Ginsenoside Re protects against kainate-induced neurotoxicity in mice by attenuating mitochondrial dysfunction through activation of the signal transducers and activators of transcription 3 signaling.

It was demonstrated that ginsenosides exert anti-convulsive potentials and interleukin-6 (IL-6) is protective from excitotoxicity induced by kainate (KA), a model of temporal lobe epilepsy. Ginsenosides-mediated mitochondrial recovery is essential for attenuating KA-induced neurotoxicity, however, little is known about the effects of ginsenoside Re (GRe), one of the major ginsenosides. In this study, GRe significantly attenuated KA-induced seizures in mice. KA-induced redox changes were more evident in mitochondrial fraction than in cytosolic fraction in the hippocampus of mice. GRe significantly attenuated KA-induced mitochondrial oxidative stress (i.e. increases in reactive oxygen species, 4-hydroxynonenal, and protein carbonyl) and mitochondrial dysfunction (i.e. the increase in intra-mitochondrial Ca2+ and the decrease in mitochondrial membrane potential). GRe or mitochondrial protectant cyclosporin A restored phospho-signal transducers and activators of transcription 3 (STAT3) and IL-6 levels reduced by KA, and the effects of GRe were reversed by the JAK2 inhibitor AG490 and the mitochondrial toxin 3-nitropropionic acid (3-NP). Thus, we used IL-6 knockout (KO) mice to investigate whether the interaction between STAT3 and IL-6 is involved in the GRe effects. Importantly, KA-induced reduction of manganese superoxide dismutase (SOD-2) levels and neurodegeneration (i.e. astroglial inhibition, microglial activation, and neuronal loss) were more prominent in IL-6 KO than in wild-type (WT) mice. These KA-induced detrimental effects were attenuated by GRe in WT and, unexpectedly, IL-6 KO mice, which were counteracted by AG490 and 3-NP. Our results suggest that GRe attenuates KA-induced neurodegeneration via modulating mitochondrial oxidative burden, mitochondrial dysfunction, and STAT3 signaling in mice.

Ginsenoside Re blocks Bay k-8644-induced neurotoxicity via attenuating mitochondrial dysfunction and PKCδ activation in the hippocampus of mice: Involvement of antioxidant potential.

Although the anticonvulsant effects of ginsenosides are recognized, little is known about their effects on the convulsive behaviors induced by the activation of L-type Ca2+ channels. Here, we investigated whether ginsenoside Re (GRe) modulates excitotoxicity induced by the L-type Ca2+ channel activator Bay k-8644. GRe significantly attenuated Bay k-8644-induced convulsive behaviors and hippocampal oxidative stress in mice. GRe-mediated antioxidant potential was more pronounced in the mitochondrial fraction than cytosolic fraction. As L-type Ca2+ channels are thought to be targets of protein kinase C (PKC), we investigated the role of PKC under excitotoxic conditions. GRe attenuated Bay k-8644-induced mitochondrial dysfunction, PKCδ activation, and neuronal loss. The PKCδ inhibition and neuroprotection mediated by GRe were comparable to those by the ROS inhibitor N-acetylcysteine, the mitochondrial protectant cyclosporin A, the microglial inhibitor minocycline, or the PKCδ inhibitor rottlerin. Consistently, the GRe-mediated PKCδ inhibition and neuroprotection were counteracted by the mitochondrial toxin 3-nitropropionic acid or the PKC activator bryostatin-1. GRe treatment did not have additional effects on PKCδ gene knockout-mediated neuroprotection, suggesting that PKCδ is a molecular target of GRe. Collectively, our results suggest that GRe-mediated anticonvulsive/neuroprotective effects require the attenuation of mitochondrial dysfunction and altered redox status and inactivation of PKCδ.

Methamphetamine-induced dopaminergic neurotoxicity as a model of Parkinson's disease.

Parkinson's disease (PD) is a progressive neurodegenerative disease with a high prevalence, approximately 1 % in the elderly population. Numerous studies have demonstrated that methamphetamine (MA) intoxication caused the neurological deficits and nigrostriatal damage seen in Parkinsonian conditions, and subsequent rodent studies have found that neurotoxic binge administration of MA reproduced PD-like features, in terms of its symptomatology and pathology. Several anti-Parkinsonian medications have been shown to attenuate the motor impairments and dopaminergic damage induced by MA. In addition, it has been recognized that mitochondrial dysfunction, oxidative stress, pro-apoptosis, proteasomal/autophagic impairment, and neuroinflammation play important roles in inducing MA neurotoxicity. Importantly, MA neurotoxicity has been shown to share a common mechanism of dopaminergic toxicity with that of PD pathogenesis. This review describes the major findings on the neuropathological features and underlying neurotoxic mechanisms induced by MA and compares them with Parkinsonian pathogenesis. Taken together, it is suggested that neurotoxic binge-type administration of MA in rodents is a valid animal model for PD that may provide knowledge on the neuropathogenesis of PD.

