Gestational folic acid supplementation does not affects the maternal behavior and the early development of rats submitted to neonatal hypoxia-ischemia but the high supplementation impairs the dam's memory and the Na+, K+ - ATPase activity in the pup's hippocampus.

Folic acid (FA) is a B-complex vitamin important to the development of the fetus, being supplemented during pregnancy. Our recent findings showed that gestation supplementation (normal and excess doses) prevented the cognitive deficits and BDNF imbalance in adult rats that were submitted to neonatal hypoxia-ischemia (HI). To better understand this protective effect, the present study aimed to evaluate whether FA supplementation could be related to (1) maternal behavior, memory and Na+, K+ - ATPase activity in the hippocampus of the dams; (2) on somatic growth, early neurobehavioral development and Na+, K+ - ATPase activity in the hippocampus of the offspring; and (3) the effects of this supplementation in pups submitted to neonatal HI. Pregnant Wistar rats were divided into three groups, according to the diet they received during gestation: standard diet (SD), supplemented with 2 mg/kg of FA (FA2 - normal dose) and supplemented with 20 mg/kg of FA (FA20 -excessive dose). At the 7th PND pups were submitted to the Levine-Vannucci model of HI. During weaning the maternal behavior, the somatic growth and the neurobehavior development of pups were assessed. After weaning, the memory of the dams (by the Ox-maze task) and the Na+, K+ - ATPase activity in the hippocampus of both dams and offspring were evaluated. Considering the dams (1), both doses of FA did not alter the maternal behavior or the Na+, K+ - ATPase activity in the hippocampus, but a memory deficit was observed in the high FA-supplemented mothers. Considering the offspring (2), both FA doses did not affect the somatic growth or the neurobehavior development, but the FA20 pups had a decreased Na+, K+ - ATPase activity in the hippocampus. The FA supplementation did not change the parameters evaluated in the HI rats (3) and did not prevent the decreased Na+, K+ - ATPase activity in the hippocampus of the HI pups. These results indicate that normal FA supplementation dose does not influence the maternal behavior and memory and does not impact on the offspring early development in rats. Further studies are needed to confirm the effects of the high FA supplementation dose in the dams' memory and in the Na+, K+ - ATPase activity in the hippocampus of the offspring.

DNA damage in homocystinuria: 8-oxo-, 8-dihydro-2'-deoxyguanosine levels in cystathionine-ß-synthase deficient patients and the in vitro protective effect of N-acetyl­L­cysteine

Introduction: Homocysteine (Hcy) tissue accumulation occurs in a metabolic disease characterized biochemically by cystathionine ß-synthase (CBS) deficiency and clinically by mental retardation, vascular problems, and skeletal abnormalities. Previous studies indicate the occurrence of DNA damage secondary to hyperhomocysteinemia and it was observed that DNA damage occurs in leukocytes from CBS-deficient patients. This study aimed to investigate whether an oxidative mechanism could be involved in DNA damage previously found and investigated the in vitro effect of N-acety-L-cysteine (NAC) on DNA damage caused by high Hcy levels. Methods: We evaluated a biomarker of oxidative DNA damage in the urine of CBS­deficient patients, as well as the in vitro effect of NAC on DNA damage caused by high levels of Hcy. Moreover, a biomarker of lipid oxidative damage was also measured in urine of CBS deficient patients. Results: There was an increase in parameters of DNA (8-oxo-7,8-dihydro-2'- deoxyguanosine) and lipid (15-F2t-isoprostanes levels) oxidative damage in CBS-deficient patients when compared to controls. In addition, a significant positive correlation was found between 15-F2t-isoprostanes levels and total Hcy concentrations. Besides, an in vitro protective effect of NAC at concentrations of 1 and 5 mM was observed on DNA damage caused by Hcy 50 µM and 200 µM. Additionally, we showed a decrease in sulfhydryl content in plasma from CBS-deficient patients when compared to controls. Discussion: These results demonstrated that DNA damage occurs by an oxidative mechanism in CBS deficiency together with lipid oxidative damage, highlighting the NAC beneficial action upon DNA oxidative process, contributing with a new treatment perspective of the patients affected by classic homocystinuria.

