The role of nitric oxide in glutaric acid-induced convulsive behavior in pup rats.

Glutaric acidaemia type I (GA-I) is a cerebral organic disorder characterized by the accumulation of glutaric acid (GA) and seizures. As seizures are precipitated in children with GA-I and the mechanisms underlying this disorder are not well established, we decided to investigate the role of nitric oxide (NO) in GA-induced convulsive behaviour in pup rats. Pup male Wistar rats (18-day-old) were anesthetized and placed in stereotaxic apparatus for cannula insertion into the striatum for injection of GA. The experiments were performed 3 days after surgery (pup rats 21-day-old). An inhibitor of NO synthesis (N-G-nitro-l-arginine methyl ester-L-NAME, 40 mg/kg) or saline (vehicle) was administered intraperitoneally 30 min before the intrastriatal injection of GA (1 µl, 1.3 µmol/striatum) or saline. Immediately after the intrastriatal injections, the latency and duration of seizures were recorded for 20 min. The administration of L-NAME significantly increased the latency to the first seizure episode and reduced the duration of seizures induced by GA in pup rats. The administration of the NO precursor l-arginine (L-ARG; 80 mg/kg) prevented the effects of L-NAME. Besides, GA significantly increased nitrate and nitrite (NOx) levels in the striatum of pup rats and the preadministration of L-NAME prevented this alteration. L-ARG blocked the reduction of striatal NOx provoked by L-NAME. These results are experimental evidence that NO plays a role in the seizures induced by GA in pup rats, being valuable in understanding the physiopathology of neurological signs observed in children with this organic acidaemia and to develop new therapeutic strategies.

Exercise pre-conditioning reduces brain inflammation and protects against toxicity induced by traumatic brain injury: behavioral and neurochemical approach.

Although the favorable effects of physical exercise in neurorehabilitation after traumatic brain injury (TBI) are well known, detailed pathologic and functional alterations exerted by previous physical exercise on post-traumatic cerebral inflammation have been limited. In the present study, it is showed that fluid percussion brain injury (FPI) induced motor function impairment, followed by increased plasma fluorescein extravasation and cerebral inflammation characterized by interleukin-1ß, tumor necrosis factor-α (TNF-α) increase, and decreased IL-10. In addition, myeloperoxidase (MPO) increase and Na⁺,K⁺-ATPase activity inhibition after FPI suggest that the opening of blood-brain barrier (BBB) followed by neurtrophils infiltration and cerebral inflammation may contribute to the failure of selected targets leading to secondary damage. In fact, Pearson's correlation analysis revealed strong correlation of MPO activity increase with Na⁺,K⁺-ATPase activity inhibition in sedentary rats. Statistical analysis also revealed that previous running exercise (4 weeks) protected against FPI-induced motor function impairment and fluorescein extravasation. Previous physical training also induced IL-10 increase per se and protected against cerebral IL-1ß, and TNF-α increase and IL-10 decrease induced by FPI. This protocol of physical training was effective against MPO activity increase and Na⁺,K⁺-ATPase activity inhibition after FPI. The present protection correlated with MPO activity decrease suggests that the alteration of cerebral inflammatory status profile elicited by previous physical training reduces initial damage and limits long-term secondary degeneration after TBI. This prophylactic effect may facilitate functional recovery in patients suffering from brain injury induced by TBI.

Adaptation to oxidative challenge induced by chronic physical exercise prevents Na+,K+-ATPase activity inhibition after traumatic brain injury.

Physical exercise is likely to alter brain function and to afford neuroprotection in several neurological diseases. Although the favorable effects of physical exercise on traumatic brain injury (TBI) patients is well known, little information is available regarding the role of free radicals in the improvement induced by physical exercise in an experimental model of TBI induced by fluid percussion injury (FPI). Thus, we investigated whether 6 weeks of swimming training protects against oxidative damage (measured by protein carbonylation and thiobarbituric acid-reactive substances-TBARS) and neurochemical alterations represented by immunodetection of alpha subunit and activity of Na(+),K(+)-ATPase after FPI in cerebral cortex of rats. Statistical analysis revealed that physical training protected against FPI-induced TBARS and protein carbonylation increase. In addition, physical training was effective against Na(+),K(+)-ATPase enzyme activity inhibition and alpha(1) subunit level decrease after FPI. Pearson's correlation analysis revealed that the decrease in levels of catalytic alpha(1) subunit of Na(+),K(+)-ATPase induced FPI correlated with TBARS and protein carbonylation content increase. Furthermore, the effective protection exerted by physical training against FPI-induced free radical correlated with the immunocontent of the catalytic alpha(1) subunit maintenance. These data suggest that TBI-induced reactive oxygen species (ROS) generation decreases Na(+),K(+)-ATPase activity by decreasing the total number of enzyme molecules, and that physical exercise protects against this effect. Therefore, the effective protection of selected targets, such as Na(+),K(+)-ATPase induced by physical training, supports the idea that physical training may exert prophylactic effects on neuronal cell dysfunction and damage associated with TBI.