GPx-1-encoded adenoviral vector attenuates dopaminergic impairments induced by methamphetamine in GPx-1 knockout mice through modulation of NF-κB transcription factor.

We suggested that selenium-dependent glutathione peroxidase (GPx) plays a protective role against methamphetamine (MA)-induced dopaminergic toxicity. We focused on GPx-1, a major selenium-dependent enzyme and constructed a GPx-1 gene-encoded adenoviral vector (Ad-GPx-1) to delineate the role of GPx-1 in MA-induced dopaminergic neurotoxicity. Exposure to Ad-GPx-1 significantly induced GPx activity and GPx-1 protein levels in GPx-1-knockout (GPx-1-KO) mice. MA-induced dopaminergic impairments [i.e., hyperthermia; increased nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) DNA-binding activity; and decreased dopamine levels, TH activity, and behavioral activity] were more pronounced in GPx-1-KO mice than in WT mice. In contrast, exposure to Ad-GPx-1 significantly attenuated MA-induced dopaminergic loss in GPx-1-KO mice. The protective effect exerted by Ad-GPx-1 was comparable to that exerted by pyrrolidine dithiocarbamate (PDTC), an NF-κB inhibitor against MA insult. Consistently, GPx-1 overexpression significantly attenuated MA dopaminergic toxicity in mice. PDTC did not significantly impact the protective effect of GPx-1 overexpression, suggesting that interaction between NF-κB and GPx-1 is critical for dopaminergic protection. Thus, NF-κB is a potential therapeutic target for GPx-1-mediated dopaminergic protective activity. This study for the first time demonstrated that Ad-GPx-1 rescued dopaminergic toxicity in vivo following MA insult. Furthermore, GPx-1-associated therapeutic interventions may be important against dopaminergic toxicity.

5-HT2A receptor-mediated PKCδ phosphorylation is critical for serotonergic impairments induced by p-chloroamphetamine in mice.

p-Chloroamphetamine (PCA), an amphetamine derivative, has been shown to induce serotonergic toxicity. However, the precise mechanism of serotonergic toxicity induced by PCA remains unclear. In this study, PCA treatment (20 mg/kg, i.p.) did not significantly change 5-HT1A receptor gene expression, but significantly increased 5-HT2A receptor gene expression. Furthermore, 5-HT2A receptor antagonist MDL11939, but not 5-HT1A receptor antagonist WAY100635, significantly attenuated PCA-induced serotonergic impairments. We investigated whether PCA activated a specific isoform of protein kinase C (PKC), since previous evidence indicated the involvement of PKC in neurotoxicity induced by amphetamines. We observed that PCA treatment significantly increased the expression levels of PKCδ among all PKC isoforms. MDL11939 treatment significantly attenuated PCA-induced phosphorylation of PKCδ. However, PCA-induced increase in 5-HT2A receptor gene expression was not altered by rottlerin (a pharmacological inhibitor of PKCδ) in mice, suggesting that 5-HT2A receptor is an upstream molecule for the activation of PKCδ. Rottlerin or PKCδ knockout significantly attenuated serotonergic behaviors. However, MDL11939 did not show any additional effects against the attenuation caused by PKCδ knockout in mice, suggesting that PKCδ gene is a molecular target for 5-HT2A receptor-mediated serotonergic effects. Our results suggest that 5-HT2A receptor mediates PCA-induced serotonergic impairments via activation of PKC.δ.

Protein kinase Cδ mediates methamphetamine-induced dopaminergic neurotoxicity in mice via activation of microsomal epoxide hydrolase.

