Fabria Chiarani

Brain-derived neurotrophic factor serum levels before and after treatment for acute mania.

Accumulating evidence suggests that reduced levels of brain-derived neurotrophic factor (BDNF) in acute mood episodes may play an important role in the pathophysiology of bipolar disorder (BD). In order to assess changes in BDNF serum levels in BD patients before and after treatment for acute mania, ten bipolar patients were prospectively examined at inpatient unit admission and discharge. Diagnoses were made using the Structured Clinical Interview for DSM-IV, SCID-I. Serum BDNF levels were measured by sandwich ELISA. The results showed that BDNF levels were decreased in BD patients during mania when compared to controls (p=0.013) but this difference was no longer significant after treatment (p=0.126). A sharp increase in BDNF levels was found after treatment of the episode of acute mania (p=0.010). These findings suggest that the changes in BDNF serum levels may be associated with treatment response in acute mania. Further studies designed to validate the use of BDNF as a marker of treatment response in bipolar disorder are warranted.

Proline reduces acetylcholinesterase activity in cerebral cortex of rats.

In the present study we investigated the in vivo and in vitro effect of proline (Pro) on acetylcholinesterase (AChE) activity in rat cerebral cortex. The action of vitamins E and C on the effects produced by Pro was also tested. Twelve-day-old rats received one s.c. injection of Pro (12.8 μmol/g body weight) or an equivalent volume of 0.9% saline solution (control) and were killed 1 h later. In another set of experiments, 5-day-old rats were pretreated for 1 week with daily i.p. administration of saline (control) or vitamins E (40 mg/kg) and C (100 mg/kg). Twelve hours after the last injection the rats received one s.c. injection of Pro (12.8 μmol/g body weight) or saline (control) and were killed 1 h later. For the in vitro studies, cerebral cortex homogenates of 12-day-old untreated rats were incubated for 1 h with various concentrations of Pro (3.0 μM–1.0 mM) or with 1.0 mM Pro, 1.0 mM trolox, or 1.0 mM Pro plus 1.0 mM trolox. Controls did not contain Pro in the incubation medium. Our results showed that the AChE activity significantly decreased (25%) in rat brain subjected to Pro administration and that the pretreatment with vitamins E and C prevented this effect. Furthermore, Pro (0.5 and 1.0 mM) also inhibits AChE activity in vitro and trolox prevented this effect. The data suggest that the inhibitory effect of Pro on AChE activity is associated with oxidative stress. Although it is difficult to extrapolate our findings to the human condition, our results may be relevant to explain, at least in part, the neurologic dysfunction associated with hyperprolinemia type II.

Methionine alters Na+,K+-ATPase activity, lipid peroxidation and nonenzymatic antioxidant defenses in rat hippocampus

In the present study we investigated the effect of methionine exposure of hippocampus homogenates on Na+,K+‐ATPase activity from synaptic plasma membrane of rats. Results showed that methionine significantly decreased this enzyme activity. We also evaluated the effect of incubating glutathione (GSH) and trolox (α‐tocopherol) alone or combined with methionine on Na+,K+‐ATPase activity. The tested antioxidants per se did not alter the enzymatic activity, but prevented the inhibitory action of methionine on Na+,K+‐ATPase activity, indicating that Met inhibitory effect was probably mediated by free radical formation. Besides, we tested the in vitro effect of methionine on some parameters of oxidative stress, namely chemiluminescence, thiobarbituric acid reactive substances (TBARS), total radical‐trapping antioxidant potential (TRAP), as well as on the antioxidant enzyme activities catalase, glutathione peroxidase and superoxide dismutase in rat hippocampus. We observed that methionine significantly increased chemiluminescence and TBARS, decreased TRAP, but did not change the activity of the antioxidant enzymes. These findings suggest that reduction of Na+,K+‐ATPase activity and induction of oxidative stress may be involved in the brain damage observed in human hypermethioninemia.

