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Organic light-emitting diodes (OLEDs) based on thermally activated delayed fluorescence (TADF) materials are promising for the realization of highly efficient light emitters. However, such devices have so far suffered from efficiency roll-off at high luminance. Here, we report the design and synthesis of two diboron-based molecules, CzDBA and tBuCzDBA, which show excellent TADF properties and yield efficient OLEDs with very low efficiency roll-off. These donor–acceptor–donor (D–A–D) type and rod-like compounds concurrently generate TADF with a photoluminescence quantum yield of ~100% and an 84% horizontal dipole ratio in the thin film. A green OLED based on CzDBA exhibits a high external quantum efficiency of 37.8 ± 0.6%, a current efficiency of 139.6 ± 2.8 cd A−1 and a power efficiency of 121.6 ± 3.1 lm W−1 with an efficiency roll-off of only 0.3% at 1,000 cd m−2. The device has a peak emission wavelength of 528 nm and colour coordinates of the Commission International de l´Eclairage (CIE) of (0.31, 0.61), making it attractive for colour-display applications.

New design of Pt(II) phosphors yield near-infrared organic light-emitting diodes with high efficiency and brightness.

Correction for ‘Superior upconversion fluorescence dopants for highly efficient deep-blue electroluminescent devices’ by Yi-Hsiang Chen et al., Chem. Sci., 2016, DOI: 10.1039/c6sc00100a.

The Sn whisker growth in Cu(top)-Sn(bottom) bilayer system upon room temperature aging was investigated by scanning electron microscope and X-ray diffraction techniques. The experimental observations indicate that the Sn whisker growth on the Cu surface in Cu-Sn bilayer system is different from that on the Sn surface in Sn-Cu bilayer system. When the Sn sublayer thickness is less than 0.5μm, the Sn whisker growth can take place in Cu-Sn system but not in Sn-Cu system. An explanation for Sn whisker growth in Cu-Sn bilayer system is given.

Rhinosinusitis is one of the most common chronic inflammatory diseases worldwide. It has been associated with many diseases, including allergic and non-allergic rhinitis, asthma, nasal polyps, aspirin sensitivity, cystic fibrosis, immotile cilia syndrome, immunodeficiency, airway infection, otitis media and dental diseases. Due to the complexity in clinical manifestation and classification of rhinosinusitis, and their associated etiology and pathophysiology, evidence based consensus on diagnosis and treatment is important in order to enhance the effectiveness of available treatment and the quality of life of the patients. Since patients with rhinosinusitis may consult a wide range of clinicians, including otolaryngologists, allergists, pulmonologists and paediatricians, clinical guidelines or consensus with the evidence based diagnostic and treatment scheme for rhinosinusitis for different specialists are very important. The first European Position Paper on Rhinosinusitis and Nasal Polyps (EP3OS) was published in 2005 by the European Academy of Allergology and Clinical Immunology in the Journal of “Rhinology”, and the updated versions were published in 2007 and 2012 in the same journal. EP3OS 2007 had been endorsed by many international medical societies, including WHO (e.g., InterAirway document), and the full document was translated into more than 15 languages, and its contents have been widely endorsed, as some chapters were significantly enriched, as extensive up-to-date information on epidemiology, predisposing factors, pediatric aspects and randomized double-blinded placebo-controlled studies in the treatment of rhinosinusitis were added, and it encourages further clinical and basic research in this field in the future.

Treatment with cytosine beta-D-arabinoside (AraC; 300 microM) induced a time-dependent accumulation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) protein in nuclei purified from cultured cerebellar granule cells, with a concomitant degradation of lamin B1, a nuclear membrane protein and a substrate of CPP32/caspase-3. Moreover, Asp-Glu-Val-Asp-fluoromethyl ketone (DEVD-fmk), a CPP32-selective antagonist, dose-dependently suppressed AraC-induced apoptosis of these neurons. Nuclear accumulation of GAPDH protein was associated with a progressive decrease in the activity of uracil-DNA glycosylase (UDG), one of the nuclear functions of GAPDH. The nuclear dehydrogenase activity of GAPDH was initially increased after treatment and then decreased parallel to UDG activity. Six GAPDH isoforms were detected in the nuclei of AraC-treated cells. The more alkaline isoforms, 1-3, constituted the bulk of the nuclear GAPDH, and the remaining isoforms, 4-6, were the minor species. Levels of all six isoforms were increased after treatment with AraC for 16 h; a 4-h treatment increased levels of only isoforms 4 and 5. Thus, it appears that various GAPDH isoforms are differentially regulated and may have distinct apoptotic roles. Pretreatment with GAPDH antisense oligonucleotide blocked the nuclear translocation of GAPDH isoforms, and the latter process occurred concurrently with a decrease in cytosolic GAPDH isoforms. Sodium nitroprusside-induced NAD labeling of nuclear GAPDH showed a 60% loss of GAPDH labeling after AraC treatment, suggesting that the active site of GAPDH may be covalently modified, denatured, or improperly folded. The unfolded protein response elicited by denatured GAPDH may contribute to AraC-induced neuronal death.