Forced Treadmill Exercise Prevents Spatial Memory Deficits in Aged Rats Probably Through the Activation of Na+, K+-ATPase in the Hippocampus.

Regular physical activity has shown to improve the quality of life and to prevent age-related memory deficits. Memory processing requires proper regulation of several enzymes such as sodium-potassium adenosine triphosphatase (Na+, K+-ATPase) and acetylcholinesterase (AChE), which have a pivotal role in neuronal transmission. The present study investigated the effects of a treadmill running protocol in young (3 months), mature (6 months) and aged (22 months) Wistar rats, on: (a) cognitive function, as assessed in the Water maze spatial tasks; (b) Na+, K+-ATPase and AChE activities in the hippocampus following cognitive training alone or treadmill running combined with cognitive training. Animals of all ages were assigned to naïve (with no behavioral or exercise training), sedentary (non-exercised, with cognitive training) and exercised (20 min of daily running sessions, 3 times per week for 4 weeks and with cognitive training) groups. Cognition was assessed by reference and working memory tasks run in the Morris Water maze; 24 h after last session of behavioral testing, hippocampi were collected for biochemical analysis. Results demonstrated that: (a) a moderate treadmill running exercise prevented spatial learning and memory deficits in aged rats; (b) training in the Water maze increased both Na+, K+-ATPase and AChE activities in the hippocampus of mature and aged rats; (c) aged exercised rats displayed an even further increase of Na+, K+-ATPase activity in the hippocampus, (d) enzyme activity correlated with memory performance in aged rats. It is suggested that exercise prevents spatial memory deficits in aged rats probably through the activation of Na+, K+-ATPase in the hippocampus.

Guanidinoacetate alters antioxidant defenses and butyrylcholinesterase activity in the blood of rats

Deficiency of guanidinoacetate methyltransferase, the first described creatine biosynthesis defect, leads to depletion of creatine and phosphocreatine, and accumulation of guanidinoacetate (GAA) in brain and body fluids. The present study aimed to investigate the influence of GAA on the activities of antioxidant enzymes, as well as on thiobarbituric acid-reactive substances (TBARS) and butyrylcholinesterase (BuChE) activity in the blood of rats. We also evaluated the effect of trolox (6-hydr oxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), GSH (glutathione) and L-NAME (NG-nitro-L-arginine methyl ester) on the alterations elicited by GAA. Methods: The rats were randomly divided into 8 groups: (1) control; (2) GAA (10, 30, 50, 100 mM/kg); (3) trolox (1 mM/kg) + control; (4) trolox (1 mM/kg) + GAA (100 mM/kg); (5) GSH (1 mM/kg) + control; (6) GSH (1 mM/kg) + GAA (100 mM/kg); (7) L-NAME (1 mM/kg) + control; (8) L-NAME + GAA (100 mM/kg). After the addition of compounds, erythrocytes and plasma were pre-incubated at 37°C for 1h and tested immediately. Results: GAA enhanced the activities of catalase (CAT) and glutathione peroxidase (GSH-Px) in the erythrocytes and BuChE activity. In addition, GAA enhanced TBARS levels in the plasma. Trolox, GSH and L-NAME addition prevented the majority of alterations in oxidative stress parameters and the increase of BuChE activity that were caused by GAA. Data suggest that GAA alters antioxidant defenses and induces lipid peroxidation in the blood, as well altering BuChE activity. However, in the presence of trolox, GSH and L-NAME some of these alterations in oxidative stress and BuChE activity were prevented. Conclusions: Our findings lend support to a potential therapeutic strategy for this condition, which may include the use of appropriate antioxidants for ameliorating the damage caused by GAA...

Homocysteine contribution to DNA damage in cystathionine ß-synthase-deficient patients.

