory signaling. A more detailed understanding of the interaction between inflammatory and anti-inflammatory pathways may reveal potential targets for ameliorating chronic inflammation and hence speed the development of therapeutics to address neurodegenerative diseases.     

Astrocytic Aβ1‐42 uptake is determined by Aβ‐aggregation state and the presence of amyloid‐associated proteins

Intracerebral accumulation of amyloid‐β (Aβ) leading to Aβ plaque formation, is the main hallmark of Alzheimer's disease and might be caused by defective Aβ‐clearance. We previously found primary human astrocytes and microglia able to bind and ingest Aβ1‐42 in vitro, which appeared to be limited by Aβ1‐42 fibril formation. We now confirm that astrocytic Aβ‐uptake depends on size and/or composition of Aβ‐aggregates as astrocytes preferably take up oligomeric Aβ over fibrillar Aβ. Upon exposure to either fluorescence‐labelled Aβ1‐42 oligomers (Aβ_oligo) or fibrils (Aβ_fib), a larger (3.7 times more) proportion of astrocytes ingested oligomers compared to fibrils, as determined by flow cytometry. Aβ‐internalization was verified using confocal microscopy and live‐cell imaging. Neither uptake of Aβ_oligo nor Aβ_fib, triggered proinflammatory activation of the astrocytes, as judged by quantification of interleukin‐6 and monocyte‐chemoattractant protein‐1 release. Amyloid‐associated proteins, including α1‐antichymotrypsin (ACT), serum amyloid P component (SAP), C1q and apolipoproteins E (ApoE) and J (ApoJ) were earlier found to influence Aβ‐aggregation. Here, astrocytic uptake of Aβ_fib increased when added to the cells in combination with SAP and C1q (SAP/C1q), but was unchanged in the presence of ApoE, ApoJ and ACT. Interestingly, ApoJ and ApoE dramatically reduced the number of Aβ_oligo‐positive astrocytes, whereas SAP/C1q slightly reduced Aβ_oligo uptake. Thus, amyloid‐associated proteins, especially ApoJ and ApoE, can alter Aβ‐uptake in vitro and hence may influence Aβ clearance and plaque formation in vivo. © 2010 Wiley‐Liss, Inc.     

Septohippocampal GABAergic neurons mediate the altered behaviors induced by n‐methyl‐D‐aspartate receptor antagonists

We hypothesize that selective lesion of the septohippocampal GABAergic neurons suppresses the altered behaviors induced by an N‐methyl‐D ‐aspartate (NMDA) receptor antagonist, ketamine or MK‐801. In addition, we hypothesize that septohippocampal GABAergic neurons generate an atropine‐resistant theta rhythm that coexists with an atropine‐sensitive theta rhythm in the hippocampus. Infusion of orexin‐saporin (ore‐SAP) into the medial septal area decreased parvalbumin‐immunoreactive (GABAergic) neurons by ～80%, without significantly affecting choline‐acetyltransferase‐immunoreactive (cholinergic) neurons. The theta rhythm during walking, or the immobility‐associated theta induced by pilocarpine, was not different between ore‐SAP and sham‐lesion rats. Walking theta was, however, more disrupted by atropine sulfate in ore‐SAP than in sham‐lesion rats. MK‐801 (0.5 mg/kg i.p.) induced hyperlocomotion associated with an increase in frequency, but not power, of the hippocampal theta in both ore‐SAP and sham‐lesion rats. However, MK‐801 induced an increase in 71–100 Hz gamma waves in sham‐lesion but not ore‐SAP lesion rats. In sham‐lesion rats, MK‐801 induced an increase in locomotion and an impairment of prepulse inhibition (PPI), and ketamine (3 mg/kg s.c.) induced a loss of gating of hippocampal auditory evoked potentials. MK‐801‐induced behavioral hyperlocomotion and PPI impairment, and ketamine‐induced auditory gating deficit were reduced in ore‐SAP rats as compared to sham‐lesion rats. During baseline without drugs, locomotion and auditory gating were not different between ore‐SAP and sham‐lesion rats, and PPI was slightly but significantly increased in ore‐SAP as compared with sham lesion rats. It is concluded that septohippocampal GABAergic neurons are important for the expression of hyperactive and psychotic symptoms an enhanced hippocampal gamma activity induced by ketamine and MK‐801, and for generating an atropine‐resistant theta. Selective suppression of septohippocampal GABAergic activity is suggested to be an effective treatment of some symptoms of schizophrenia. © 2012 Wiley Periodicals, Inc.     

