Cardiac myocyte hypertrophy and proliferating cell nuclear antigen expression in Wistar rats infected with Trypanosoma cruzi

Chagasic cardiomyopathy is a major life-threatening complication of Trypanosoma cruzi infection in human beings. This study focuses on the hypertrophic and hyperplastic mechanisms underlying the structural changes of the heart during experimental infection. Proliferating cell nuclear antigen (PCNA) expression, transversal diameter, nuclear area, and number of nuclei per unit volume were determined in the ventricular myocytes of T. cruzi-infected Wistar rats. PCNA expression was enhanced throughout the inflamed myocardium and in the spared areas of the left ventricular wall and the septum. Myocyte width increased from 26 to 75% at the inflammation-free myocardium (P < 0.0001), whereas it decreased 25% at the inflamed left ventricular wall areas (P < 0.001). Nuclear size increased in the inflammation-free myocardium of the left ventricle and the septum (> 10-36%, P < 0.01 and >0.2-32%, P < 0.03, respectively) and decreased at the inflamed areas of the left ventricular wall (10-22%. P < 0.02) with respect to the controls. The number of nuclei per unit volume decreased at the inflamed myocardium regardless of topographical location (36-65%) with respect to the controls (P < 0.0001) and in the inflammation-free myocardium of the right ventricle and the septum (<21-37%, P < 0.002 and <8-39%, P < 0.002, respectively). These results show that the heart responds to T. cruzi infection with DNA repair and cell multiplication in the inflamed sites and with hypertrophy of the unaffected myocardium.     

A comparative study between a brain Na+,K(+)-ATPase inhibitor (endobain E) and ascorbic acid

In the search of Na⁺,K⁺-ATPase modulators, we have reported the isolation by gel filtration and HPLC of a brain fraction, termed endobain E, which highly inhibits Na⁺,K⁺-ATPase activity. In the present study we compared some properties of endobain E with those of ascorbic acid. Kinetic experiments assaying synaptosomal membrane K⁺-p-nitrophenylphosphatase (K⁺-p-NPPase) activity in the presence of endobain E or ascorbic acid showed that in neither case did enzyme inhibition prove competitive in nature versus K⁺ or p-NPP concentration. At pH 5.0, endobain E and ascorbic acid maximal UV absorbance was 266 and 258 nm, respectively; alkalinization to pH 14.0 led to absorption drop and shift for endobain E but to absorbance disappearance for ascorbic acid. After cysteine treatment, endobain E absorbance decreased, whereas that of ascorbic acid remained unaltered; iodine treatment led to absorbance drop and shift for endobain E but to absorbance disappearance for ascorbic acid. HPLC analysis of endobain E disclosed the presence of two components: one eluting with retention time and UV spectrum indistinguishable from those of ascorbic acid and a second, as yet unidentified, both exerting Na⁺,K⁺-ATPase inhibition.     

K(+)-p-nitrophenylphosphatase inhibition by neurotensin involves high affinity neurotensin receptor: influence of potassium concentration and enzyme phosphorylation

Neurotensin (NT), a 13-amino acid peptide, is widely distributed in the brain and peripheral tissues of several mammalian species including man. In adult rat brain NT can bind to two distinct sites, one of high and the other of low affinity, corresponding to NT(1) and NT(2) receptor, respectively; structurally unrelated to these two, a third NT receptor (NT(3)) has been described. We have previously shown that Na(+), K(+)-ATPase is inhibited by NT when using ATP as substrate. In order to determine whether K(+)-stimulated dephosphorylation of this enzyme is involved, we tested NT effect by using p-nitrophenylphosphate, a non-natural substrate. K(+)-p-nitrophenylphosphatase activity was inhibited 42% by NT at 8.6 x 10(-6) M using an incubation medium containing 2 mM KCl but was unaffected in the presence of 5 or 20 mM KCl; however, with such KCl concentrations, NT was enabled to inhibit enzyme activity ( congruent with 35%) provided a suitable ATP:NaCl mixture (0.6:45.0 mM) was added. Mg(2+)-p-nitrophenylphosphatase activity remained unaltered at all conditions tested. Since SR 48692, a selective non-peptide NT(1) antagonist, abolished NT effect, involvement of NT(1) receptor in enzyme inhibition is suggested.     