In an earlier study, we demonstrated that PAN-811 (3-aminopyridine-2-carboxaldehyde thiosemicarbazone), a novel neuroprotectant, provides protection against glutamate, staurosporine, veratridine, or hypoxia/hypoglycemia toxicities in primary cortical neuronal cultures by upregulating Bcl-2 expression [R.-W. Chen, C. Yao, X.C. Lu, Z.-G. Jiang, R. Whipple, Z. Liao, H.A. Ghanbari, B. Almassian, F.C. Tortella, J.R. Dave. PAN-811 (3-aminopyridine-2-carboxaldehyde thiosemicarbazone), a novel neuroprotectant, elicits its function in primary neuronal cultures by upregulating Bcl-2 expression. Neuroscience 135 (2005) 191–201]. Both JNK (c-Jun N-terminal kinase) and p38 MAP (mitogen-activated protein) kinase activation have a direct inhibitory action on Bcl-2 by phosphorylation. In the present study, we continued to explore the mechanism of PAN-811 neuroprotection. Our results indicate that treatment of cultured cortical neurons with glutamate (100 μM) induces phosphorylation of both JNK and p38 MAPK. Specifically, pretreatment of neurons with 10 μM PAN-811 (an optimal neuroprotective concentration) for 1 h, 4 h, or 24 h significantly suppresses glutamate-mediated activation of both JNK and p38 MAPK. Furthermore, the p38 MAPK-specific inhibitor SB203580 and the JNK-specific inhibitor SP600125 prevented glutamate-induced neuronal death in these primary cultures. Our results demonstrate that glutamate-induced phosphorylation of JNK and p38 MAPK is suppressed by PAN-811, which might contribute to Bcl-2 upregulation and PAN-811 neuroprotection.

Cell cycle reentry has been found during apoptosis of postmitotic neurons under certain pathological conditions. To evaluate whether nuclear factor-kappaB (NF-kappaB) activation promotes cell cycle entry and neuronal apoptosis, we studied the relation among NF-kappaB-mediated cyclin induction, bromodeoxyuridine (BrdU) incorporation, and apoptosis initiation in rat striatal neurons following excitotoxic insult. Intrastriatally injected N-methyl-D-aspartate receptor agonist quinolinic acid (QA, 60 nmol) elicited a rise in cyclin D1 mRNA and protein levels (P

Acute seizures following brain injury have been associated with a worsening of patient outcome, but they are often undiagnosed and untreated when they occur without motor convulsions. Here, we sought to compare the antiseizure profile of ethosuximide (EXM; 125-312.5 mg/kg i.v.) and gabapentin (GBP; 0.3-50 mg/kg. i.v.) in a rat model of nonconvulsive seizures (NCS) induced by brain ischemia. Seizures were detected by continuous electroencephalographic monitoring for 24 h following permanent middle cerebral artery occlusion (MCAo). Both "preseizure" and "postseizure" treatment effects were evaluated. Control rats experienced a 91% incidence of NCS (averaging 10-11 NCS/rat), which was significantly reduced following preseizure treatment (delivered 20 min post-MCAo) with either EXM (ED(50) = 161 mg/kg) or GBP (ED(50) = 10.5 mg/kg). In contrast to preseizure treatment effects, only GBP reduced NCS when given after the first seizure event. A further, albeit nonsignificant, 20% reduction in NCS incidence was measured when given in combination postseizure. Drug treatment also reduced infarct volume, which was positively correlated to the number of NCS events (r = 0.475; P0.001). EXM and GBP treatment of cultured neurons exposed to neurotoxic or ischemic insults showed no neuroprotective effects, suggesting that in vivo neuroprotection can be attributed to anti-seizure effects. We conclude that EXM and GBP significantly attenuate NCS in a dose-related manner and may help to improve patient outcome from brain ischemia-induced seizure activity.