We previously demonstrated that activation of protein kinase Cδ (PKCδ) is critical for methamphetamine (MA)-induced dopaminergic toxicity. It was recognized that microsomal epoxide hydrolase (mEH) also induces dopaminergic neurotoxicity. It was demonstrated that inhibition of PKC modulates the expression of mEH. We investigated whether MA-induced PKCδ activation requires mEH induction in mice. MA treatment (8 mg/kg, i.p., × 4; 2 h interval) significantly enhanced the level of phosphorylated PKCδ in the striatum of wild type (WT) mice. Subsequently, treatment with MA resulted in significant increases in the expression of cleaved PKCδ and mEH. Treatment with MA resulted in enhanced interaction between PKCδ and mEH. PKCδ knockout mice exhibited significant attenuation of the enhanced mEH expression induced by MA. MA-induced hyperthermia, oxidative stress, proapoptotic potentials, and dopaminergic impairments were attenuated by PKCδ knockout or mEH knockout in mice. However, treating mEH knockout in mice with PKCδ inhibitor, rottlerin did not show any additive beneficial effects, indicating that mEH is a critical mediator of neurotoxic potential of PKCδ. Our results suggest that MA-induced PKCδ activation requires mEH induction as a downstream signaling pathway and that the modulation of the PKCδ and mEH interaction is important for the pharmacological intervention against MA-induced dopaminergic neurotoxicity.

Methiopropamine, a methamphetamine analogue, produces neurotoxicity via dopamine receptors.

Methiopropamine (MPA) is structurally categorized as a thiophene ring-based methamphetamine (MA) derivative. Although abusive potential of MPA was recognized, little is known about the neurotoxic potential of MPA up to now. We investigated whether MPA induces dopaminergic neurotoxicity, and whether MPA activates a specific dopamine receptor. Here, we observed that treatment with MPA resulted in dopaminergic neurotoxicity in a dose-dependent manner. MPA treatment potentiated oxidative parameters (i.e., increases in the level of reactive oxygen species, 4-hydroxynonenal, and protein carbonyl), M1 phenotype-related microglial activity, and pro-apoptotic property (i.e., increases in Bax- and cleaved caspase-3-expressions, while a decrease in Bcl-2-expression). Moreover, treatment with MPA resulted in significant impairments in dopaminergic parameters [i.e., changes in dopamine level, dopamine turnover rate, tyrosine hydroxylase (TH) levels, dopamine transporter (DAT) expression, and vesicular monoamine transporter-2 (VMAT-2) expression], and in behavioral deficits. Both dopamine D1 receptor antagonist SCH23390 and D2 receptor antagonist sulpiride protected from these neurotoxic consequences. Therefore, our results suggest that dopamine D1 and D2 receptors simultaneously mediate MPA-induced dopaminergic neurodegeneration in mice via oxidative burdens, microgliosis, and pro-apoptosis.

Significance of protein kinase C in the neuropsychotoxicity induced by methamphetamine-like psychostimulants.

The abuse of methamphetamine (MA), an amphetamine (AMPH)-type stimulant, has been demonstrated to be associated with various neuropsychotoxicity, including memory impairment, psychiatric morbidity, and dopaminergic toxicity. Compelling evidence from preclinical studies has indicated that protein kinase C (PKC), a large family of serine/threonine protein kinases, plays an important role in MA-induced neuropsychotoxicity. PKC-mediated N-terminal phosphorylation of dopamine transporter has been identified as one of the prerequisites for MA-induced synaptic dopamine release. Consistently, it has been shown that PKC is involved in MA (or AMPH)-induced memory impairment and mania-like behaviors as well as MA drug dependence. Direct or indirect regulation of factors related to neuronal plasticity seemed to be critical for these actions of PKC. In addition, PKC-mediated mitochondrial dysfunction, oxidative stress or impaired antioxidant defense system has been suggested to play a role in psychiatric and cognitive disturbance induced by MA (or AMPH). In MA-induced dopaminergic toxicity, particularly PKCδ has been shown to trigger oxidative stress, mitochondrial dysfunction, pro-apoptotic changes, and neuroinflammation. Importantly, PKCδ may be a key mediator in the positive feedback loop composed of these detrimental events to potentiate MA-induced dopaminergic toxicity. This review outlines the role of PKC and its individual isozymes in MA-induced neuropsychotoxicity. Better understanding on the molecular mechanism of PKCs might provide a great insight for the development of potential therapeutic or preventive candidates for MA (or AMPH)-associated neuropsychotoxicity.

Role of protein kinase Cδ in dopaminergic neurotoxic events.