Desacyl‐ghrelin and synthetic GH‐secretagogues modulate the production of inflammatory cytokines in mouse microglia cells stimulated by β‐amyloid fibrils

Data from Alzheimer's disease (AD) patients and AD animal models demonstrate the accumulation of inflammatory microglia at sites of insoluble fibrillar β‐amyloid protein (fAβ) deposition. It is known that fAβ binds to CD36, a type B scavenger receptor also involved in internalization of oxidized low‐density lipoprotein (LDL), and initiate a signaling cascade that regulates microglial recruitment, activation, and secretion of inflammatory mediators leading to neuronal dysfunction and death. The recent demonstration of a binding site for the growth hormone secretagogues (GHS) on CD36 prompted us to ascertain whether ghrelin and synthetic GHS could modulate the synthesis of inflammatory cytokines in fAβ‐activated microglia cells. We demonstrate that N9 microglia cells express the CD36 and are a suitable model to study the activation of inflammatory cytokines synthesis. In fact, in N9 cells exposed to fAβ_25–35 for 24 hr, the expression of interleukin (IL)‐1β and IL‐6 mRNA significantly increased. Interestingly, 10^?7 M desacyl‐ghrelin, hexarelin, and EP80317 in the nanomolar range effectively counteracted fAβ_25–35 stimulation of IL‐6 mRNA levels, whereas ghrelin was ineffective. Similarly, the effects of fAβ_25–35 on IL‐1β mRNA levels were attenuated by desacyl‐ghrelin, hexarelin, and EP80317, but not ghrelin. Because we have observed that the specific GHS receptor GHS‐R1a is not expressed in N9 cells, the actions of GHS should be mediated by different receptors. Reportedly, hexarelin and EP80317 are capable of binding the CD36 in mouse macrophages and reducing atherosclerotic plaque deposition in mice. We conclude that desacyl‐ghrelin, hexarelin, and EP80317 might interfere with fAβ activation of CD36 in microglia cells. © 2009 Wiley‐Liss, Inc.     

In vivo [123I]CNS‐1261 binding to D‐serine‐activated and MK801‐blocked NMDA receptors: A storage phosphor imaging study in rats

Disturbances of activity of the glutamatergic neurotransmitter system in the brain are present in many neuropsychiatric disorders. The N‐methyl‐D ‐aspartate (NMDA) receptor is the most abundant receptor of the glutamatergic system. In the neurodegenerative events of Alzheimer's disease, excessive activation of NMDA receptors may contribute to neuronal death. Inhibition of NMDA receptor activation may have neuroprotective effects and (semi)quantitative imaging of the activated system may help in the selection of patients for such inhibition therapies. In this study we evaluated [¹²³I]CNS‐1261 binding in the rat brain. This radiotracer binds in vivo to the MK801 binding site of activated NMDA receptors. To determine the optimal time point for ex vivo assessments after bolus injection [¹²³I]CNS‐1261 binding in rats, we performed a time course biodistribution study using dissection techniques. [¹²³I]CNS‐1261 binding was also studied in the rat brain using autoradiography by means of storage phosphor imaging, with prior facilitation of NMDA receptor activation by injection of the potent coagonist D ‐serine and after blocking of the NMDA receptor binding site by MK801 injection in D ‐serine pretreated rats. Measurements of [¹²³I]CNS‐1261 uptake matched the distribution of similar tracers for the MK801 binding site of the NMDA receptor and revealed an optimal time point of 2 h post injection for the assessment of tracer distribution in the rat brain. The blocking experiments indicated specific binding of [¹²³I]CNS‐1261 to NMDA receptors but also a considerable amount of nonspecific binding. Facilitation of NMDA receptor activation by D ‐serine did not result in an enhancement of binding of the radiotracer in the NMDA receptor‐rich rat hippocampus compared to the untreated group, as measured by autoradiography. In conclusion, our study has shown that [¹²³I]CNS‐1261 binding is influenced by NMDA receptor availability. However, high nonspecific binding limits quantification and small changes in receptor availability are unlikely to be detected. Synapse 63:557–564, 2009. © 2009 Wiley‐Liss, Inc.     