Arginine Administration Decreases Cerebral Cortex Acetylcholinesterase and Serum Butyrylcholinesterase Probably by Oxidative Stress Induction

In the present study we investigated the action of vitamins E and C on the inhibition of acetylcholinesterase and butyrylcholinesterase activities provoked by arginine in cerebral cortex and serum of 60-day-old rats. Animals were pretreated for 1 week with daily intraperitoneal administration of saline (control) or vitamins E (40 mg/kg) and C (100 mg/kg). Twelve hours after the last injection, animals received one injection of arginine (0.8 μM/g of body weight) or saline. Results showed that acetylcholinesterase and butyrylcholinesterase activities were decreased in the arginine-treated rats. Furthermore, pretreatment with vitamins E and C prevented these effects. The data indicate that the reduction of acetylcholinesterase and butyrylcholinesterase activities caused by arginine was probably mediated by oxidative stress. Assuming the possibility that these effects might also occur in the human condition, our findings may be relevant to explain, at least in part, the neurological dysfunction associated with hyperargininemia and might support a novel therapeutic strategy to slow the progression of neurodegeneration in this disorder.

In vivo and in vitro effects of proline on some parameters of oxidative stress in rat brain.

Hyperprolinemia type II is an autosomal recessive disorder caused by the severe deficiency of delta(1)-pyrroline-5-carboxylate dehydrogenase activity leading to tissue accumulation of proline (Pro). Most patients detected so far show neurological manifestations including epilepsy and mental retardation, whose pathophysiology is not yet fully established. In the present study, we determined the in vivo and in vitro effects of Pro on some parameters of oxidative stress, namely chemiluminescence, total radical-trapping antioxidant potential (TRAP) and the activity of the antioxidant enzymes catalase (CAT), glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) from cerebral cortex of 29-day-old Wistar rats. Results showed that acute administration of Pro provoked a significant increase of chemiluminescence and a decrease of TRAP, whereas chronic administration of the metabolite did not alter these parameters. Furthermore, in vitro brain exposure to Pro resulted in increased chemiluminescence and decreased TRAP at Pro concentrations similar to those observed in tissues of hyperprolinemic patients (0.5-1.0 mM). As regards to the antioxidant enzymes, acute injection of Pro significantly decreased CAT activity and did not alter SOD and GSH-Px activities, whereas chronic Pro administration provoked a significant increase of CAT activity, a decrease of GSH-Px activity and did not modify SOD activity. Furthermore, CAT, GSH-Px and SOD activities were not affected by the presence of Pro in the incubation medium. The data indicate that Pro induces oxidative stress in vivo and in vitro, which may be involved in the brain dysfunction observed in hyperprolinemic patients.

Evaluation of the mechanism underlying the inhibitory effect of guanidinoacetate on brain Na+, K+-ATPase activity

Guanidinoacetate methyltransferase deficiency (GAMT‐deficiency) is an inherited neurometabolic disorder clinically characterized by epilepsy and mental retardation and biochemically by accumulation of guanidinoacetate (GAA) and depletion of creatine. Although the neurological symptoms are predominant, the pathogenesis of the brain dysfunction in this disorder is not yet established. In the present study we investigated the in vitro effect of GAA on Na+, K+‐ATPase and Mg2+‐ATPase activities in synaptic plasma membrane from hippocampus of young rats. Results showed that GAA significantly inhibited Na+, K+‐ATPase activity without affecting Mg2+‐ATPase activity. We also evaluated the effect of glutathione (GSH), trolox, Nϖ‐nitro‐l‐arginine methyl ester (L‐NAME) and taurine (Tau) on the inhibition elicited by GAA on Na+, K+‐ATPase activity. GSH, trolox, L‐NAME and Tau per se did not alter Na+, K+‐ATPase activity. However, L‐NAME and taurine prevented the inhibitory effect of GAA on this enzyme activity. Our findings suggest that the inhibition of Na+, K+‐ATPase activity caused by GAA is possibly mediated by nitric oxide (NO) formation and/or synaptic membrane alteration. The present data may contribute to the understanding of the neurological dysfunction characteristic of GAMT‐deficient patients.