Methylmalonate-induced seizures are attenuated in inducible nitric oxide synthase knockout mice.

Methylmalonic acidemias consist of a group of inherited neurometabolic disorders caused by deficiency of methylmalonyl-CoA mutase activity clinically and biochemically characterized by neurological dysfunction, methylmalonic acid (MMA) accumulation, mitochondrial failure and increased reactive species production. Although previous studies have suggested that nitric oxide (NO) plays a role in the neurotoxicity of MMA, the involvement of NO-induced nitrosative damage from inducible nitric oxide synthase (iNOS) in MMA-induced seizures are poorly understood. In the present study, we showed a decrease of time spent convulsing induced by intracerebroventricular administration of MMA (2 micromol/2 microL; i.c.v.) in iNOS knockout (iNOS(-/-)) mice when compared with wild-type (iNOS(+/+)) littermates. Visual analysis of electroencephalographic recordings (EEG) showed that MMA injection induced the appearance of high-voltage synchronic spike activity in the ipsilateral cortex which spreads to the contralateral cortex while quantitative electroencephalographic analysis showed larger wave amplitude during MMA-induced seizures in wild-type mice when compared with iNOS knockout mice. We also report that administration of MMA increases NOx (NO(2) plus NO(3) content) and 3-nitrotyrosine (3-NT) levels in a greater extend in iNOS(+/+) mice than in iNOS(-/-) mice, indicating that NO overproduction and NO-mediated damage to proteins are attenuated in iNOS knockout mice. In addition, the MMA-induced decrease in Na(+), K(+)-ATPase activity, but not in succinate dehydrogenase (SDH) activity, was less pronounced in iNOS(-/-) when compared with iNOS(+/+) mice. These results reinforce the assumption that metabolic collapse contributes for the secondary toxicity elicited by MMA and suggest that oxidative attack by NO derived from iNOS on selected target such as Na(+), K(+)-ATPase enzyme might represent an important role in this excitotoxicity induced by MMA. Therefore, these results may be of value in understating the pathophysiology of the neurological features observed in patients with methylmalonic acidemia and in the development of new strategies for treatment of these patients.

Creatine decreases convulsions and neurochemical alterations induced by glutaric acid in rats.

Glutaric acidemia type I (GA-I) is an inherited metabolic disease characterized by striatal degeneration, seizures, and accumulation of glutaric acid (GA). Considering that GA impairs energy metabolism and induces reactive species generation, we investigated whether the acute administration of creatine, an amino acid with antioxidant and ergogenic properties, protects against the seizures and neurochemical alterations (inhibition of Na(+),K(+)-ATPase and increased protein carbonylation) induced by the intrastriatal injection of GA (4 micromol/striatum). We also investigated whether creatine protected against the GA-induced inhibition of glutamate uptake in vitro. Creatine administration (300 mg/kg, p.o.) decreased seizures (evidenced by electrographic changes), protein carbonylation and Na(+),K(+)-ATPase inhibition induced by GA. However, creatine, at a dose capable of fully preventing GA-induced protein carbonylation (50 and 150 mg/kg, p.o.), did not prevent convulsions and Na(+),K(+)-ATPase inhibition, suggesting that the anticonvulsant activity of creatine in this experimental model is not related to its antioxidant action. Creatine also protected against the GA-induced inhibition of l-[(3)H]glutamate uptake in synaptosomes, suggesting that creatine may reduce the deleterious effects of GA by maintaining glutamate uptake in the synaptic cleft. Therefore, considering that creatine significantly attenuates the deleterious effects of GA assessed by behavioral and neurochemical measures, it is plausible to propose the use of this amino acid as an adjuvant therapy in the management of glutaric acidemia.