RESUMEN
Dextromethorphan (DM) abuse produces mania-like symptoms in humans. ERK/Akt signaling activation involved in manic potential can be attenuated by the inhibition of ouabain-like cardiac steroids. In this study, increased phosphorylations of ERK/Akt and hyperlocomotion induced by DM (30 mg/kg, i.p./day × 7) were significantly protected by the ouabain inhibitor rostafuroxin (ROSTA), suggesting that DM induces the manic potential. ROSTA significantly attenuated DM-induced protein kinase C δ (PKCδ) phosphorylation, GluN2B (i.e., MDA receptor subunit) expression, and phospho-PKCδ/GluN2B interaction. DM instantly upregulated the nuclear factor erythroid-2-related factor 2 (Nrf2)-dependent system. However, DM reduced Nrf2 nuclear translocation, Nrf2 DNA binding activity, γ-glutamylcysteine mRNA expression, and subsequent GSH/GSSG level and enhanced oxidative parameters following 1-h of administration. ROSTA, PKCδ inhibitor rottlerin, and GluN2B inhibitor traxoprodil significantly attenuated DM-induced alterations in Nrf2-related redox parameters and locomotor activity induced by DM in wild-type mice. Importantly, in PKCδ knockout mice, DM failed to alter the above parameters. Further, ROSTA and traxoprodil also failed to enhance PKCδ depletion effect, suggesting that PKCδ is a critical target for the anti-manic potential of ROSTA or GluN2B antagonism. Our results suggest that ROSTA inhibits DM-induced manic potential by attenuating ERK/Akt activation, GluN2B/PKCδ signalings, and Nrf2-dependent system.
Asunto(s)
Androstanoles/farmacología , Trastorno Bipolar , Dextrometorfano/efectos adversos , Ouabaína/antagonistas & inhibidores , Animales , Trastorno Bipolar/inducido químicamente , Trastorno Bipolar/metabolismo , Modelos Animales de Enfermedad , Locomoción/efectos de los fármacos , Masculino , Ratones , Transducción de Señal/efectos de los fármacosRESUMEN
It has been recognized that serotonin 2A receptor (5-HT2A) agonist 2,5-dimethoxy-4-iodo-amphetamine (DOI) impairs serotonergic homeostasis. However, the mechanism of DOI-induced serotonergic behaviors remains to be explored. Moreover, little is known about therapeutic interventions against serotonin syndrome, although evidence suggests that ginseng might possess modulating effects on the serotonin system. As ginsenoside Re (GRe) is well-known as a novel antioxidant in the nervous system, we investigated whether GRe modulates 5-HT2A receptor agonist DOI-induced serotonin impairments. We proposed that protein kinase Cδ (PKCδ) mediates serotonergic impairments. Treatment with GRe or 5-HT2A receptor antagonist MDL11939 significantly attenuated DOI-induced serotonergic behaviors (i.e., overall serotonergic syndrome behaviors, head twitch response, hyperthermia) by inhibiting mitochondrial translocation of PKCδ, reducing mitochondrial glutathione peroxidase activity, mitochondrial dysfunction, and mitochondrial oxidative stress in wild-type mice. These attenuations were in line with those observed upon PKCδ inhibition (i.e., pharmacologic inhibitor rottlerin or PKCδ knockout mice). Furthermore, GRe was not further implicated in attenuation mediated by PKCδ knockout in mice. Our results suggest that PKCδ is a therapeutic target for GRe against serotonergic behaviors induced by DOI.
Asunto(s)
Ginsenósidos/farmacología , Proteína Quinasa C-delta/metabolismo , Antagonistas de la Serotonina/farmacología , Síndrome de la Serotonina/prevención & control , Acetofenonas/farmacología , Anfetaminas/toxicidad , Animales , Conducta Animal/efectos de los fármacos , Conducta Animal/fisiología , Benzopiranos/farmacología , Hipotálamo/efectos de los fármacos , Hipotálamo/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Estrés Oxidativo/efectos de los fármacos , Piperidinas/farmacología , Proteína Quinasa C-delta/deficiencia , Proteína Quinasa C-delta/genética , Inhibidores de Proteínas Quinasas/farmacología , Serotonina/fisiología , Agonistas de Receptores de Serotonina/farmacología , Síndrome de la Serotonina/inducido químicamente , Síndrome de la Serotonina/fisiopatologíaRESUMEN
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 (8â¯mg/kg, i.p.,â¯×â¯4; 2â¯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.