High blood levels of homocysteine (Hcy) are found in patients affected by homocystinuria, a genetic disorder caused by deficiency of cystathionine ß-synthase (CBS) activity, as well as in nutritional deficiencies (vitamin B12 or folate) and in abnormal renal function. We previously demonstrated that lipid and protein oxidative damage is increased and the antioxidant defenses diminished in plasma of CBS-deficient patients, indicating that oxidative stress is involved in the pathophysiology of this disease. In the present work, we extended these investigations by evaluating DNA damage through the comet assay in peripheral leukocytes from CBS-deficient patients, as well as by analyzing of the in vitro effect of Hcy on DNA damage in white blood cells. We verified that DNA damage was significantly higher in the CBS-deficient patients under treatment based on a protein-restricted diet and pyridoxine, folic acid, betaine and vitamin B12 supplementation, when compared to controls. Furthermore, the in vitro study showed a concentration-dependent effect of Hcy inducing DNA damage. Taken together, the present data indicate that DNA damage occurs in treated CBS-deficient patients, possibly due to high Hcy levels.

Homocysteine and other markers of cardiovascular risk during a manic episode in patients with bipolar disorder.

OBJECTIVE: To evaluate serum levels of different biomarkers associated with cardiovascular disease in patients with bipolar disorder (BD). Patients were prospectively evaluated in two separate instances: during acute mania and after remission of manic symptoms. All measurements were compared with those of healthy controls. METHODS: The study included 30 patients with BD and 30 healthy controls, matched for gender and age. Biochemical parameters evaluated included homocysteine (Hcy), folic acid, vitamin B12, ferritin, creatine kinase (CK) and C-reactive protein (CRP). RESULTS: Hcy levels were significantly higher in the BD patients, both during mania and after achieving euthymia. When Hcy was adjusted for body mass index, there was no significant difference between patients and controls. Ferritin was the only marker that showed a significant decrease during mania when compared to both euthymic patients and controls. There were no significant differences for folate, vitamin B12, CK and CRP. CONCLUSIONS: These findings do not show an association between alterations of markers of cardiovascular risk during manic episodes. Further studies are necessary to determine factors and mechanisms associated with cardiovascular risk in patients with BD.

Homocysteine and other markers of cardiovascular risk during a manic episode in patients with bipolar disorder

Objective: To evaluate serum levels of different biomarkers associated with cardiovascular disease in patients with bipolar disorder (BD). Patients were prospectively evaluated in two separate instances: during acute mania and after remission of manic symptoms. All measurements were compared with those of healthy controls. Methods: The study included 30 patients with BD and 30 healthy controls, matched for gender and age. Biochemical parameters evaluated included homocysteine (Hcy), folic acid, vitamin B12, ferritin, creatine kinase (CK) and C-reactive protein (CRP). Results: Hcy levels were significantly higher in the BD patients, both during mania and after achieving euthymia. When Hcy was adjusted for body mass index, there was no significant difference between patients and controls. Ferritin was the only marker that showed a significant decrease during mania when compared to both euthymic patients and controls. There were no significant differences for folate, vitamin B12, CK and CRP. Conclusions: These findings do not show an association between alterations of markers of cardiovascular risk during manic episodes. Further studies are necessary to determine factors and mechanisms associated with cardiovascular risk in patients with BD. .

Chronic methylphenidate administration alters antioxidant defenses and butyrylcholinesterase activity in blood of juvenile rats.

Methylphenidate (MPH), a psychostimulant that affects both dopaminergic and noradrenergic systems, is one of the most frequently prescribed treatments for attention-deficit hyperactivity disorder. The present study investigated the effects of chronic administration of MPH on some parameters of oxidative stress, as well as on butyrylcholinesterase (BuChE) activity in blood of young rats. Rats received intraperitoneal injections of MPH (2.0 mg/kg) once a day, from the 15th to the 45th day of age or an equivalent volume of 0.9% saline solution (controls). Two hours after the last injection, animals were euthanized, and blood was collected. Results demonstrated that MPH did not alter the dichlorofluorescein formed, decreased both thiobarbituric acid reactive substances and total non-enzymatic radical-trapping antioxidant, and increased superoxide dismutase and catalase activities, suggesting that this psychostimulant may alter antioxidant defenses. BuChE activity was increased in blood of juvenile rats subjected to chronic MPH administration. These findings suggest that MPH may promote peripheral oxidative adaptations and cholinergic changes.