The pro-apoptotic role of Protein kinase Cδ (PKCδ), a member of the novel PKC subfamily, has been well-documented in various pathological conditions. In the central nervous system, the possible role of PKCδ has been studied, mainly in the condition of dopaminergic loss. It has been suggested that the phosphorylation of PKCδ at tyrosine 311 residue (Tyr311) by redox-sensitive Src family kinases (SFKs) is critical for the caspase-3-mediated proteolytic cleavage, which produces the constitutively active cleaved form of PKCδ. Mitochondrial translocation of cleaved PKCδ has been suggested to facilitate mitochondria-derived apoptosis and oxidative burdens. Moreover, it has been suggested that PKCδ contribute to neuroinflammation through the transformation of microglia into the pro-inflammatory M1 phenotype and the assembly of membrane NADPH oxidase in dopaminergic impairments. Interestingly, mitochondrial respiratory chain inhibitors or neuroinflammogens have shown to induce PKCδ activation in dopaminergic systems. Thus, PKCδ activation may be one of the pivotal causes of neuropathologic events, and could amplify these processes further in a positive feedback manner. Furthermore, PKCδ may play an intermediary role in connecting each neuropathologic event. This review affords insight into the role of PKCδ in various dopaminergic neurotoxic models, which could provide a potential target for mitigating dopaminergic neurotoxicity.

Ginsenoside Re protects methamphetamine-induced dopaminergic neurotoxicity in mice via upregulation of dynorphin-mediated κ-opioid receptor and downregulation of substance P-mediated neurokinin 1 receptor.

BACKGROUND: We previously reported that ginsenoside Re (GRe) attenuated against methamphetamine (MA)-induced neurotoxicity via anti-inflammatory and antioxidant potentials. We also demonstrated that dynorphin possesses anti-inflammatory and antioxidant potentials against dopaminergic loss, and that balance between dynorphin and substance P is important for dopaminergic neuroprotection. Thus, we examined whether GRe positively affects interactive modulation between dynorphin and substance P against MA neurotoxicity in mice. METHODS: We examined changes in dynorphin peptide level, prodynorphin mRNA, and substance P mRNA, substance P-immunoreactivity, homeostasis in enzymatic antioxidant system, oxidative parameter, microglial activation, and pro-apoptotic parameter after a neurotoxic dose of MA to clarify the effects of GRe, prodynorphin knockout, pharmacological inhibition of κ-opioid receptor (i.e., nor-binaltorphimine), or neurokinin 1 (NK1) receptor (i.e., L-733,060) against MA insult in mice. RESULTS: GRe attenuated MA-induced decreases in dynorphin level, prodynorphin mRNA expression in the striatum of wild-type (WT) mice. Prodynorphin knockout potentiated MA-induced dopaminergic toxicity in mice. The imbalance of enzymatic antioxidant system, oxidative burdens, microgliosis, and pro-apoptotic changes led to the dopaminergic neurotoxicity. Neuroprotective effects of GRe were more pronounced in prodynorphin knockout than in WT mice. Nor-binaltorphimine, a κ-opioid receptor antagonist, counteracted against protective effects of GRe. In addition, we found that GRe significantly attenuated MA-induced increases in substance P-immunoreactivity and substance P mRNA expression in the substantia nigra. These increases were more evident in prodynorphin knockout than in WT mice. Although, we observed that substance P-immunoreactivity was co-localized in NeuN-immunreactive neurons, GFAP-immunoreactive astrocytes, and Iba-1-immunoreactive microglia. NK1 receptor antagonist L-733,060 or GRe selectively inhibited microgliosis induced by MA. Furthermore, L-733,060 did not show any additive effects against GRe-mediated protective activity (i.e., antioxidant, antimicroglial, and antiapoptotic effects), indicating that NK1 receptor is one of the molecular targets of GRe. CONCLUSIONS: Our results suggest that GRe protects MA-induced dopaminergic neurotoxicity via upregulatgion of dynorphin-mediated κ-opioid receptor and downregulation of substance P-mediated NK1 R.

Role of dopamine D1 receptor in 3-fluoromethamphetamine-induced neurotoxicity in mice.