Cellular and molecular pathways underlying ischemic neurotoxicity are multifaceted and complex. Although many potentially neuroprotective agents have been investigated, the simplicity of their protective mechanisms has often resulted in insufficient clinical utility. We describe a previously uncharacterized class of potent neuroprotective compounds, represented by PAN-811, that effectively block both ischemic and hypoxic neurotoxicity. PAN-811 disrupts neurotoxic pathways by at least two modes of action. It causes a reduction of intracellular-free calcium as well as free radical scavenging resulting in a significant decrease in necrotic and apoptotic cell death. In a rat model of ischemic stroke, administration of PAN-811 i.c.v. 1 h after middle cerebral artery occlusion resulted in a 59% reduction in the volume of infarction. Human trials of PAN-811 for an unrelated indication have established a favorable safety and pharmacodynamic profile within the dose range required for neuroprotection warranting its clinical trial as a neuroprotective drug.

Neurotoxicity in primary neurons was induced using hypoxia/hypoglycemia (H/H), veratridine (10microM), staurosporine (1microM) or glutamate (100microM), which resulted in 72%, 67%, 75% and 66% neuronal injury, respectively. 3-Aminopyridine-2-carboxaldehyde thiosemicarbazone (PAN-811; 10microM; Panacea Pharmaceuticals, Gaithersburg, MD) pretreatment for 24 h provided maximal neuroprotection of 89%, 42%, 47% and 89% against these toxicities, respectively. Glutamate or H/H treatment of cells increased cytosolic cytochrome c levels, which was blocked by pretreatment of cells with PAN-811. Pretreatment of neurons with PAN-811 produced a time-dependent increase in the protein level of Bcl-2, which was evident even after glutamate or H/H treatments. An up-regulation in the expression of the p53 and Bax genes was also observed following exposure to these neurotoxic insults; however, this increase was not suppressed by PAN-811 pretreatment. Functional inhibition of Bcl-2 by HA14-1 reduced the neuroprotective efficacy of PAN-811. PAN-811 treatment also abolished glutamate or H/H-mediated internucleosomal DNA fragmentation.

Cyclooxygenase-2 (COX-2) activity has been implicated in the pathogenesis of neuronal cell death in ischemia and other diseases, but the mechanism by which COX-2 exacerbates cell death is unknown. COX-2 activity is known to induce expression of cyclin D1 in neoplastic cells, and cyclin D1 expression can induce cell death in postmitotic neurons. In the present study, the role of COX-2 and cyclin D1 in neuronal cell death induced by anoxia and ischemia was examined. Treatment with the COX-2 specific inhibitor (NS 398 25 microM) and cyclin D1 inhibitor (flavopiridol 1 microM) increased neuronal survival and inhibited DNA fragmentation after anoxia. NS-398 suppressed anoxia-induced expression of cyclin D1. Flavopiridol inhibited the anoxia-induced increased expression of cyclin D1, but had no effect on COX-2 expression. Treatment with the selective COX-2 inhibitor, SC58125, had no affect on COX-2 expression but partially suppressed cyclin D1 expression in the cortex following middle cerebral artery occlusion in vivo. These results show that COX-2 activity is required for cyclin D1 expression after ischemia in vivo and anoxia in vitro. These data provide support for the hypothesis that cyclin D1 expression is an important mechanism by which COX-2 activity exacerbates ischemic neuronal death.

Highly efficient deep-blue electroluminescent devices are realized via an efficient triplet–triplet annihilation process of styrylpyrene-based emitters., In this study, we revealed a new approach for the development of new triplet–triplet annihilation (TTA) materials with highly efficient deep-blue fluorescence via the incorporation of a styrylpyrene core and an electron-donating group. The resulting deep-blue emitters (PCzSP, DFASP, and DPASP) exhibit intramolecular charge transfer emissions with remarkably high emission quantum yields. The electroluminescent devices based on these three fluorophores as dopants using CBP as a host exhibit very high device efficiencies; in particular, the DPASP-doped device reveals an extremely high EQE of 12%, reaching the limit of a TTA-based device. The EL characteristics of DPASP-doped CBP-based devices at various doping concentrations (0–5%) suggest that the dopant DPASP is responsible for the TTA-type delayed fluorescence in the device; no delayed fluorescence was observed for the device using CBP as the host emitter. Moreover, when using DMPPP with ambipolar characteristics as the host, the deep-blue DPASP-doped device also gives outstanding performance with an EQE of nearly 11% with an extremely small efficiency roll-off, which was ascribed to the excellent charge balance in the emitting layer of the EL device. The TTA process of the SP-based dopants accounts significantly for the superior efficiencies of the EL devices.