Apolipoproteins E and J interfere with amyloid‐beta uptake by primary human astrocytes and microglia in vitro

Defective clearance of the amyloid‐β peptide (Aβ) from the brain is considered a strong promoter in Alzheimer's disease (AD) pathogenesis. Astrocytes and microglia are important mediators of Aβ clearance and Aβ aggregation state and the presence of amyloid associated proteins (AAPs), such as Apolipoproteins E and J (ApoE and ApoJ), may influence Aβ clearance by these cells. Here we set out to investigate whether astrocytes and microglia differ in uptake efficiency of Aβ oligomers (Aβ_oligo) and Aβ fibrils (Aβ_fib), and whether the Aβ aggregation state and/or presence of AAPs affect Aβ uptake in these cells in vitro. Adult human primary microglia and astrocytes, isolated from short delay post‐mortem brain tissue, were exposed to either Aβ_oligo or Aβ_fib alone or combined with a panel of certain AAPs whereafter Aβ‐positive cells were quantified using flow cytometry. Upon exposure to Aβ combined with ApoE, ApoJ, α1‐antichymotrypsin (ACT) and a combination of serum amyloid P and complement C1q (SAP‐C1q), a clear reduction in astrocytic but not microglial Aβ_oligo uptake, was observed. In contrast, Aβ_fib uptake was strongly reduced in the presence of AAPs in microglia, but not in astrocytes. These data provide the first evidence of distinct roles of microglia and astrocytes in Aβ clearance. More importantly we show that Aβ clearance by glial cells is negatively affected by AAPs like ApoE and ApoJ. Thus, targeting the association of Aβ with AAPs, such as ApoE and ApoJ, could serve as a therapeutic strategy to increase Aβ clearance by glial cells. GLIA 2014;62:493–503     

Methyl 3,4‐dihydroxybenzoate protects primary cortical neurons against Aβ25–35‐induced neurotoxicity through mitochondria pathway

Amyloid‐β peptides (Aβ), which can aggregate into oligomers or fibrils in neurons, play a critical role in the pathogenesis of Alzheimer's disease (AD). Methyl 3,4‐dihydroxybenzoate (MDHB), a phenolic acid compound, has been reported to have antioxidative and neurotrophic effects. The present study investigated the neuroprotective effects of MDHB against Aβ‐induced apoptosis in rat primary cortical neutons. The primary cortical neurons were pretreated with different concentrations of MDHB for 24 hr, then incubated with 10 μM Aβ_25–35 for 24 hr. The results showed that Aβ_25–35 could induce neurotoxicity as evidenced by the decreased cell viability and the increased apoptotic rate. In parallel, Aβ_25–35 significantly increased the reactive oxygen species accumulation and decreased mitochondrial membrane potential. However, pretreatment of the primary cortical neurons with MDHB could effectively suppress these cellular events caused by Aβ_25–35 exposure. In addition, MDHB could increase the level of Bcl‐2, decrease the level of Bax, and inhibit the activation of caspase‐9 and caspase‐3 in Aβ_25–35‐treated primary cortical neurons. All these results were beneficial in their protective effect against Aβ‐induced neurotoxicity. Our results suggest that MDHB has a neuroprotective effect that provides a pharmacological basis for its clinical use in the treatment of AD. © 2013 Wiley Periodicals, Inc.     