Asunto(s)
Neuronas Dopaminérgicas/metabolismo , Epóxido Hidrolasas/metabolismo , Metanfetamina/efectos adversos , Síndromes de Neurotoxicidad/metabolismo , Proteína Quinasa C-delta/metabolismo , Acetofenonas/farmacología , Animales , Benzopiranos/farmacología , Neuronas Dopaminérgicas/efectos de los fármacos , Epóxido Hidrolasas/genética , Fiebre/genética , Técnicas de Inactivación de Genes , Locomoción/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Síndromes de Neurotoxicidad/genética , Estrés Oxidativo/genética , Proteína Quinasa C-delta/genéticaRESUMEN
Serotonin syndrome is an adverse reaction due to increased serotonin (5-hydroxytryptophan: 5-HT) concentrations in the central nervous system (CNS). The full 5-HT1A receptor (5-HT1AR) agonist (±)-8-hydroxy-dipropylaminotetralin (8-OH-DPAT) has been recognized to elicit traditional serotonergic behaviors. Treatment with 8-OH-DPAT selectively increased PKCδ expression out of PKC isoforms and 5-HT turnover rate in the hypothalamus of wild-type mice. Treatment with 8-OH-DPAT resulted in oxidative burdens, co-immunoprecipitation of 5-HT1AR and PKCδ, and phosphorylation and membrane translocation of p47phox. Importantly, p47phox also interacted with 5-HT1AR or PKCδ in the presence of 8-OH-DPAT. Consistently, the interaction and oxidative burdens were attenuated by 5-HT1AR antagonism (i.e., WAY100635), PKCδ inhibition (i.e., rottlerin and genetic depletion of PKCδ), or NADPH oxidase/p47phox inhibition (i.e., apocynin and genetic depletion of p47phox). However, WAY100635, apocynin, or rottlerin did not exhibit any additive effects against the protective effect by inhibition of PKCδ or p47phox. Furthermore, apocynin, rottlerin, or WAY100635 also significantly protected from pro-inflammatory/pro-apoptotic changes induced by 8-OH-DPAT. Therefore, we suggest that 8-OH-DPAT-induced serotonergic behaviors requires oxidative stress, pro-inflammatory, and pro-apoptotic changes, that PKCδ or p47phox mediates the serotonergic behaviors induced by 8-OH-DPAT, and that the inhibition of PKCδ-dependent p47phox activation is critical for protecting against serotonergic behaviors.
Asunto(s)
NADPH Oxidasas/metabolismo , Proteína Quinasa C-delta/metabolismo , Receptor de Serotonina 5-HT1A/metabolismo , Agonistas del Receptor de Serotonina 5-HT1/administración & dosificación , Síndrome de la Serotonina/tratamiento farmacológico , Serotonina/metabolismo , 8-Hidroxi-2-(di-n-propilamino)tetralin , Animales , Conducta Animal/efectos de los fármacos , Hipotálamo/efectos de los fármacos , Hipotálamo/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , NADPH Oxidasas/genética , Fosforilación/efectos de los fármacos , Unión Proteica , Proteína Quinasa C-delta/genética , Receptor de Serotonina 5-HT1A/genética , Síndrome de la Serotonina/genética , Síndrome de la Serotonina/metabolismo , Síndrome de la Serotonina/psicologíaRESUMEN
We investigated whether protein kinase Cδ (PKCδ) mediates cocaine-induced hepatotoxicity in mice. Cocaine treatment (60â¯mg/kg, i.p.) significantly increased cleaved PKCδ expression in the liver of wild-type (WT) mice, and led to significant increases in oxidative parameters (i.e., reactive oxygen species, 4-hydroxylnonenal and protein carbonyl). These cocaine-induced oxidative burdens were attenuated by pharmacological (i.e., rottlerin) or genetic depletion of PKCδ. We also demonstrated that treatment with cocaine resulted in significant increases in nuclear factor erythroid-2-related factor 2 (Nrf-2) nuclear translocation and increased Nrf-2 DNA-binding activity in wild-type (WT) mice. These increases were more pronounced in the rottlerin-treated WT or PKCδ knockout mice than in the saline-treated WT mice. Although cocaine treatment increased Nrf-2 nuclear translocation, DNA binding activity, and γ-glutamyl cysteine ligases (i.e., GCLc and GCLm) mRNA expressions, while it reduced the glutathione level and GSH/GSSG ratio. These decreases were attenuated by PKCδ depletion. Cocaine treatment significantly increased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in the serum of WT mice signifying the hepatic damage. These increases were also attenuated by PKCδ depletion. In addition, cocaine-induced hepatic degeneration in WT mice was evident 1â¯d post-cocaine. At that time, cocaine treatment decreased Bcl-2 and Bcl-xL levels, and increased Bax, cytosolic cytochrome c, and cleaved caspase-3 levels. Pharmacological or genetic depletion of PKCδ significantly ameliorated the pro-apoptotic properties and hepatic degeneration. Therefore, our results suggest that inhibition of PKCδ, as well as activation of Nrf-2, is important for protecting against hepatotoxicity induced by cocaine.
Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas/genética , Cocaína/toxicidad , Proteína Quinasa C-delta/deficiencia , Proteína Quinasa C-delta/genética , Animales , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 2 Relacionado con NF-E2/metabolismo , Proteína Quinasa C-delta/metabolismoRESUMEN
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.