RESUMO
The present study shows that intranigral injection of dicoumarol, a DT-diaphorase inhibitor, potentiates the neurotoxic effect of salsolinol (salsolinol 1.25 nmoles plus dicoumarol 2 nmoles; in 2 microl). Rats treated with dicoumarol plus salsolinol presented a characteristic contralateral rotational behaviour when they were stimulated with apomorphine (0.5 mg/kg, s.c.), similar to rats injected unilaterally with 6-hydroxydopamine (6-OHDA). These rats also exhibited impairment of motor and cognitive behaviours. The results support the hypothesis that DT-diaphorase plays a protective role in the nigrostriatal dopaminergic systems.
Assuntos
Inibidores Enzimáticos/administração & dosagem , Isoquinolinas/toxicidade , NAD(P)H Desidrogenase (Quinona)/antagonistas & inibidores , Substância Negra/efeitos dos fármacos , Animais , Dicumarol/administração & dosagem , Sinergismo Farmacológico , Injeções Intraventriculares , Masculino , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , NAD(P)H Desidrogenase (Quinona)/metabolismo , Ratos , Ratos Sprague-Dawley , Substância Negra/fisiologiaRESUMO
The exact mechanism of cell death in neurodegenerative diseases remains obscure, although there is evidence that their pathogenesis may involve the formation of free radicals originating from the oxidative metabolism of catecholamines. The purpose of this study was to evaluate the degree of neurodegenerative changes and behavioral impairments induced by unilateral injection into the rat substantia nigra of cyclized o-quinones, aminochrome and dopachrome, derived from oxidizing dopamine and L-DOPA, respectively, with Mn(3+)-pyrophosphate complex. The behavioral changes were compared with those induced after selective lesions of dopaminergic neurons with 6-hydroxydopamine (6-OHDA). Intranigral injection of aminochrome and dopachrome produced impairment in motor and cognitive behaviors. The behavioral impairment was also revealed by apomorphine-induced rotational asymmetry. Apomorphine (0.5 mg/kg sc) significantly increased rotational behavior in rats injected with aminochrome and dopachrome. These rats presented a clear motor bias showing a significant contralateral rotation activity, similar but less vigorous that in rats injected with 6-OHDA. The avoidance conditioning was seriously impaired in rats injected with aminochrome and dopachrome although only dopachrome-injected rats showed a similar hypomotility to 6-OHDA-injected rats. The behavioral effects were correlated to the extent of striatal tyrosine hydroxylase (TH)-positive fiber loss. Rats receiving unilateral intranigral aminochrome and dopachrome injections exhibited a 47.9+/-5.1% and a 39.7+/-4.4% reduction in nigrostriatal TH-positive fiber density. In conclusion, this study provided evidence that oxidizing DA and L-DOPA to cytotoxic quinones, aminochrome and dopachrome appears to be an important mediator of oxidative damage in vivo.
Assuntos
Aprendizagem da Esquiva/efeitos dos fármacos , Indolquinonas , Indóis/farmacologia , Atividade Motora/efeitos dos fármacos , Quinonas/farmacologia , Substância Negra/efeitos dos fármacos , Animais , Aprendizagem da Esquiva/fisiologia , Atividade Motora/fisiologia , Oxidopamina/toxicidade , Ratos , Ratos Sprague-Dawley , Substância Negra/fisiologiaRESUMO
Intracerebral manganese administration together with the DT-diaphorase inhibitor dicoumarol [Mn(III) 40 nmol + -dicoumarol 2 nmol; in 4 micro l] into the left medial forebrain bundle (MFB) produced a behavioural pattern characterized by contralateral behaviour when the rats were stimulated with apomorphine (0.5 mg/kg s.c.), in a manner similar to that when administered to unilaterally 6-hydroxy-dopamine-lesioned animals. The same animals rotated towards the opposite side (ipsilaterally) when stimulated with d-amphetamine (2 mg/kg s.c.). These results support the idea that DT-diaphorase plays a protective role in the dopaminergic systems.
RESUMO
Monoamine oxidase-A (MAO-A) [amiflamine (AMF) and 4-methylthioamphetamine (MTA)] and MAO-B (L-deprenyl) inhibitors were found to be cytotoxic in a concentration-dependent manner for RCHT cells derived from adult rat hypothalamus. The cytotoxic effects were increased when the inhibitors were co-incubated with dicoumarol and especially with 25 micro M AMF+100 micro M dicoumarol (2.5-fold; P <0.001). The treatment of RCHT cells solely with AMF induced a marked decrease in the expression of DT-diaphorase mRNA.