Tripchlorolide protects neuronal cells from microglia‐mediated β‐amyloid neurotoxicity through inhibiting NF‐κB and JNK signaling

Recent research has focused on soluble oligomeric assemblies of β‐amyloid peptides (Aβ) as the proximate cause of neuroinflammation, synaptic loss, and the eventual dementia associated with Alzheimer's disease (AD). In this study, tripchlorolide (T₄), an extract of Tripterygium wilfordii Hook. F (TWHF), was studied as a novel agent to suppress neuroinflammatory process in microglial cells and to protect neuronal cells against microglia‐mediated oligomeric Aβ toxicity. T₄ significantly attenuated oligomeric Aβ(1‐42)‐induced release of inflammatory productions such as tumor necrosis factor‐α, interleukin‐1β, nitric oxide (NO), and prostaglandin E₂. It also downregulated the protein levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2) in microglial cells. Further molecular mechanism study demonstrated that T₄ inhibited the nuclear translocation of nuclear factor‐κB (NF‐κB) without affecting I‐κBα phosphorylation. It repressed Aβ‐induced JNK phosphorylation but not ERK or p38 MAPK. The inhibition of NF‐κB and JNK by T₄ is correlated with the suppression of inflammatory mediators in Aβ‐stimulated microglial cells. These results suggest that T₄ protects neuronal cells by blocking inflammatory responses of microglial cells to oligomeric Aβ(1‐42) and that T₄ acts on the signaling of NF‐κB and JNK, which are involved in the modulation of inflammatory response. Therefore, T₄ may be an effective agent in modulating neuroinflammatory process in AD. © 2009 Wiley‐Liss, Inc.     

Mechanisms of status epilepticus: α‐Amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid receptor hypothesis

Prolonged seizures of status epilepticus (SE) result from failure of mechanisms of seizure termination or activation of mechanisms that sustain seizures. Reduced γ‐aminobutyric acid type A receptor–mediated synaptic transmission contributes to impairment of seizure termination. However, mechanisms that sustain prolonged seizures are not known. We propose that insertion of GluA1 subunits at the glutamatergic synapses causes potentiation of α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic receptor (AMPAR)‐mediated neurotransmission, which helps to spread and sustain seizures. The AMPAR‐mediated neurotransmission of CA1 pyramidal neurons was increased in animals in SE induced by pilocarpine. The surface membrane expression of GluA1 subunit–containing AMPARs on CA1 pyramidal neurons was also increased. Blockade of N‐methyl‐d ‐aspartate receptors 10 minutes after the onset of continuous electrographic seizure activity prevented the increase in the surface expression of GluA1 subunits. N‐methyl‐d ‐aspartate receptor antagonist MK‐801 in conjunction with diazepam also terminated seizures that were refractory to MK‐801 or diazepam alone. Future studies using mice lacking the GluA1 subunit expression will provide further insights into the role of GluA1 subunit–containing AMPAR plasticity in sustaining seizures of SE.     

Cannabinoids prevent the amyloid β‐induced activation of astroglial hemichannels: A neuroprotective mechanism

The mechanisms involved in Alzheimer's disease are not completely understood and how astrocytes and their gliotransmission contribute to this neurodegenerative disease remains to be fully elucidated. Previous studies have shown that amyloid‐β peptide (Aβ) induces neuronal death by a mechanism that involves the excitotoxic release of ATP and glutamate associated to astroglial hemichannel opening. We have demonstrated that synthetic and endogenous cannabinoids (CBs) reduce the opening of astrocyte Cx43 hemichannels evoked by activated microglia or inflammatory mediators. Nevertheless, whether CBs could prevent the astroglial hemichannel‐dependent death of neurons evoked by Aβ is unknown. Astrocytes as well as acute hippocampal slices were treated with the active fragment of Aβ alone or in combination with the following CBs: WIN, 2‐AG, or methanandamide (Meth). Hemichannel activity was monitored by single channel recordings and by time‐lapse ethidium uptake while neuronal death was assessed by Fluoro‐Jade C staining. We report that CBs fully prevented the hemichannel activity and inflammatory profile evoked by Aβ in astrocytes. Moreover, CBs fully abolished the Aβ‐induced release of excitotoxic glutamate and ATP associated to astrocyte Cx43 hemichannel activity, as well as neuronal damage in hippocampal slices exposed to Aβ. Consequently, this work opens novel avenues for alternative treatments that target astrocytes to maintain neuronal function and survival during AD. GLIA 2016 GLIA 2017;65:122–137     