Lithium has long been a primary drug used to treat bipolar mood disorder, even though the drug's therapeutic mechanisms remain obscure. Recent studies demonstrate that lithium has neuroprotective effects against glutamate-induced excitotoxicity in cultured neurons and in vivo . The present study was undertaken to examine whether postinsult treatment with lithium reduces brain damage induced by cerebral ischemia. We found that s.c. injection of lithium dose dependently (0.5–3 mEq/kg) reduced infarct volume in the rat model of middle cerebral artery occlusion/reperfusion. Infarct volume was reduced at a therapeutic dose of 1 mEq/kg even when administered up to 3 h after the onset of ischemia. Neurological deficits induced by ischemia were also reduced by daily administration of lithium over 1 week. Moreover, lithium treatment decreased the number of neurons showing DNA damage in the ischemic brain. These neuroprotective effects were associated with an up-regulation of cytoprotective heat shock protein 70 (HSP70) in the ischemic brain hemisphere as determined by immunohistochemistry and Western blotting analysis. Lithium-induced HSP70 up-regulation in the ischemic hemisphere was preceded by an increase in the DNA binding activity of heat shock factor 1, which regulates the transcription of HSP70. Physical variables and cerebral blood flow were unchanged by lithium treatment. Our results suggest that postinsult lithium treatment reduces both ischemia-induced brain damage and associated neurological deficits. Moreover, the heat shock response is likely to be involved in lithium's neuroprotective actions. Additionally, our studies indicate that lithium may have clinical utility for the treatment of patients with acute stroke.

In rat cerebellar granule cells, glutamate induced rapid activation of c-Jun N-terminal kinase (JNK) and p38 kinase to phosphorylate c-Jun (at Ser63) and p53 (at Ser15), respectively, and a subsequent marked increase in activator protein-1 (AP-1) binding that preceded apoptotic death. These glutamate-induced effects and apoptosis could largely be prevented by long-term (7 days) pretreatment with 0.5–2 mm lithium, an antibipolar drug. Glutamate's actions could also be prevented by known blockers of this pathway, MK-801 (an NMDA receptor blocker), SB 203580 (a p38 kinase inhibitor) and curcumin (an AP-1 binding inhibitor). The concentration- and time-dependent suppression of glutamate's effects by lithium and curcumin correlated well with their neuroprotective effects. These results suggest a prominent role of JNK and p38, as well as their downstream AP-1 binding activation and p53 phosphorylation in mediating glutamate excitotoxicity. Moreover, the neuroprotective effects of lithium are mediated, at least in part, by suppressing NMDA receptor-mediated activation of the mitogen-activated protein kinase pathway.

Lithium, the major drug used to treat manic depressive illness, robustly protects cultured rat brain neurons from glutamate excitotoxicity mediated by N-methyl-D-aspartate (NMDA) receptors. The lithium neuroprotection against glutamate excitotoxiciy is long-lasting, requires long-term pretreatment and occurs at therapeutic concentrations of this drug. The neuroprotective mcchanisms involve inactivation of NMDA receptors, decreased expression of pro-apoptotic proteins, p53 and Bax, enhanced expression of the cytoprotective protein, Bcl-2, and activation of the cell survival kinase, Akt. In addition, lithium pretreatment suppresses glutamate-induced loss of the activities of Akt, cyclic AMP-response element binding protein (CREB), c-Jun - N-terminal kinase (JNK) and p38 kinase. Lithium also reduces brain damage in animal models of neurodegenerative diseases in which excitotoxicity has been implicated. In the rat model of stroke using middle cerebral artery occlusion, lithium markedly reduces neurologic deficits and decreases brain infarct volume even when administered after the onset of ischemia. In a rat Huntington's disease model, lithium significantly reduces brain lesions resulting from intrastriatal infusion of quinolinic acid, an excitotoxin. Our results suggest that lithium might have utility in the treatment of neurodegenerative disorders in addition to its common use for the treatment of bipolar depressive patients.

This study was undertaken to investigate the molecular mechanisms underlying the neuroprotective actions of lithium against glutamate excitotoxicity with a focus on the role of proapoptotic and antiapoptotic genes. Long term, but not acute, treatment of cultured cerebellar granule cells with LiCl induces a concentration-dependent decrease in mRNA and protein levels of proapoptotic p53 and Bax; conversely, mRNA and protein levels of cytoprotective Bcl-2 are remarkably increased. The ratios of Bcl-2/Bax protein levels increase by approximately 5-fold after lithium treatment for 5–7 days. Exposure of cerebellar granule cells to glutamate induces a rapid increase in p53 and Bax mRNA and protein levels with no apparent effect on Bcl-2 expression. Pretreatment with LiCl for 7 days prevents glutamate-induced increase in p53 and Bax expression and maintains Bcl-2 in an elevated state. Glutamate exposure also triggers the release of cytochrome c from the mitochondria into the cytosol. Lithium pretreatment blocks glutamate-induced cytochrome c release and cleavage of lamin B1, a nuclear substrate for caspase-3. These results strongly suggest that lithium-induced Bcl-2 up-regulation and p53 and Bax down-regulation play a prominent role in neuroprotection against excitotoxicity. Our results further suggest that lithium, in addition to its use in the treatment of bipolar depressive illness, may have an expanded use in the intervention of neurodegeneration.