RESUMO
The endogenous dopamine-derived neurotoxin salsolinol was found to decrease survival in the dopaminergic neuronal cell line RCSN-3, derived from adult rat substantia nigra in a concentration-dependent manner (208 microM salsolinol induced a 50% survival decrease). Incubation of RCSN-3 cells with 100 micro;M dicoumarol and salsolinol significantly decreased cell survival by 2.5-fold (P < 0.001), contrasting with a negligible effect on RCHT cells, which exhibited nearly a 5-fold lower nomifensine-insensitive dopamine uptake. The levels of catalase and glutathione peroxidase mRNA were decreased when RCSN-3 cells were treated with 100 microM salsolinol alone or in the presence of 100 microM dicoumarol. In vitro oxidation of salsolinol to o-quinone catalyzed by lactoperoxidase gave the quinone methide and 1,2-dihydro-1-methyl-6,7-isoquinoline diol as final products of salsolinol oxidation as determined by NMR analysis. Evidence of the formation of salsolinol o-semiquinone radical has been provided by ESR studies during one-electron oxidation of salsolinol catalyzed by lactoperoxidase.
Assuntos
Sobrevivência Celular/efeitos dos fármacos , Dopamina/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Indolquinonas , Indóis/farmacologia , Isoquinolinas/farmacologia , Neurônios/efeitos dos fármacos , Quinonas/farmacologia , Animais , Transporte Biológico/efeitos dos fármacos , Catalase/genética , Linhagem Celular , Dicumarol/farmacologia , Espectroscopia de Ressonância de Spin Eletrônica , Glutationa Peroxidase/genética , Neurônios/citologia , Neurônios/metabolismo , RNA Mensageiro/genética , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Substância Negra/citologia , Superóxido Dismutase/genética , Transcrição Gênica/efeitos dos fármacosRESUMO
The mechanism of copper (Cu) neurotoxicity was studied in the RCSN-3 neuronal dopaminergic cell line, derived from substantia nigra of an adult rat. The formation of a Cu-dopamine complex was accompanied by oxidation of dopamine to aminochrome. We found that the Cu-dopamine complex mediates the uptake of (64)CuSO(4) into the Raúl Caviedes substantia nigra-clone 3 (RCSN3) cells, and it is inhibited by the addition of excess dopamine (2 m M) (63%, p < 0.001) and nomifensine (2 microM) (77%, p < 0.001). Copper sulfate (1 m M) alone was not toxic to RCSN-3 cells, but was when combined with dopamine or with dicoumarol (95% toxicity; p < 0.001) which inhibits DPNH and TPNH (DT)-diaphorase. Electron spin resonance (ESR) spectrum of the 5,5-dimethylpyrroline-N-oxide (DMPO) spin trap adducts showed the presence of a C-centered radical when incubating cells with dopamine, CuSO(4) and dicoumarol. A decrease in the expression of CuZn-superoxide dismutase and glutathione peroxidase mRNA was observed when RCSN-3 cells were treated with CuSO(4), dopamine, or CuSO(4) and dopamine. However, the mRNA expression of glutathione peroxidase remained at control levels when the cells were treated with CuSO(4), dopamine and dicoumarol. The regulation of catalase was different since all the treatments with CuSO(4) increased the expression of catalase mRNA. Our results suggest that copper neurotoxicity is dependent on: (i) the formation of Cu-dopamine complexes with concomitant dopamine oxidation to aminochrome; (ii) dopamine-dependent Cu uptake; and (iii) one-electron reduction of aminochrome.
Assuntos
Sulfato de Cobre/toxicidade , Dopamina/farmacologia , Indolquinonas , Indóis/metabolismo , Transporte de Íons/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Substância Negra/citologia , Animais , Catalase/biossíntese , Catalase/genética , Linhagem Celular , Sulfato de Cobre/metabolismo , Sulfato de Cobre/farmacologia , Dicumarol/toxicidade , Espectroscopia de Ressonância de Spin Eletrônica , Indução Enzimática/efeitos dos fármacos , Glutationa Peroxidase/biossíntese , Glutationa Peroxidase/genética , Metalotioneína/metabolismo , NAD(P)H Desidrogenase (Quinona)/biossíntese , NAD(P)H Desidrogenase (Quinona)/genética , Neurônios/metabolismo , Nomifensina/farmacologia , Oxirredução , Estresse Oxidativo , Doença de Parkinson/metabolismo , RNA Mensageiro/biossíntese , Ratos , Superóxido Dismutase/biossíntese , Superóxido Dismutase/genéticaRESUMO
Human glutathione transferase M2-2 prevents the formation of neurotoxic aminochrome and dopachrome by catalyzing the conjugation of dopamine and dopa o-quinone with glutathione. NMR analysis of dopamine and dopa o-quinone-glutathione conjugates revealed that the addition of glutathione was at C-5 to form 5-S-glutathionyl-dopamine and 5-S-glutathionyl-dopa, respectively. Both conjugates were found to be resistant to oxidation by biological oxidizing agents such as O(2), H(2)O(2), and O(*-)(2), and the glutathione transferase-catalyzed reaction can therefore serve a neuroprotective antioxidant function.