Tyrosine phosphatase STEP61 negatively regulates amyloid β‐mediated ERK/CREB signaling pathways via α7 nicotinic acetylcholine receptors

Striatal‐enriched phosphatase 61 (STEP₆₁) plays an essential role in synaptic plasticity and has recently been implicated in neurodegenerative disease. Here we characterized a possible role of STEP₆₁ in Alzheimer's disease (AD) pathology using a mouse model of AD (Tg‐APPswe/PSEN1dE9, APP/PS1 mice) and an in vitro model of AD [cortical neurons treated with amyloid β (Aβ)_1–42 peptides]. Our data indicate age‐related elevation of STEP₆₁ levels and the proportion of dephosphorylated STEP₆₁ (active STEP₆₁) in wild‐type mice, which was enhanced in APP/PS1 mice. Furthermore, the increased STEP₆₁ levels and active STEP₆₁ were observed in the hippocampus and cortex from 12‐month‐old APP/PS1 mice and in Aβ_1–42‐treated cortical neurons. An α7 nicotinic acetylcholine receptors (nAChRs) antagonist, α‐bungarotoxin (BTX), inhibited the Aβ_1–42‐induced increase of STEP₆₁ expression and activation. In addition, extracellular signal‐regulated kinase 1/2 (ERK1/2) and cAMP response element binding (CREB) were impaired in Aβ_1–42‐treated cortical neurons, and knockdown of STEP₆₁ enhanced the activation of ERK1/2 and CREB. Collectively, these findings indicate two alternate pathological pathways effecting STEP₆₁ regulation in AD. First, Aβ regulating STEP₆₁ activity is mediated by Aβ binding to α7 nAChRs. Second, STEP₆₁ negatively regulates Aβ‐mediated ERK/CREB pathway, an important signaling cascade involved in memory formation. © 2013 Wiley Periodicals, Inc.     

Neonatal exposure to MK‐801 reduces mRNA expression of mGlu3 receptors in the medial prefrontal cortex of adolescent rats

Schizophrenia is considered as a “neurodegenerative” and “neurodevelopmental” disorder, the pathophysiology of which may include hypofunction of the N‐methyl‐d ‐aspartate receptor (NMDA‐R) or subsequent pathways. Accordingly, administration of NMDA‐R antagonists to rodents during the perinatal period may emulate some core pathophysiological aspects of schizophrenia. The effect of 4‐day (postnatal day; PD 7–10) administration of MK‐801, a selective NMDA‐R antagonist, on gene expression in the medial prefrontal cortex (mPFC), hippocampus, and amygdala was evaluated using quantitative polymerase chain reaction methods. Specifically, we sought to determine whether genes related to Glu transmissions, for example those encoding for NMDA‐Rs, metabotropic Glu receptors (mGluRs), or Glu transporters, were altered by neonatal treatment with MK‐801. Model rats showed downregulation of the mGluR3 subtype in the mPFC around puberty, especially at PD 35 in response to MK‐801 or during ontogenesis without pharmacological manipulations. Genes encoding for other mGluRs subtypes, that is NMDA‐Rs and Glu transporters, were not affected by the neonatal insult. These results suggest that NMDA‐R antagonism in the early course of development modulates the expression of mGluR3 in mPFC around puberty. Thus, mGluR3 may serve as a potential target to prevent the onset and progression of schizophrenia. Synapse 68:202–208, 2014 . © 2014 Wiley Periodicals, Inc.