Proline reduces brain cytochrome c oxidase : prevention by antioxidants

In the present study, we initially investigated the in vivo (acute and chronic) and in vitro effects of proline on cytochrome c oxidase (complex IV) activity in rat cerebral cortex to test the hypothesis that proline might alter energy metabolism and that this alteration could be provoked by oxidative stress. The action of α‐tocopherol and ascorbic acid on the effects produced by proline was also evaluated. For acute administration, 29‐ and 60‐day‐old rats received one subcutaneous injection of proline (18.2 μmol/g body weight) or an equivalent volume of 0.9% saline solution (control) and were sacrificed 1 h later. For chronic treatment, proline was injected subcutaneously twice a day at 10 h intervals from the 6th to the 28th day of age. Rats were sacrificed 12 h (29th) or 31 days (60th) after the last injection. Results showed that acute administration of proline significantly diminished the activity of cytochrome c oxidase in the cerebral cortex of 29‐ and 60‐day‐old rats. On the other hand, chronic hyperprolinemia reduced this complex activity only on day 29, but not on the 60th day of life. In another set of experiments, 22‐day‐old rats or 53‐day‐old rats were pretreated for 1 week with daily intraperitoneal administration of α‐tocopherol (40 mg/kg) and ascorbic acid (100 mg/kg) or saline. Twelve hours after the last antioxidant injection, rats received a single injection of proline or saline and were killed 1 h later. In parallel to chronic treatment, rats received a daily intraperitoneal injection of α‐tocopherol and ascorbic acid from the 6th to the 28th day of life and were killed 12 h after the last injection. Results showed that the pretreatment with α‐tocopherol and ascorbic acid before acute proline administration or concomitant to chronic proline administration significantly prevented these effects. We also observed that proline (3.0 μM–1.0 mM) when added to the incubation medium (in vitro studies) did not alter cytochrome c oxidase activity. Data suggest that the inhibitory effect of proline on cytochrome c oxidase activity is possibly associated with oxidative stress and that this parameter may be involved in the brain dysfunction observed in hyperprolinemia.

Effect of hyperprolinemia on acetylcholinesterase and butyrylcholinesterase activities in rat

Summary.We observed here that acute proline (Pro) administration provoked a decrease (32%) of acetylcholinesterase (AChE) activity in cerebral cortex and an increase (22%) of butyrylcholinesterase (BuChE) activity in the serum of 29-day-old rats. In contrast, chronic administration of Pro did not alter AChE or BuChE activities. Furthermore, pretreatment of rats with vitamins E and C combined or alone, Nϖ-nitro-L-arginine methyl ester or melatonin prevented the reduction of AChE activity caused by acute Pro administration, suggesting the participation of oxidative stress in such effects.

Protective effect of antioxidants on brain oxidative damage caused by proline administration

We have previously demonstrated that acute and chronic hyperprolinemia induce oxidative stress in cerebral cortex of rats. In the present study, we investigated the action of Vitamins E and C on the oxidative damage elicited by acute and chronic administration of proline (Pro) in rat cerebral cortex. Results showed that treatment with Vitamins E and C prevented the alterations caused by acute and chronic administration of proline on chemiluminescence, total radical-trapping antioxidant potential (TRAP) and on the activities of catalase and glutathione peroxidase. If these effects also occur in the human condition, it is possible that antioxidant administration might serve as a potential adjuvant therapy to avoid the progression of the neuropsychiatric dysfunction observed in hyperprolinemic patients.

Effect of hypoxanthine on Na+,K+-ATPase activity and some parameters of oxidative stress in rat striatum

The main objective of this study was to investigate the effects of preincubation of rat striatum homogenate in the presence of hypoxanthine, a metabolite accumulated in Lesch-Nyhan disease, on Na+,K+-ATPase activity and on some parameters of oxidative stress namely thiobarbituric acid-reactive substances (TBA-RS), total radical-trapping antioxidant parameter (TRAP) and membrane protein thiol content. Results showed that hypoxanthine significantly increased TBA-RS and reduced Na+,K+-ATPase activity, TRAP and membrane protein thiol content. In addition, we also evaluated the effect of glutathione, trolox, allopurinol and Nvarpi-nitro-L-arginine methyl ester (L-NAME) on the inhibitory effect of hypoxanthine on Na+,K+-ATPase activity in the same rat cerebral structure. All tested compounds per se did not alter Na+,K+-ATPase activity, but only glutathione and trolox prevented the effect of hypoxanthine on the enzyme activity. The effect of glutathione and trolox on hypoxanthine-induced increase of TBA-RS levels was also investigated. These antioxidants alone or combined with hypoxanthine reduced TBA-RS levels. Our present findings show that hypoxanthine induces oxidative stress in rat striatum and that the inhibition of Na+,K+-ATPase activity caused by this oxypurine was probably mediated by reactive oxygen species. It is presumed that these results might be associated with the neuronal dysfunction of patients affected by Lesch-Nyhan disease.