3-Fluoromethamphetamine (3-FMA) is an illegal designer drug of methamphetamine (MA) derivative. Up to date, little is known about the neurotoxic potential of 3-FMA. In the present study, we investigated the role of dopamine receptors in neurotoxicity induced by 3-FMA in comparison with MA (35 mg/kg, i.p.) as a control drug. Here we found that 3-FMA (40, 60 or 80 mg/kg, i.p.) produced mortality in a dose-dependent manner in mice. Treatment with 3-FMA (40 mg/kg, i.p.) resulted in significant hyperthermia, oxidative stress and microgliosis (microglial differentiation into M1 phenotype) followed by pro-apoptotic changes and the induction of terminal deoxynucleotidyl transferase dUDP nick end labeling (TUNEL)-positive cells. Moreover, 3-FMA significantly produced dopaminergic impairments [i.e., increase in dopamine (DA) turnover rate and decreases in DA level, and in the expression of tyrosine hydroxylase (TH), dopamine transporter (DAT), and vesicular monoamine transporter 2 (VMAT-2)] with behavioral impairments. These dopaminergic neurotoxic effects of 3-FMA were comparable to those of MA. SCH23390, a dopamine D1 receptor antagonist, but not sulpiride, a dopamine D2 receptor antagonist significantly attenuated 3-FMA-induced neurotoxicity. Although both SCH23390 and sulpiride attenuated MA-induced dopaminergic neurotoxicity, sulpiride is more effective than SCH23390 on the dopaminergic neurotoxicity. Interestingly, SCH23390 treatment positively modulated 3-FMA-induced microglial activation (i.e., SCH23390 inhibited M1 phenotype from 3-FMA insult, but activated M2 phenotype). Therefore, our results suggest that the activation of dopamine D1 receptor is critical to 3-FMA-induced neurotoxicity, while both dopamine D1 and D2 receptors (dopamine D2 receptor > dopamine D1 receptor) mediate MA-induced dopaminergic neurotoxicity.

PKCδ-dependent p47phox activation mediates methamphetamine-induced dopaminergic neurotoxicity.

Protein kinase C (PKC) has been recognized to activate NADPH oxidase (PHOX). However, the interaction between PKC and PHOX in vivo remains elusive. Treatment with methamphetamine (MA) resulted in a selective increase in PKCδ expression out of PKC isoforms. PKCδ co-immunoprecipitated with p47phox, and facilitated phosphorylation and membrane translocation of p47phox. MA-induced increases in PHOX activity and reactive oxygen species were attenuated by knockout of p47phox or PKCδ. In addition, MA-induced impairments in the Nrf-2-related glutathione synthetic system were also mitigated by knockout of p47phox or PKCδ. Glutathione-immunoreactivity was co-localized in Iba-1-labeled microglial cells and in NeuN-labeled neurons, but not in GFAP-labeled astrocytes, reflecting the necessity for self-protection against oxidative stress by mainly microglia. Buthionine-sulfoximine, an inhibitor of glutathione biosynthesis, potentiated microglial activation and pro-apoptotic changes, leading to dopaminergic losses. These neurotoxic processes were attenuated by rottlerin, a pharmacological inhibitor of PKCδ, genetic inhibitions of PKCδ [i.e., PKCδ knockout mice (KO) and PKCδ antisense oligonucleotide (ASO)], or genetic inhibition of p47phox (i.e., p47phox KO or p47phox ASO). Rottlerin did not exhibit any additive effects against the protective activity offered by genetic inhibition of p47phox. Therefore, we suggest that PKCδ is a critical regulator for p47phox activation induced by MA, and that Nrf-2-dependent GSH induction via inhibition of PKCδ or p47phox, is important for dopaminergic protection against MA insult.

Trichloroethylene and Parkinson's Disease: Risk Assessment.

This study was conducted to investigate the mechanism of action and extent of selective dopaminergic neurodegeneration caused by exposure to trichloroethylene (TCE) leading to the endogenous formation of the neurotoxin 1-trichloromethyl-1,2,3,4-tetrahydro-ß-carboline (TaClo) in rodents. Beginning at 3 months of age, male C57BL/6 mice received oral TCE dissolved in vehicle for 8 months. Dopaminergic neuronal loss was assessed by nigral tyrosine hydroxylase (TH) immunoreactivity. Selective dopaminergic neurodegeneration was determined based on histological analysis of non-dopaminergic neurons in the brain. Behavioral assays were evaluated using open field activity and rotarod tests. Mitochondrial complex I activity, oxidative stress markers, and microglial activation were also examined in the substantia nigra. The level of TaClo was detected using HPLC-electrospray ionization tandem mass spectrometry. Dopaminergic neurotoxicity of TaClo was determined in midbrain organotypic cultures from rat pups. Following 8 months of TCE treatment, there was a progressive and selective loss of 50% of the dopaminergic neurons in mouse substantia nigra (SN) and about 50% loss of dopamine and 72% loss of 3,4-dihydroxyphenylacetic acid in the striatum, respectively. In addition, motor deficits, mitochondrial impairment, oxidative stress, and inflammation were measured. TaClo content was quantified in the brain after TCE treatment. In organotypic cultures, TaClo rather than TCE induced dopaminergic neuronal loss, similar to MPP+. TCE exposure may stimulate the endogenous formation of TaClo, which is responsible for dopaminergic neurodegeneration. However, even prolonged administration of TCE was insufficient for producing a greater than 50% loss of nigral dopamine neurons, indicating that additional co-morbid factors would be needed for mimicking the profound loss of dopamine neurons seen in Parkinson's disease.