Glycine-proline-glutamate (GPE) is an N-terminal tripeptide endogenously cleaved from insulin-like growth factor-1 in the brain and is neuroprotective against hypoxic-ischemic brain injury and neurodegeneration. NNZ-2566 is an analog of GPE designed to have improved bioavailability. In this study, we tested NNZ-2566 in a rat model of penetrating ballistic-type brain injury (PBBI) and assessed its effects on injury-induced histopathology, behavioral deficits, and molecular and cellular events associated with inflammation and apoptosis. In the initial dose-response experiments, NNZ-2566 (0.01-3 mg/kg/h x 12 h intravenous infusion) was given at 30 min post-injury and the therapeutic time window was established by delaying treatments 2-4 h post-injury, but with the addition of a 10- or 30-mg/kg bolus dose. All animals survived 72 h. Neuroprotection was evaluated by balance beam testing and histopathology. The effects of NNZ-2566 on injury-induced changes in Bax and Bcl-2 proteins, activated microgliosis, neutrophil infiltration, and astrocyte reactivity were also examined. Behavioral results demonstrated that NNZ-2566 dose-dependently reduced foot faults by 19-66% after acute treatments, and 35-55% after delayed treatments. Although gross lesion volume was not affected, NNZ-2566 treatment significantly attenuated neutrophil infiltration and reduced the number of activated microglial cells in the peri-lesion regions of the PBBI. PBBI induced a significant upregulation in Bax expression (36%) and a concomitant downregulation in Bcl-2 expression (33%), both of which were significantly reversed by NNZ-2566. Collectively, these results demonstrated that NNZ-2566 treatment promoted functional recovery following PBBI, an effect related to the modulation of injury-induced neural inflammatory and apoptotic mechanisms.

Recent efforts have been aimed at developing a panel of protein biomarkers for the diagnosis/prognosis of the neurological damage associated with acute brain injury.This study utilized high-throughput immunoblotting (HTPI) technology to compare changes between two animal models of acute brain injury: penetrating ballistic-like brain injury (PBBI) which mimics the injury created by a gunshot wound and transient middle cerebral artery occlusion (MCAo) which is a model of stroke. Brain and blood were collected at 24-hours post-injury.This study identified the changes in 18 proteins following PBBI and 17 proteins following MCAo out of a total of 998 screened proteins. Distinct differences were observed between the two models: five proteins were up- or down-regulated in both models, 23 proteins changed in only one model and one protein was differentially expressed. Western blots were used to verify HTPI results for selected proteins with measurable changes observed in both blood and brain for the proteins STAT3, Tau, PKA RII beta, 14-3-3 epsilon and p43/EMAPII.These results suggest distinct post-injury protein profiles between brain injury types (traumatic vs. ischemic) that will facilitate strategies aimed at the differential diagnosis and prognosis of acute brain injury.

The objective of this study was to determine levels of DNA fragmentation in blood leukocytes and parietal cortex from guinea pigs following repeated low-level exposure to the chemical warfare nerve agent (CWNA) sarin. Guinea pigs were injected (s.c.) once a day for 10 days with saline, or 0.1, 0.2, or 0.4 LD50 (50% mean lethal dose) sarin dissolved in sterile physiological saline. Blood and parietal cortex was collected after injection at 0, 3, and 17 days recovery and evaluated for DNA fragmentation using single-cell gel electrophoresis (Comet assay). Cells were imaged using comet analysis software and three parameters of DNA fragmentation measured: tail length, percent DNA in the tail, and tail moment arm. Repeated low-dose exposure to sarin produced a dose-dependent response in leukocytes at 0 and 3 days post-exposure. There was a significant increase in all measures of DNA fragmentation at 0.2 and 0.4 LD50, but not at 0.1 LD50. There was no significant increase in DNA fragmentation in any of the groups at 17 days post-exposure. Sarin did not produce a systematic dose-dependent response in parietal cortex at any of the time points. However, significant increases in DNA fragmentation at 0.1 and 0.4 LD50 were observed at 0 and 3 days post-exposure. All measures of DNA fragmentation in both leukocytes and neurons returned to control levels by 17 days post-exposure, indicating a small and non-persistent increase in DNA fragmentation following repeated low-level exposure to sarin.

