Renata Franzon

Pretreatment with vitamins E and C prevent the impairment of memory caused by homocysteine administration in rats

Homocystinuria is a metabolic disorder caused by deficiency of cystathione β-synthase activity leading to tissue accumulation of homocysteine (Hcy); affected patients present neurological dysfunction. Considering that Hcy induces free radical formation and that memory is impaired by oxidative stress, in the present study we investigated the effect of an acute administration of Hcy on retrieval of step-down inhibitory avoidance in adult rats. The action of vitamins E and C on the effects produced by Hcy was also tested. Adult Wistar rats were pretreated for 1 week with daily i.p. administration of saline (control group) and vitamins E and C (vitamin E 40 mg/kg and vitamin C 100 mg/kg). Hcy (11 mmol/kg) or an equivalent volume of 0.9% saline were administered 1 h before training, 1 h before testing, or immediately after training sessions. Memory was significantly impaired in Hcy-treated group, whereas the rats chronically treated with vitamins E and C had this effect prevented. Present data strongly indicate that Hcy administration impairs memory, an effect probably mediated by oxidative stress since treatment with vitamins E and C prevented amnesia. Assuming the possibility that this might occur in the human condition, reported results may be relevant to explain, at least in part, neurologic dysfunction associated with homocystinuria.

Inhibition of rat brain Na+, K+-ATPase activity induced by homocysteine is probably mediated by oxidative stress.

The objective of the present study was to investigate the effects of preincubation of hippocampus homogenates in the presence of homocysteine or methionine on Na+, K+-ATPase and Mg2+-ATPase activities in synaptic membranes of rats. Homocysteine significantly inhibited Na+, K+-ATPase activity, whereas methionine had no effect. Mg2+-ATPase activity was not altered by the metabolites. We also evaluated the effect of incubating glutathione, cysteine, dithiothreitol, trolox, superoxide dismutase and GM1 ganglioside alone or incubation with homocysteine on Na+, K+-ATPase activity. Tested compounds did not alter Na+, K+-ATPase and Mg2+-ATPase activities, but except for trolox, prevented the inhibitory effect of homocysteine on Na+, K+-ATPase activity. These results suggest that inhibition of this enzyme activity by homocysteine is possibly mediated by free radicals and may contribute to the neurological dysfunction found in homocystinuric patients.

Na+,K+-ATPase activity is reduced in hippocampus of rats submitted to an experimental model of depression: Effect of chronic lithium treatment and possible involvement in learning deficits

This study was undertaken to verify the effects of chronic stress and lithium treatments on the hippocampal Na+,K(+)-ATPase activity of rats, as well as to investigate the effects of stress interruption and post-stress lithium treatment on this enzyme activity and on spatial memory. Two experiments were carried out; in the first experiment, adult male Wistar rats were divided into two groups: control and submitted to a chronic variate stress paradigm, and subdivided into treated or not with LiCl. After 40 days of treatment, rats were killed, and Na+,K(+)-ATPase activity was determined. In the second experiment, rats were stressed during 40 days, and their performance was evaluated in the Water Maze task. The stressed group was then subdivided into four groups, with continued or interrupted stress treatment and treated or not with lithium for 30 additional days. After a second evaluation of performance in the Water Maze, rats were killed and Na+,K(+)-ATPase activity was also measured. Results showed an impairment in Na+,K(+)-ATPase activity and in Water Maze performance of chronically stressed rats, which were prevented by lithium treatment and reversed by lithium treatment and by stress interruption. These results suggest that the modulation of Na+,K(+)-ATPase activity may be one of the mechanisms of action of lithium in the treatment of mood disorders.

Nitric Oxide Synthase Inhibition by L-NAME Prevents the Decrease of Na+,K+-ATPase Activity in Midbrain of Rats Subjected to Arginine Administration

In the present study we investigated the effect of acute administration of L-arginine on Na+,K+-ATPase and Mg2+-ATPase activities and on some parameters of oxidative stress (chemiluminescence and total radical-trapping antioxidant parameter-TRAP) in midbrain of adult rats. We also tested the effect of L-NAME on the effects produced by arginine. Sixty-day-old rats were treated with an acute intraperitoneal injection of saline (group I, control), arginine (0.8 g/kg) (group II), L-NAME (2 mg/kg) (group III) or arginine (0.8 g/kg) plus L-NAME (2 mg/kg) (group IV). Na+,K+-ATPase activity was significantly reduced in the arginine-treated rats, but was not affected by other treatments. In contrast, Mg2+-ATPase activity was not altered by any treatment. Furthermore, chemiluminescence was significantly increased and TRAP was significantly decreased in arginine-treated rats, whereas the simultaneous injection of L-NAME prevented these effects. These results demonstrate that in vivo arginine administration reduces Na+,K+-ATPase activity possibly through free radical generation induced by NO formation.