Ginsenoside Re protects against phencyclidine-induced behavioral changes and mitochondrial dysfunction via interactive modulation of glutathione peroxidase-1 and NADPH oxidase in the dorsolateral cortex of mice.

We investigated whether ginsenoside Re (Re) modulates phencyclidine (PCP)-induced sociability deficits and recognition memory impairments to extend our recent finding. We examined the role of GPx-1 gene in the pharmacological activity of Re against mitochondrial dysfunction induced by PCP in the dorsolateral cortex of mice. Since mitochondrial oxidative stress activates NADPH oxidase (PHOX), we applied PHOX inhibitor apocynin for evaluating interactive modulation between GPx-1 and PHOX against PCP neurotoxicity. Sociability deficits and recognition memory impairments induced by PCP were more pronounced in GPx-1 knockout (KO) than in wild type (WT) mice. PCP-induced mitochondrial oxidative stress, mitochondrial dysfunction, and membrane translocation of p47phox were more evident in GPx-1 KO than in WT. Re treatment significantly attenuated PCP-induced neurotoxic changes. Re also significantly attenuated PCP-induced sociability deficits and recognition memory impairments. The attenuation by Re was comparable to that by apocynin. The attenuation was more obvious in GPx-1 KO than in WT. Importantly, apocynin did not show any additional positive effects on the neuroprotective activity of Re, indicating that PHOX is a molecular target for therapeutic activity of Re. Our results suggest that Re requires interactive modulation between GPx activity and PHOX (p47phox) to exhibit neuroprotective potentials against PCP insult.

The role of system Xc- in methamphetamine-induced dopaminergic neurotoxicity in mice.

The cystine/glutamate antiporter (system Xc-, Sxc) transports cystine into cell in exchange for glutamate. Since xCT is a specific subunit of Sxc, we employed xCT knockout mice and investigated whether this antiporter affected methamphetamine (MA)-induced dopaminergic neurotoxicity. MA treatment significantly increased striatal oxidative burdens in wild type mice. xCT inhibitor [i.e., S-4-carboxy-phenylglycine (CPG), sulfasalazine] or an xCT knockout significantly protected against these oxidative burdens. MA-induced increases in Iba-1 expression and Iba-1-labeled microglial immunoreactivity (Iba-1-IR) were significantly attenuated by CPG or sulfasalazine administration or xCT knockout. CPG or sulfasalazine significantly attenuated MA-induced TUNEL-positive cell populations in the striatum of Taconic ICR mice. The decrease in excitatory amino acid transporter-2 (or glutamate transporter-1) expression and increase in glutamate release were attenuated by CPG, sulfasalazine or xCT knockout. In addition, CPG, sulfasalazine or xCT knockout significantly protected against dopaminergic loss (i.e., decreases in tyrosine hydroxylase expression and immunoreactivity, and an increase in dopamine turnover rate) induced by MA. However, CPG, sulfasalazine or xCT knockout did not significantly affect the impaired glutathione system [i.e., decrease in reduced glutathione (GSH) and increase in oxidized glutathione (GSSG)] induced by MA. Our results suggest that Sxc mediates MA-induced neurotoxicity via facilitating oxidative stress, microgliosis, proapoptosis, and glutamate-related toxicity.

YY-1224, a terpene trilactone-strengthened Ginkgo biloba, attenuates neurodegenerative changes induced by ß-amyloid (1-42) or double transgenic overexpression of APP and PS1 via inhibition of cyclooxygenase-2.