Cellular injury can involve the aberrant stimulation of cell cycle proteins in part through activation of phosphodiesterases (PDEs) and downstream expression of cell-cycle components such as cyclin D1. In mature non-proliferating cells activation of the cell cycle can lead to the induction of programmed cell death. In the present study, we investigated the in vitro neuroprotective efficacy and mechanism of action of vinpocetine (PDE1 inhibitor), trequinsin (PDE3 inhibitor), and rolipram (PDE4 inhibitor) in four mechanistically-distinct models of injury to primary rat cortical neurons as related to cell cycle regulation and apoptosis. Cellular injury was induced by hypoxia/hypoglycemia, veratridine (10 microM), staurosporine (1 microM), or glutamate (100 microM), resulting in average neuronal cell death rates of 43-48% as determined by MTT assay. Treatment with each PDE inhibitor (PDEI) resulted in a similar concentration-dependent neuroprotection profile with maximal effective concentrations of 5-10 microM (55-77% neuroprotection) in all four neurotoxicity models. Direct cytotoxicity due to PDE inhibition alone was not observed at concentrations below 100 microM. Further studies indicated that PDEIs can suppress the excitotoxic upregulation of cyclin D1 similar to the effects of flavopiridol, a cyclin-dependent kinase inhibitor, including suppression of pro-apoptotic caspase-3 activity. Overall, these data indicate that PDEIs are broad-spectrum neuroprotective agents acting through modulation of cell cycle elements and may offer a novel mode of therapy against acute injury to the brain.

Valproic acid (VPA), used to treat bipolar mood disorder and seizures, also inhibits histone deacetylase (HDAC). Here, we found that VPA and other HDAC inhibitors, butyrate and trichostatin A, robustly protected mature cerebellar granule cell cultures from excitotoxicity induced by SYM 2081 ((2S, 4R)-4-methylglutamate), an inhibitor of excitatory amino-acid transporters and an agonist of low-affinity kainate receptors. These neuroprotective effects required protracted treatment and were correlated with enhanced acetylated histone levels, indicating HDAC inhibition. SYM-induced excitotoxicity was blocked by MK-801 ((5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate), supporting that the toxicity was largely N-methyl-D-aspartate receptor dependent. SYM excitotoxicity had apoptotic characteristics and was prevented by a caspase inhibitor. SYM-induced apoptosis was associated with a rapid and robust nuclear accumulation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a housekeeping gene previously shown to be proapoptotic. VPA pretreatment suppressed SYM 2081-induced GAPDH nuclear accumulation, concurrent with its neuroprotective effects. Chromatin immunoprecipitation (ChIP) revealed that GAPDH is copresent with acetylated histone H3, including Lys9-acetylated histone, and that VPA treatment caused a time-dependent decrease in the levels of nuclear GAPDH with a concomitant increase in acetylated histones in the ChIP complex. Our results strongly suggest that VPA protects neurons from excitotoxicity through inhibition of HDAC activity and that this protective effect may involve suppression of excitotoxicity-induced accumulation of GAPDH protein in the nucleus.

Lithium, the major drug used to treat manic depressive illness, robustly protects cultured rat brain neurons from glutamate excitotoxicity mediated by N-methyl-D-aspartate (NMDA) receptors. The lithium neuroprotection against glutamate excitotoxiciy is long-lasting, requires long-term pretreatment and occurs at therapeutic concentrations of this drug. The neuroprotective mcchanisms involve inactivation of NMDA receptors, decreased expression of pro-apoptotic proteins, p53 and Bax, enhanced expression of the cytoprotective protein, Bcl-2, and activation of the cell survival kinase, Akt. In addition, lithium pretreatment suppresses glutamate-induced loss of the activities of Akt, cyclic AMP-response element binding protein (CREB), c-Jun - N-terminal kinase (JNK) and p38 kinase. Lithium also reduces brain damage in animal models of neurodegenerative diseases in which excitotoxicity has been implicated. In the rat model of stroke using middle cerebral artery occlusion, lithium markedly reduces neurologic deficits and decreases brain infarct volume even when administered after the onset of ischemia. In a rat Huntington's disease model, lithium significantly reduces brain lesions resulting from intrastriatal infusion of quinolinic acid, an excitotoxin. Our results suggest that lithium might have utility in the treatment of neurodegenerative disorders in addition to its common use for the treatment of bipolar depressive patients.