Experimental evidence of oxidative stress in plasma of homocystinuric patients: a possible role for homocysteine.

Homocystinuria is an inherited disorder biochemically characterized by high urinary excretion of homocystine and increased levels of homocysteine (Hcy) and methionine in biological fluids. Affected patients usually have a variety of clinical and pathologic manifestations. Previous experimental data have shown a relationship between Hcy and oxidative stress, although very little was reported on this process in patients with homocystinuria. Therefore, in the present study we evaluated parameters of oxidative stress, namely carbonyl formation, malondialdehyde (MDA) levels, sulfhydryl content and total antioxidant status (TAS) in patients with homocystinuria at diagnosis and under treatment with a protein restricted diet supplemented by pyridoxine, folate, betaine, and vitamin B(12). We also correlated plasma Hcy and methionine concentrations with the oxidative stress parameters examined. We found a significant increase of MDA levels and carbonyl formation, as well as a reduction of sulfhydryl groups and TAS in plasma of homocystinuric patients at diagnosis relatively to healthy individuals (controls). We also verified that Hcy levels were negatively correlated with sulfhydryl content and positively with MDA levels. Furthermore, patients under treatment presented a significant reduction of the content of MDA, Hcy and methionine concentrations relatively to patients at diagnosis. Taken together, the present data indicate that lipid and protein oxidative damages are increased and the antioxidant defenses diminished in plasma of homocystinuric patients, probably due to increased reactive species elicited by Hcy. It is therefore presumed that oxidative stress participates at least in part in the pathogenesis of homocystinuria.

Homocysteine induces cytoskeletal remodeling and production of reactive oxygen species in cultured cortical astrocytes.

Homocysteine (Hcy) is an excitatory amino acid which markedly enhances the vulnerability of neuronal cells to excitotoxicity and oxidative injury. Patients with severe hyperhomocysteinemia exhibit a wide range of clinical manifestations including neurological abnormalities such as mental retardation, cerebral atrophy, and seizures. In this study we treated cortical astrocytes and neurons in culture with 10 and 100 µM Hcy and after 24h exposure cytoskeletal remodeling was analyzed by immunocytochemistry. We observed dramatically altered actin cytoskeleton in astrocytes exposed to 100 µM Hcy, with concomitant change of morphology to fusiform and/or flattened cells with retracted cytoplasm. Moreover, we observed disruption of the glial fibrillary acidic protein (GFAP) meshwork, supporting misregulation of actin cytoskeleton. Induction of reactive oxygen species (ROS) in astrocytes showed fluctuating levels along 24h exposure to both Hcy concentrations. Actin remodeling induced by 100 µM Hcy was prevented by the antioxidants folate (5 µM) or trolox (80 µM). Unlike astrocyte cytoskeleton, results evidence little susceptibility of neuron cytoskeleton until 24h of treatment, since immunocytochemical analysis showed that 10 and 100 µM Hcy-treated neurons presented unaltered neurite arborization. Moreover, alterations in astrocyte and neuron viability were not observed along the 24h of exposure to Hcy. Neuron/astrocyte co-cultures evidence an anchorage dependence for neuronal survival over long exposure to Hcy. Taken together, these findings indicate, that the cytoskeleton of cortical astrocytes, but not of neurons in culture, is a target to Hcy and such effects are mediated by redox signaling. Astrocytes were able to respond to Hcy (100 µM) reorganizing their cytoskeleton, surviving, and protecting neurons from Hcy damage. Moreover our results suggest a protective role for astrocytes remodeling the cytoskelon, and probably generating signals that would assure neuronal survival in response to the damage induced by Hcy.

Hyperhomocysteinemia selectively alters expression and stoichiometry of intermediate filament and induces glutamate- and calcium-mediated mechanisms in rat brain during development.