Evidence that oxidative stress is involved in the inhibitory effect of proline on Na+,K+-ATPase activity in synaptic plasma membrane of rat hippocampus

In the present study, we investigated the effect of Vitamins E and C on the inhibition of Na+,K+‐ATPase activity provoked by proline (Pro) administration in rat hippocampus. Five‐day‐old rats were pretreated for 1 week with daily i.p. administration of saline (control) or Vitamin E (40 mg/kg) and Vitamin C (100 mg/kg). Twelve hours after the last injection, animals received one single injection of Pro (12.8 μmol/g of body weight) or saline and were killed 1 h later. Results showed that Na+,K+‐ATPase activity was decreased in the Pro‐treated rats and that the pretreatment with Vitamins E and C prevented this effect. In another set of experiments, we investigated the in vitro effect of 1.0 mM Pro on Na+,K+‐ATPase activity from synaptic membranes of hippocampus of rats. Pro significantly inhibited (30%) Na+,K+‐ATPase activity. We also evaluated the effect of preincubating glutathione, trolox and Nϖ‐nitro‐l‐arginine methyl ester (l‐NAME) alone or combined with Pro on Na+,K+‐ATPase activity. Tested drugs did not alter Na+,K+‐ATPase activity, but glutathione prevented the inhibitory effect of Pro on this enzyme activity. These results suggest that the in vivo and in vitro inhibitory effect of Pro on Na+,K+‐ATPase activity is probably mediated by free radicals that may be involved in the neurological dysfunction found in hyperprolinemic patients.

Inhibition of Na+,K+-ATPase Activity from Rat Hippocampus by Proline

Na+,K+-ATPase and Mg2+-ATPase activities were determined in the synaptic plasma membranes from hippocampus of rats subjected to chronic and acute proline administration. Na+,K+-ATPase activity was significantly reduced in chronic and acute treatment by 33% and 40%, respectively. Mg2+-ATPase activity was not altered by any treatment. In another set of experiments, synaptic plasma membranes were prepared from hippocampus and incubated with proline or glutamate at final concentrations ranging from 0.2 to 2.0 mM. Na+,K+-ATPase, but not Mg2+-ATPase was inhibited (30%) by the two amino acids. In addition, competition between proline and glutamate for the enzyme activity was observed, suggesting a common binding site for these amino acids. Considering that Na+,K+-ATPase activity is critical for normal brain function, the results of the present study showing a marked inhibition of this enzyme by proline may be associated with the neurological dysfunction found in patients affected by type II hyperprolinemia.

Reduction of butyrylcholinesterase activity in rat serum subjected to hyperhomocysteinemia.

In the present study we investigate the effect of homocysteine (Hcy) administration, the main metabolite accumulating in homocystinuria, on butyrylcholinesterase (BuChE) activity in serum of rats. For the acute treatment, 29-day-old Wistar rats received one subcutaneous injection of Hcy (0.6 μmol/g) or saline (control) and were killed 1 h later. For the chronic treatment, Hcy was administered subcutaneously to rats from the 6th to the 28th day of life. Control rats received saline. The rats were killed 12 h after the last injection. In another set of experiments, rats were pretreated for one week with vitamins E and C or saline and 12 h after the last injection received one single injection of Hcy or saline, being killed 1 h later. Serum was used to determine BuChE activity. Our results showed that acute and chronic administration of Hcy significantly decreased BuChE activity. Furthermore, vitamins E and C per se did not alter BuChE activity, but prevented the reduction of this enzyme activity caused by acute administration of Hcy. The data suggest that the inhibitory effect of Hcy on BuChE activity is probably mediated by free radicals, since vitamins E and C administration prevented such effect.

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.

Outcomes of one-step incomplete and complete excavation in primary teeth: a 24-month randomized controlled trial.

Aim: To compare 24-month pulp health outcomes of partial caries removal (PCR) and total caries removal (TCR) with composite restoration in primary molars. Methods: 48 children aged 3-8 years with at least one molar with a deep carious lesion were included. 120 teeth were randomized to control (TCR; n = 54; 69% class II) and test (PCR; n = 66; 63% class II) groups. Total absence of carious tissue was confirmed using a blunt-tipped probe in the TCR group. For PCR, excavation was stopped when hardened, dried dentin with a leathery consistency was achieved. Pulpotomy was performed in cases of pulp exposure. Results: Pulp exposure occurred in 2 and 27.5% of teeth treated with PCR and TCR, respectively (p < 0.01). The operative time was significantly higher for TCR than PCR. Success rates were 92 and 96% in the PCR and TCR groups, respectively (p = 0.34). The success rate tended to be lower in occlusoproximal (92%) than in occlusal (100%) lesions (p = 0.08). Conclusion: The clinical and radiographic success rates of PCR and TCR in primary teeth with deep carious lesions were high and did not differ significantly, indicating that PCR is a reliable minimally invasive approach in primary teeth and that the retention of carious dentin does not interfere with pulp vitality. Moreover, PCR provided other clinically relevant advantages over TCR, especially lower incidence of pulp exposure and lower operative time.

Effects of electroconvulsive seizures on Na(+),K(+)-ATPase activity in the rat hippocampus.

Although several advances have occurred concerning the use of electroconvulsive therapy, little progress has been made in understanding the mechanisms underlying its therapeutic or side effects. Na(+),K(+)-ATPase is an important enzyme of central nervous system, responsible for ionic gradient maintenance and consumption of approximately 40-50% of brain ATP. This work was performed in order to determine Na(+),K(+)-ATPase activity after acute and chronic electroconvulsive shock. Results showed an inhibition of Na(+),K(+)-ATPase activity in the hippocampus 48 h, 7, 30, 60 and 90 days after a single electroconvulsive shock. Chronic treatment diminished the enzyme activity in the hippocampus 7 and 30 days after electroconvulsive (ECS) sessions. Our findings demonstrated that Na(+),K(+)-ATPase activity is altered by ECS.