We have studied neurotoxicity induced by pharmacological concentrations of 3-hydroxykynurenine (3-HK), an endogenous toxin implicated in certain neurodegenerative diseases, in cerebellar granule cells, PC12 pheochromocytoma cells, and GT1-7 hypothalamic neurosecretory cells. In all three cell types, the toxicity was induced in a dose-dependent manner by 3-HK at high micromolar concentrations and had features characteristic of apoptosis, including chromatin condensation and internucleosomal DNA cleavage. In cerebellar granule cells, the 3-HK neurotoxicity was unaffected by xanthine oxidase inhibitors but markedly potentiated by superoxide dismutase and its hemelike mimetic, MnTBAP [manganese(III) tetrakis(benzoic acid)porphyrin chloride]. Catalase blocked 3-HK neurotoxicity in the absence and presence of superoxide dismutase or MnTBAP. The formation of H2O2 was demonstrated in PC12 and GT1-7 cells treated with 3-HK, by measuring the increase in the fluorescent product, 2′,7′-dichlorofluorescein. In both PC12 and cerebellar granule cells, inhibitors of the neutral amino acid transporter that mediates the uptake of 3-HK failed to block 3-HK toxicity. However, their toxicity was slightly potentiated by the iron chelator, deferoxamine. Taken together, our results suggest that neurotoxicity induced by pharmacological concentrations of 3-HK in these cell types is mediated primarily by H2O2, which is formed most likely by auto-oxidation of 3-HK in extracellular compartments. 3-HK-induced death of PC12 and GT1-7 cells was protected by dantrolene, an inhibitor of calcium release from the endoplasmic reticulum. The protection by dantrolene was associated with a marked increase in the protein level of Bcl-2, a prominent antiapoptotic gene product. Moreover, overexpression of Bcl-2 in GT1-7 cells elicited by gene transfection suppressed 3-HK toxicity. Thus, dantrolene may elicit its neuroprotective effects by mechanisms involving up-regulation of the level and function of Bcl-2 protein.

Treatment of rat pheochromocytoma cells (PC 12) cells with beta-amyloid peptide-(1-42) for 24 h induced a concentration-dependent decrease in cellular redox activity in the dose range of 1 to 20 microM. These effects were markedly attenuated by pretreatment with 2 mM LiCl for 7 days, whereas 1-day pretreatment was ineffective. Measurements of live and dead cells by double-staining with fluorescein diacetate and propidium iodide, respectively revealed that protracted lithium pretreatment attenuated PC 12 cell death induced by beta-amyloid-(1-42) and cerebellar granule cell death induced by beta-amyloid-(25-35). Preceding PC 12 cell death, beta-amyloid peptide elicited a slight decrease in protein levels of Bcl-2. Conversely, 7-day pretreatment with lithium resulted in an approximate doubling of Bcl-2 protein levels in cells treated with or without beta-amyloid peptide-(1-42). Lithium-induced Bcl-2 upregulation was temporally associated with the cytoprotective effects of this drug. Thus, lithium protection against beta-amyloid peptide neurotoxicity might involve Bcl-2 overexpression, and lithium treatment for Alzheimer's disease should be reexamined.

Interactions between glutamatergic mechanisms mediated by receptors of the ionotropic and metabotropic classes in the central nervous system are complex and incompletely understood. To explore the consequences of these interactions on excitotoxicity, we examined the influence of group II and group III selective metabotropic glutamate receptor agonists on the N-methyl-D-aspartate-induced apoptotic destruction of GABAergic neurons in rat striatum. The intrastriatal administration of a group III metabotropic glutamate receptor agonist (amino-4-phosphonobutyric acid, 900-1800 nmol), but not of a group II agonist [(2S,1'S,2'S)-(carboxycyclopropyl)glycine, 100-1800 nmol] produced internucleosomal DNA fragmentation. Similarly, amino-4-phosphonobutyric acid (600 nmol) but not (2S,1'S,2'S)-(carboxycyclopropyl)glycine (100-1800 nmol) destroyed some striatal neurons as indicated by a loss of D1 dopamine receptors and 67,000 mol. wt glutamate decarboxylase (glutamate decarboxylase-67) messenger RNA. On the other hand, the intensity of internucleosomal DNA fragmentation induced by N-methyl-D aspartate (150 nmol) was substantially decreased by the intrastriatal co-administration of either (2S,1'S,2'S)-(carboxycyclopropyl)glycine or amino-4-phosphonobutyric acid (100-600 nmol). Both (2S, 1'S,2'S)-(carboxycyclopropyl)glycine and amino-4-phosphonobutyric acid also reduced the N-methyl-D-aspartate-induced loss of striatal D1 dopamine receptors by 67% and 68% (both P0.001), and glutamate decarboxylase-67 messenger RNA by 68% and 61%, respectively. Furthermore, both (2S,1'S,2'S)-(carboxycyclopropyl)glycine and amino-4-phosphonobutyric acid also attenuated the N-methyl-D-aspartate-induced decline in striatal IKB-alpha protein levels by 62% and 37%, as well as the increase in nuclear transcription factor nuclear factor-kappaB binding activity by 135% and 94% (both P0.001), and the subsequent rise in p53 and c-Myc protein levels. These results suggest that stimulation of cyclic-AMP-linked metabotropic glutamate receptors inhibits ionotropic glutamate receptor-mediated activation of apoptotic cascades involving IkappaB-alpha degradation and nuclear factor-kappaB nuclear translocation, as well as p53 and c-Myc induction. Certain selective metabotropic glutamate receptor agonists might thus find utility as adjuncts to N-methyl-D-aspartate antagonists in the protection against the neurotoxicity initiated by excessive ionotropic glutamate receptor stimulation.