The aim of the present work was to investigate the actions of a chemically induced chronic hyperhomocysteinemia model on intermediate filaments (IFs) of cortical and hippocampal neural cells and explore signaling mechanisms underlying such effects. Results showed that in hyperhomocysteinemic rats the expression of neural IF subunits was affected. In cerebral cortex, glial fibrillary acidic protein (GFAP) expression was donwregulated while in hippocampus high and middle molecular weight neurofilament subunits (NF-H and NF-M, respectively) were up-regulated. Otherwise, the immunocontent of IF proteins was unaltered in cerebral cortex while in hippocampus the immunocontent of cytoskeletal-associated low molecular weight neurofilament (NF-L) and NF-H subunits suggested a stoichiometric ratio consistent with a decreased amount of core filaments enriched in lateral projections. These effects were not accompanied by an alteration in IF phosphorylation. In vitro results showed that 500muM Hcy-induced protein phosphatases 1-, 2A- and 2B-mediated hypophosphorylation of NF subunits and GFAP in hippocampal slices of 17-day-old rats without affecting the cerebral cortex, showing a window of vulnerability of cytoskeleton in developing hippocampus. Ionotropic and metabotropic glutamate receptors were involved in this action, as well as Ca(2+) release from intracellular stores through ryanodine receptors. We propose that the mechanisms observed in the hippocampus of 17-day-old rats could support the neural damage observed in these animals.

Homocysteine decreases extracellular nucleotide hydrolysis in rat platelets.

Hyperhomocysteinemia is an independent risk factor for atherothrombotic disease. Platelets play an important role in cardiovascular disease and release pro-aggregates mediators when activated, such as ADP, a physiological agonist involved in normal hemostasis and thrombosis. NTPDases and 5'-nucleotidase are ecto-enzymes that hydrolyze ATP, ADP and AMP to adenosine playing an important role on blood flow and thrombogenesis by regulating ADP catabolism. The aim of the present study was evaluate extracellular adenine nucleotide hydrolysis of rat platelets exposed to homocysteine in vitro and in vivo. In vitro homocysteine (Hcy) in the concentration range of 20 to 500 microM caused a significant decrease on ATP (around 30%) and ADP (around 45%) hydrolysis, respectively, while AMP hydrolysis was not altered. Hcy was not able to inhibit the hydrolysis of ATP and ADP catalyzed by purified apyrase at the same concentrations tested in vitro on platelets, suggesting an indirect effect. The inhibitory effect of Hcy on platelets was prevented by antioxidants agents in vitro and in vivo. Furthermore homocysteine treatment increased platelet aggregation induced by ADP. Based on the results presented herein, we propose that inhibition of extracellular ATP and ADP hydrolysis caused by homocysteine was probably due oxidative stress, since antioxidants prevented such effects. These findings may contribute to an increase platelet response to ADP and consequence development of thrombotic risk attributed to hyperhomocysteinemia.

Exercise effects on activities of Na(+),K(+)-ATPase, acetylcholinesterase and adenine nucleotides hydrolysis in ovariectomized rats.

Hormone deficiency following ovariectomy causes activation of Na(+),K(+)-ATPase and acetylcholinesterase (AChE) that has been related to cognitive deficits in experimental animals. Considering that physical exercise presents neuroprotector effects, we decide to investigate whether exercise training would affect enzyme activation in hippocampus and cerebral cortex, as well as adenosine nucleotide hydrolysis in synaptosomes from cerebral cortex of ovariectomized rats. Female adult Wistar rats were assigned to one of the following groups: sham (submitted to surgery without removal of the ovaries), exercise, ovariectomized (Ovx) and Ovx plus exercise. Thirty days after surgery, animals were submitted to one month of exercise training, three times per week. After, rats were euthanized, blood serum was collected and hippocampus and cerebral cortex were dissected. Data demonstrated that exercise reversed the activation of Na(+),K(+)-ATPase and AChE activities both in hippocampus and cerebral cortex of ovariectomized rats. Ovariectomy decreased AMP hydrolysis in cerebral cortex and did not alter adenine nucleotides hydrolysis in blood serum. Exercise per se decreased ADP and AMP hydrolysis in cerebral cortex. On the other hand, AMP hydrolysis in blood serum was increased by exercise in ovariectomized adult rats. Present data support that physical exercise might have beneficial effects and constitute a therapeutic alternative to hormone replacement therapy for estrogen deprivation.

