Uticaj NG-nitro-L-arginin metil estra na kliničke i biohemijske efekte metilen plavog kod konvulzija izazvanih pentilentetrazolom

Abstract

Background/Aim. Despite years of research in a number of experimental models the question whether nitric oxide (NO) and methylene blue (MB) have pro- or anticonvulsant effects remains to be fully resolved. Methods. In adult Wistar rats the influence of a nonselective inhibitor of nitric oxide synthase NG-nitro-L-arginine methyl ester (LNAME, 10µg) on clinical and biochemical effects of MB (10µg) given before the intraperitoneally administered chemical convulsant pentylenetetrazole (PTZ, 80 mg/kg) was examined. MB and L-NAME were applied intracerebroventricularly. PTZ application was followed by a 4- minute observation time, after which rats were sacrificed and elements of oxido-reductive balance were measured in a crude mitochondrial fraction of forebrain cortex, hippocampus and striatum. Results. Convulsive responses (forelimb dystonia - FLD, generalised clonic- and clonic-tonic convulsions - GCC and GCTC respectively) were observed in all rats received PTZ, together with significantly decreased lipid peroxidation in the forebrain cortex and striatum and increased superoxide dismutase activity in the hippocampus, in comparison to controls (saline treated). It was registered anticonvulsant effects of L-NAME pretreatment. However, these effects were insignificant. In the hippocampus of these animals there was decreased lipid peroxidation (p < 0.01, p < 0.05 vs saline-treated and PTZ-treated rats, respectively) and reverted PTZ-induced increase of superoxide dismutase activity. But MB individually pretreatment significantly decreased the incidence of CTCs and GCCs (FLD: p = 0.0513), prolonged the convulsive latent time for FLD, GCTCs and GCCs, in all the examined brain regions increased lipid peroxidation and decreased the level of superoxide anion. Administration of L-NAME 10 minutes before MB reverted all MB-evoked clinical and biochemical effects. Conclusion. Methylene blue applied individually before PTZ has strong anticonvulsant effects that were eliminated by L-NAME pretreatment. These effects and changed biochemical parameters in the brains of animals treated by L-NAME before MB in comparison to MBtreated group suggest involvement of NO in MB's effects in the animal model of PTZ-evoked convulsions.

Uvod/Cilj. I pored višegodišnjeg istraživanja na različitim eksperimentalnim modelima, nije potpuno odgovoreno na pitanje da li azot-oksid (NO) i metilen plavo (MP) deluju konvulzivno ili antikonvulzivno. Metode. Na odraslim pacovima Vistar soja ispitivan je uticaj NG-nitro-L-arginin metil estra (L-NAME, 10 µg), neselektivnog inhibitora azot oksid sintaze, na kliničke i biohemijske efekte metilen plavog (MP, 10 µg) datog intracerebroventrikularno pre hemijskog konvulziva pentilentetrazola (PTZ, 80 mg/kg), primenjenog intraperitonealno. Pacovi su posmatrani četiri minuta posle davanja PTZ-a, posle čega su žrtvovani i u neprečišćenoj mitohondrijskoj frakciji prednjeg mozga, hipokampusa i strijatuma određivani su parametri oksidoreduktivne ravnoteže. Rezultati. Posle primene PTZ-a, konvulzivni odgovor (distonija prednjih nogu - DPN, generalizovane klonične - GKK i generalizovane kloničnotonične konvulzije - GKTK) bio je ispoljen kod svih životinja, kao i statistički značajno sniženje lipidne peroksidacije u kori prednjeg mozga i strijatuma, i povećanje aktivnosti superoksid dizmutaze (SOD) u hipokampusu, u poređenju sa kontrolnom grupom (dobila fiziološki rastvor NaCl). Registrovani su antikonvulzivni efekti L-NAME koji nisu bili statistički značajni. U hipokampusu ovih životinja bila je snižena lipidna peroksidacija (p < 0,01 u poređenju sa kontrolnom grupom, p < 0,05 u poređenju sa životinjama koje su dobile PTZ), kao i aktivnost SOD u poređenju sa životinjama koje su dobile PTZ. Samo metilen plavo dovelo je do statistički značajnog smanjenja incidencije GKK I GKTK (DPN: p = 0,0513), produžilo je latentni period DPN, GKK i GKTK, a u svim ispitivanim strukturama mozga bila je povećana lipidna peroksidacija i smanjen nivo superoksidnog anjona. Svi klinički i biohemijski efekti izazvani primenom MP u potpunosti su odstranjeni primenom L-NAME 10 minuta pre davanja MP. Zaključak. Metilen plavo, dat samostalno pre PTZ, ispoljio je snažne antikonvulzivne efekte. Nestanak ovih efekata i izmenjeni biohemijski parametri u mozgovima pacova koji su pre MP dobili L-NAME, sugerišu da je NO uključen u efekte MP ispoljene na životinjskom modelu konvulzija izazvanih primenom PTZ-a.