We recently reported that cytosine arabinoside (AraC)-induced apoptosis of cerebellar neurons involves the overexpression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The present study was undertaken to investigate whether p53 and/or Bax overexpression participates in the AraC-induced apoptosis of cerebellar granule cells and, if so, the relationship between p53 induction and GAPDH overexpression in these cells.AraC-induced apoptosis of cerebellar granule cells was preceded by an increase in levels of p53 mRNA and protein detected between 1 and 8 hr after treatment. The mRNA level for a p53 target gene, Bax, was also increased. The increase in GAPDH mRNA lasted longer than that of either p53 or Bax, and the level of GAPDH protein in the particulate fraction increased after induction of GAPDH mRNA. The antisense oligonucleotide to p53 protected granule cells from AraC-induced chromatin condensation, internucleosomal cleavage, and apoptotic death. The inhibition of p53 expression by the p53 antisense oligonucleotide not only blocked the expression of Bax but also partially suppressed the increased GAPDH mRNA and protein levels. Conversely, the suppression of GAPDH expression and subsequent attenuation of apoptosis of granule cells by GAPDH antisense oligonucleotide did not influence the expression of p53 or Bax. Cerebellar granule cells prepared from p53 knock-out mice were resistant to AraC toxicity, and the p53 gene knock-out suppressed AraC-upregulated GAPDH expression. Moreover, infection of PC12 cells with an adenoviral vector containing p53 gene dramatically increased GAPDH expression and triggered cell apoptosis. These results suggest that AraC-induced apoptosis of cerebellar granule cells involves the expression of both GAPDH and p53 and that, similar to Bax, GAPDH is upregulated by p53 after exposure to the apoptotic insult.

Nuclear factor kappaB (NF-kappaB) appears to participate in the excitotoxin-induced apoptosis of striatal medium spiny neurons. To elucidate molecular mechanisms by which this transcription factor contributes to NMDA receptor-triggered apoptotic cascades in vivo, rats were given the NMDA receptor agonist quinolinic acid (QA) by intrastriatal infusion, and the role of NF-kappaB in the induction of apoptosis-related genes and gene products was evaluated. QA administration induced time-dependent NF-kappaB nuclear translocation. The nuclear NF-kappaB protein after QA treatment was comprised mainly of p65 and c-Rel subunits as detected by gel supershift assay. Levels of c-Myc and p53 mRNA and protein were markedly increased at the time of QA-induced NF-kappaB nuclear translocation. Immunohistochemical analysis showed that c-Myc and p53 induction occurred in the excitotoxin-sensitive medium-sized striatal neurons. NF-kappaB nuclear translocation was blocked in a dose-dependent manner by the cell-permeable recombinant peptide NF-kappaB SN50, but not by the NF-kappaB SN50 control peptide. NF-kappaB SN50 significantly inhibited the QA-induced elevation in levels of c-Myc and p53 mRNA and protein. Pretreatment or posttreatment with NF-kappaB SN50, but not the control peptide, also substantially reduced the intensity of QA-induced internucleosomal DNA fragmentation. The results suggest that NF-kappaB may promote an apoptotic response in striatal medium-sized neurons to excitotoxic insult through upregulation of c-Myc and p53. This study also provides evidence indicating an unique signaling pathway from the cytoplasm to the nucleus, which regulates p53 and c-Myc levels in these neurons during apoptosis.