Effects of cysteamine on oxidative status in cerebral cortex of rats.

Cystinosis is a systemic genetic disease caused by a lysosomal transport deficiency accumulating cystine in most tissues. Tissue damage depends on cystine accumulation, but the mechanisms of this damage are still obscure. Cysteamine administration depletes cystine accumulated, increasing survive of affected patients. Studies performed in fibroblasts of cystinotic patients suggest that apoptosis is enhanced in this disease. Considering that oxidative stress is a known apoptosis inducer, our main objective was to investigate a possible antioxidant effect of cysteamine on several parameters of oxidative stress in the brain of young rats. Animals received three subcutaneous injections at 3-h intervals of a buffered solution (pH 7.4) of 10 mg/kg body weight cysteamine and were sacrificed 1 h after the last injection. Cysteamine decreased lipoperoxidation and glutathione peroxidase activity, and increased the carbonyl content of proteins and catalase activity. In vitro studies showed that cysteamine reduced lipoperoxidation, 2',7'-dihydrodichlorofluorescein oxidation, carbonyl content of proteins and catalase activity, and increased glutathione peroxidase activity. These results suggest that cysteamine may act as a scavenger of superoxide free radicals and hydrogen peroxide. Therefore, it is possible that cysteamine may extend life of cystinotic patients acting not only as a cystine depleting drug, but also as a free radical scavenger, reducing cell damage by apoptosis.

Effects of an acute treatment with L-thyroxine on memory, habituation, danger avoidance, and on Na+, K+ -ATPase activity in rat brain.

Thyroid hormones (THs) have a relevant action on brain development and maintenance. By using an acute treatment to induce a hyperthyroid animal model, we aimed at investigating the effect of an altered THs levels on learning and memory and on the activity of Na(+), K(+)-ATPase in the rat brain. Our results have shown that the acute treatment with L-T4 did not alter the retrieval of the inhibitory avoidance task, but had a significant effect on the elevated plus maze and on open-field performance in rats. We suggest that animals subjected to L-T4 administration improved the habituation to a novel environment as well as a better evaluation of a dangerous environment, respectively. Na(+), K(+)-ATPase activity is increased in parietal cortex (30%), but it is not altered in hippocampus in L-T4 treated group. These both brain structures are involved in memory processing and it was previously demonstrated that there is a double dissociation between them for spatial location information, perceptual and episodic memory. We propose the hypothesis that this increase of Na(+), K(+)-ATPase activity in parietal cortex may be correlated to our results in behavior tests, which suggest a role of THs as well as of the Na(+), K(+)-ATPase in the cognitive process.

Intrastriatal hypoxanthine reduces Na(+),K (+)-ATPase activity and induces oxidative stress in the rats.

The main objective of this study was to investigate the effects of a single intrastriatal injection of hypoxanthine, a metabolite accumulated in Lesch Nyhan disease and possibly involved in its neuropathology, on Na(+),K(+)-ATPase activity, as well as on some parameters of oxidative stress, namely chemiluminescence (an index of lipid peroxidation), total radical-trapping antioxidant parameter--TRAP (an index of total antioxidant capacity of the tissue) and total thiol protein membrane content, in striatum, cerebral cortex and hippocampus of rats. Results show that hypoxanthine significantly decreased Na(+),K(+)-ATPase activity and TRAP while increased chemiluminescence in all ipsislateral structures tested. However, no effect on total thiol protein membrane content was detected. We suggest that hypoxanthine induces oxidative stress in all cerebral structures studied (striatum, hippocampus and cerebral cortex) and that the reduction of Na(+),K(+)-ATPase activity was probably mediated by reactive oxygen species.

Intrastriatal injection of hypoxanthine reduces striatal serotonin content and impairs spatial memory performance in rats.