BACKGROUND: Ginkgo biloba has been reported to possess free radical-scavenging antioxidant activity and anti-inflammatory properties. In our pilot study, YY-1224, a terpene trilactone-strengthened extract of G. biloba, showed anti-inflammatory, neurotrophic, and antioxidant effects. RESULTS: We investigated the pharmacological potential of YY-1224 in ß-amyloid (Aß) (1-42)-induced memory impairment using cyclooxygenase-2 (COX-2) knockout (-/-) and APPswe/PS1dE9 transgenic (APP/PS1 Tg) mice. Repeated treatment with YY-1224 significantly attenuated Aß (1-42)-induced memory impairment in COX-2 (+/+) mice, but not in COX-2 (-/-) mice. YY-1224 significantly attenuated Aß (1-42)-induced upregulation of platelet-activating factor (PAF) receptor gene expression, reactive oxygen species, and pro-inflammatory factors. In addition, YY-1224 significantly inhibited Aß (1-42)-induced downregulation of PAF-acetylhydrolase-1 (PAF-AH-1) and peroxisome proliferator-activated receptor γ (PPARγ) gene expression. These changes were more pronounced in COX-2 (+/+) mice than in COX-2 (-/-) mice. YY-1224 significantly attenuated learning impairment, Aß deposition, and pro-inflammatory microglial activation in APP/PS1 Tg mice, whereas it significantly enhanced PAF-AH and PPARγ expression. A preferential COX-2 inhibitor, meloxicam, did not affect the pharmacological activity by YY-1224, suggesting that the COX-2 gene is a critical mediator of the neuroprotective effects of YY-1224. The protective activity of YY-1224 appeared to be more efficacious than a standard G. biloba extract (Gb) against Aß insult. CONCLUSIONS: Our results suggest that the protective effects of YY-1224 against Aß toxicity may be associated with its PAF antagonistic- and PPARγ agonistic-potential as well as inhibition of the Aß-mediated pro-inflammatory switch of microglia phenotypes through suppression of COX-2 expression.

Genetic or pharmacological depletion of cannabinoid CB1 receptor protects against dopaminergic neurotoxicity induced by methamphetamine in mice.

Accumulating evidence suggests that cannabinoid ligands play delicate roles in cell survival and apoptosis decisions, and that cannabinoid CB1 receptors (CB1R) modulate dopaminergic function. However, the role of CB1R in methamphetamine (MA)-induced dopaminergic neurotoxicity in vivo remains elusive. Multiple high doses of MA increased phospho-ERK and CB1R mRNA expressions in the striatum of CB1R (+/+) mice. These increases were attenuated by CB1R antagonists (i.e., AM251 and rimonabant), an ERK inhibitor (U0126), or dopamine D2R antagonist (sulpiride). In addition, treatment with MA resulted in dopaminergic impairments, which were attenuated by CB1R knockout or CB1R antagonists (i.e., AM251 and rimonabant). Consistently, MA-induced oxidative stresses (i.e., protein oxidation, lipid peroxidation and reactive oxygen species) and pro-apoptotic changes (i.e., increases in Bax, cleaved PKCδ- and cleaved caspase 3-expression and decrease in Bcl-2 expression) were observed in the striatum of CB1R (+/+) mice. These toxic effects were attenuated by CB1R knockout or CB1R antagonists. Consistently, treatment with four high doses of CB1R agonists (i.e., WIN 55,212-2 36mg/kg and ACEA 16mg/kg) also resulted in significant oxidative stresses, pro-apoptotic changes, and dopaminergic impairments. Since CB1R co-immunoprecipitates PKCδ in the presence of MA or CB1R agonists, we applied PKCδ knockout mice to clarify the role of PKCδ in the neurotoxicity elicited by CB1Rs. CB1R agonist-induced toxic effects were significantly attenuated by CB1R knockout, CB1R antagonists or PKCδ knockout. Therefore, our results suggest that interaction between D2R, ERK and CB1R is critical for MA-induced dopaminergic neurotoxicity and that PKCδ mediates dopaminergic damage induced by high-doses of CB1R agonist.

Current understanding of methamphetamine-associated dopaminergic neurodegeneration and psychotoxic behaviors.

Clinical and preclinical studies have indicated that chronic methamphetamine (MA) use is associated with extensive neurodegeneration, psychosis, and cognitive impairment. Evidence from animal models has suggested a considerable role of excess dopamine or glutamate, oxidative stress, neuroinflammation, and apoptosis in MA-induced neurotoxicity, and that protein kinase Cδ might mediate the interaction among these factors. In addition, the relatively long-lasting and recurrent nature of MA psychosis has been reproduced in animals treated with various dosing regimens of MA, which have shown behavioral sensitization, sociability deficits, and impaired prepulse inhibition. Genetic predisposition as well as dopaminergic and glutamatergic alterations might be important in the development of MA psychosis. Neuroimaging studies have identified functional and morphological changes related to the cognitive dysfunction shown in chronic MA users. Failure in the task-evoked phosphorylation of extracellular signal-related kinase likely underlies MA-induced memory impairment. Recent progress has suggested certain roles of oxidative stress and neuroinflammation in the psychosis and cognitive deficits induced by repeated low doses of MA. This review provides a comprehensive description of pertinent findings from human and animal studies, with an emphasis on the current understanding of the underlying mechanisms of MA neuropsychotoxicity and its relevance to Parkinson's disease or schizophrenia.

