Long-term treatment with 3, 4-methylenedioxymethamphetamine caused retina damage in C57BL/6 mice / Tratamento a longo prazo com 3, 4-metilenedioximetanfetamina causando dano na retina em camundongos C57BL/6

ABSTRACT Purpose: As a class of psychostimulant drugs, amphetamines are widely abused for their stimulant, euphoric, and hallucinogenic properties. Many of these effects result from acute increases in dopamine and serotonin neurotransmission. Following the onset of these effects, 3,4 methylenedioxymethamphetamine produces persistent damage to dopamine and serotonin nerve terminals, resulting in long-lasting neurotoxicity. The purpose of this investigation was to assess the effects of treatment with low dose of methylenedioxymethamphetamine on retinal function of C57BL/6 mice and its underlying mechanisms. Methods: C57BL/6 mice were divided randomly into two groups (n=10): one group was treated with phosphate buffered saline by intraperitoneal injection daily; the other group was treated with 1 mg/kg methylenedioxymethamphetamine by intraperitoneal injection daily for three months. Electroretinography was used to test retinal function every month. H&E staining and terminal deoxynucleotidyl transferase assay were used to evaluate the retinal morphology and histology. Enzyme-linked immunosorbent assay assays were used to measure markers of oxidative stress and inflammatory factors. Gene and protein expression was detected by real-time PCR and western blot. Results: Three-month treatment with methylenedioxymethamphetamine induced significant retinal dysfunction via photoreceptor cell apoptosis by oxidative stress and inflammatory responses. Conclusions: These results suggest that long-term treatment with methylenedioxymethamphetamine increases inflammatory responses in photoreceptor cells resulting in retinal dysfunction in C57BL/6 mice. Thus, this investigation provides preclinical rationale for the retina damage caused by the methylenedioxymethamphetamine abuse.

RESUMO Objetivos: Como uma classe de drogas psicoesti mulantes, as anfetaminas são amplamente usadas por suas propriedades estimulantes, eufóricas e alucinógenas. Muitos desses efeitos resultam de aumentos agudos na neurotransmissão da dopamina e da serotonina. Após o início desses efeitos, a 3,4-metilenedioximetanfetamina produz danos persistentes nos terminais nervosos de dopamina e serotonina, resultando em neurotoxicidade duradoura. O objetivo desta investigação foi avaliar os efeitos do tratamento baixa dose de metilenedioximetanfetamina na função da retina em camundongos C57BL/6 e seus mecanismos subjacentes. Métodos: Camundongos C57BL/6 foram divididos aleatoriamente em dois grupos (n=10): um grupo foi tratado com solução salina tamponada de fosfato por injeção intraperitoneal diária; o outro grupo foi tratado com 1 mg/kg de metilenedioximetanfetamina por injeção intraperitoneal diária durante 3 meses. Eletroretinografia foi utilizada para testar a função da retina a cada mês. A coloração H&E e análise com deoxinucleotidil terminal transferase foram utilizados para avaliar a morfologia e histologia da retina. Testes de imunoabsorção enzimática foram utilizados para medir marcadores de estresse oxidativo e fatores inflamatórios. A expressão de genes e proteínas foi detectada por PCR em tempo real e western blot. Resultados: O tratamento de três meses com metilenedioximetanfetamina induziu disfunção de retina significativa por apoptose de células fotorreceptoras por estresse oxidativo e resposta inflamatória. Conclusões: Estes resultados sugerem que o tratamento a longo prazo com metilenedioximetanfetamina aumenta as respostas inflamatórias em células fotorreceptoras, resultando em disfunção de retina em camundongos C57BL/6. Assim, a investigação foence uma justificação pré-clínica para os danos na retina causados pelo abuso de metilenedioximetanfetamina.

Delay of Photoreceptor Cell Degeneration in rd Mice by Systemically Administered Phenyl-N-tert-butylnitrone

PURPOSE: To study the effect of systemic administration of phenyl-N-tert-butylnitrone (PBN) on the degeneration of photoreceptor cells in rd mice. METHODS: PBN was injected intraperitoneally into FVB/rd mice on postnatal days (P) 5 to 14 (group A), and P10 to 18 (group B). At days P14, 16, 18, 20 and 27, morphological changes and apoptosis were analyzed by staining with hematoxylin and eosin or DAPI. The effect of PBN on apoptosis was analyzed in retinal pigment epithelial (RPE) cells by the measurement of caspase-3 activity. RESULTS: In control and group B mice, the outer nuclear layer (ONL) of the retina was composed of 8-10 rows at P12, and rapidly decreased to one row at P18. In group A mice, the ONL was preserved with 5-7 rows at P18, and decreased to one row at P22. PBN inhibited caspase-3 activity in cultured RPE cells. CONCLUSIONS: PBN delayed, but did not block, the degeneration of photoreceptor cells in rd mice. PBN may exert its inhibitory effect during the early phase of photoreceptor cell degeneration.