The aim of this study was to investigate the effects of intrastriatal injection of hypoxanthine, a metabolite accumulated in Lesch-Nyhan disease, on rats' performance in the Morris water maze tasks, along with the monoamine content in striatum of rats. Male adult Wistar rats were divided in two groups: (1) saline-injected and (2) hypoxanthine-injected group. Seven days after solutions infusion, animals were trained in the Morris Water Maze or were sacrificed for evaluation of the striatal monoamine content. Results show that hypoxanthine administration caused impairment on spatial navigation in the acquisition phase in reference memory task in the Morris Water Maze, as well as in the latency to cross over the platform location in probe trial, when compared to the saline group (control). Hypoxanthine also altered rat performance in the working memory. Although striatal dopamine metabolites content did not change, treated animals showed a reduction of tissue levels of serotonin (5-HT) and 5- hydroxyl-indoleacetic acid (5-HIAA). These results show that intra-striatal hypoxanthine administration provoked impairment of spatial learning/memory in rats without affecting striatal dopaminergic system, although serotonergic pathways seem to have been affected.

Alpha-tocopherol and ascorbic acid administration prevents the impairment of brain energy metabolism of hyperargininemic rats.

1. We have previously demonstrated that arginine administration induces oxidative stress and compromises energy metabolism in rat hippocampus. In the present study we initially investigated the influence of pretreatment with alpha-tocopherol and ascorbic acid on the effects produced by arginine on hippocampus energy metabolism. We also tested the effect of acute administration of arginine on various parameters of energy metabolism, namely glucose uptake, lactate release and on the activities of succinate dehydrogenase, complex II and cytochrome c oxidase in rat cerebellum, as well as the influence of pretreatment with alpha-tocopherol and ascorbic acid on the effects elicited by arginine on this structure. 2. Sixty-day-old female Wistar rats were treated with a single i.p. injection of saline (control) or arginine (0.8 g/kg) and were killed 1 h later. In another set of experiments, the animals were pretreated for 1 week with daily i.p. administration of saline (control) or alpha-tocopherol (40 mg/kg) and ascorbic acid (100 mg/kg). Twelve hours after the last injection of the antioxidants the rats received one i.p. injection of arginine (0.8 g/kg) or saline and were killed 1 h later. 3. Results showed that arginine administration significantly increased lactate release and diminished glucose uptake and the activities of succinate dehydrogenase and complex II in rat cerebellum. In contrast, complex IV (cytochrome c oxidase) activity was not changed by this amino acid. Furthermore, pretreatment with alpha-tocopherol and ascorbic acid prevented the impairment of energy metabolism caused by hyperargininemia in cerebellum and hippocampus of rats.

The effects of the interactions between amino acids on pyruvate kinase activity from the brain cortex of young rats.

Considering that pyruvate kinase activity, a crucial enzyme for glucose metabolism and energy liberation in brain, may be regulated by some amino acids, it is possible that diminution of this enzyme activity may contribute to the brain damage caused by amino acids accumulated in metabolic diseases, such as phenylalanine, tryptophan and cystine. Therefore, the present study was undertaken to investigate the effect of these amino acids on pyruvate kinase activity in the brain cortex of rats. We also investigated the effect of serine and alanine on pyruvate kinase activity in the same tissue. The results suggested that phenylalanine, tryptophan, cystine, alanine, and serine act at the same site on the enzyme, phenylalanine, tryptophan, and cystine causing inhibition, and alanine and serine preventing this effect. Cystine also inhibited the enzyme activity through a different mechanism, possibly acting on the enzyme thiol groups. Considering that this enzyme is a target for amino acids accumulated in some metabolic diseases of amino acid metabolism, it is possible that its inhibition may contribute to the brain damage found in these diseases.

Chronic hyperprolinemia provokes a memory deficit in the Morris water maze task.

In the present study we investigated the effect of chronic proline (Pro) administration on rat performance in the Morris water maze task. Rats received s.c. injections of Pro twice a day at 8 h intervals from the 6th to the 28th days of age and equivalent volume of 0.9% saline solution (control). On the 60th day of life, rats were subjected to the water maze task. Results showed that chronic Pro administration provokes impairment on spatial learning, as shown by the increase of latency in acquisition and retention and by a reduced efficiency to find the platform position in the working memory test. Present results suggest that hyperprolininemia causes cognitive dysfunction and might be relevant to explain, at least in part, the neurological dysfunction associated with hyperprolinemia.