PKCδ knockout mice are protected from para-methoxymethamphetamine-induced mitochondrial stress and associated neurotoxicity in the striatum of mice.

Para-methoxymethamphetamine (PMMA) is a para-ring-substituted amphetamine derivative sold worldwide as an illegal psychotropic drug. Although PMMA use has been reported to lead to severe intoxication and even death, little is known about the mechanism(s) by which PMMA exerts its neurotoxic effects. Here we found that PMMA treatment resulted in phosphorylation of protein kinase Cδ (PKCδ) and subsequent mitochondrial translocation of cleaved PKCδ. PMMA-induced oxidative stress was more pronounced in mitochondria than in the cytosol. Moreover, treatment with PMMA consistently resulted in significant reductions in mitochondrial membrane potential, mitochondrial complex I activity, and mitochondrial Mn superoxide dismutase-immunoreactivity. In contrast, PMMA treatment led to a significant increase in intramitochondrial Ca2+ level. Treatment with PMMA also significantly increased ionized calcium binding adaptor molecule 1 (Iba-1)-labeled microglial activation and upregulated tumor necrosis factor alpha (TNF-α) gene expression. PKCδ knockout attenuated these mitochondrial effects and dampened the neurotoxic effects of PMMA. Importantly, TNF-α knockout mice were significantly protected from PMMA-induced increases in phospho-PKCδ expression, mitochondrial translocation of cleaved PKCδ, and Iba-1-labeled microgliosis. Both rottlerin, a pharmacological inhibitor of PKCδ, and etanercept, a pharmacological inhibitor of TNF-α, significantly protected against PMMA-mediated induction of apoptosis, as assessed by terminal deoxynucleotidyl transferase dUDP nick end labeling (TUNEL) assays. In addition, PKCδ knockout and TNF-α knockout both resulted in decreased PMMA-mediated induction of dopaminergic loss. Therefore, our results suggest that PKCδ mediates PMMA-induced neurotoxicity by facilitating oxidative stress (mitochondria > cytosol), mitochondrial dysfunction, microglial activation, and pro-apoptotic signaling. Our results also indicate that PMMA-induced PKCδ activation requires the proinflammatory cytokine TNF-α.

Apocynin prevents mitochondrial burdens, microglial activation, and pro-apoptosis induced by a toxic dose of methamphetamine in the striatum of mice via inhibition of p47phox activation by ERK.

BACKGROUND: Activation of NADPH oxidase (PHOX) plays a critical role in mediating dopaminergic neuroinflammation. In the present study, we investigated the role of PHOX in methamphetamine (MA)-induced neurotoxic and inflammatory changes in mice. METHODS: We examined changes in mitogen-activated protein kinases (MAPKs), mitochondrial function [i.e., mitochondrial membrane potential, intramitochondrial Ca(2+) accumulation, mitochondrial oxidative burdens, mitochondrial superoxide dismutase expression, and mitochondrial translocation of the cleaved form of protein kinase C delta type (cleaved PKCδ)], microglial activity, and pro-apoptotic changes [i.e., cytosolic cytochrome c release, cleaved caspase 3, and terminal deoxynucleotidyl transferase dUDP nick-end labeling (TUNEL) positive populations] after a neurotoxic dose of MA in the striatum of mice to achieve a better understanding of the effects of apocynin, a non-specific PHOX inhibitor, or genetic inhibition of p47phox (by using p47phox knockout mice or p47phox antisense oligonucleotide) against MA-induced dopaminergic neurotoxicity. RESULTS: Phosphorylation of extracellular signal-regulated kinases (ERK1/2) was most pronounced out of MAPKs after MA. We observed MA-induced phosphorylation and membrane translocation of p47phox in the striatum of mice. The activation of p47phox promoted mitochondrial stresses followed by microglial activation into the M1 phenotype, and pro-apoptotic changes, and led to dopaminergic impairments. ERK activated these signaling pathways. Apocynin or genetic inhibition of p47phox significantly protected these signaling processes induced by MA. ERK inhibitor U0126 did not exhibit any additional positive effects against protective activity mediated by apocynin or p47phox genetic inhibition, suggesting that ERK regulates p47phox activation, and ERK constitutes the crucial target for apocynin-mediated inhibition of PHOX activation. CONCLUSIONS: Our results indicate that the neuroprotective mechanism of apocynin against MA insult is via preventing mitochondrial burdens, microglial activation, and pro-apoptotic signaling process by the ERK-dependent activation of p47phox.