The role of vocal individuality in conservation

Identifying the individuals within a population can generate information on life history parameters, generate input data for conservation models, and highlight behavioural traits that may affect management decisions and error or bias within census methods. Individual animals can be discriminated by features of their vocalisations. This vocal individuality can be utilised as an alternative marking technique in situations where the marks are difficult to detect or animals are sensitive to disturbance. Vocal individuality can also be used in cases were the capture and handling of an animal is either logistically or ethically problematic. Many studies have suggested that vocal individuality can be used to count and monitor populations over time; however, few have explicitly tested the method in this role. In this review we discuss methods for extracting individuality information from vocalisations and techniques for using this to count and monitor populations over time. We present case studies in birds where vocal individuality has been applied to conservation and we discuss its role in mammals.     

The Effect of Endogenous Modulator Endobain E on NMDA Receptor Is Interfered by Zn22+ but Is Independent of Modulation by Spermidine

A brain endogenous factor, termed endobain E, allosterically decreases [3H]dizocilpine binding to NMDA receptor. Such effect depends on receptor activation by the coagonists glutamate and glycine and is interfered by channel blockers, suggesting its interaction with the inner surface of the associated channel. To further analyze endobain E effect on NMDA receptor, in the current study competitive [3H]dizocilpine binding assays to brain membranes were performed with Zn2+ to block the associated channel, as well as with spermidine (SPD), which exerts positive allosteric modulation of NMDA receptor. Partially or nonadditive effects on [3H]dizocilpine binding were recorded, respectively, in the presence of endobain E at a concentration that inhibits binding 25% plus IC25 Zn2+ or endobain E at a concentration that inhibits binding 50% plus IC50 Zn2+. With an endobain E concentration that decreases 25% ligand binding, SPD potentiated binding over a wide concentration range but failed to modify endobain E effect. Similarly, [3H]dizocilpine binding reduction over a wide endobain E concentration range remained unaltered by high SPD concentrations. Additive effects were observed with endobain E at a concentration that decreases binding 25% plus IC25 SPD site antagonists arcaine or ifenprodil. Zn2+ experiments indicated that endobain E effect is interfered by channel blockade produced by this ion. Although endobain E effect is dependent on NMDA receptor activation by glutamate and glycine, it proves independent of the positive modulation exerted by SPD. Thus the endogenous modulator seems not to interact at NMDA receptor polyamine site, favoring the hypothesis that endobain E binds inside the associated channel.     

An Endogenous Na+, K+-ATPase Inhibitor Enhances Phosphoinositide Hydrolysis in Neonatal but Not in Adult Rat Brain Cortex

The effect of an endogenous Na⁺, K⁺-ATPase inhibitor, termed endobain E, on phosphoinositide hydrolysis was studied in rat brain cortical prisms and compared with that of ouabain. As already shown for ouabain, a transient effect was obtained with endobain E; maximal accumulation of inositol phosphates induced by endobain E was 604 ± 138% and 186 ± 48% of basal values in neonatal and adult rats, respectively. The concentration-response plot for the interaction between endobain E and phosphoinositide turnover differed from that of ouabain, thus suggesting the involvement of distinct mechanisms. In the presence of endobain E plus ouabain at saturating concentrations, no additive effect was recorded, suggesting that both substances share at least a common step in their activation mechanism of inositol phosphates metabolism or that they enhance phosphatidylinositol 4,5-biphosphate breakdown from the same membrane precursor pool, until its exhaustion. Experiments with benzamil, a potent blocker of Na⁺/Ca²⁺ exchanger, showed that it partially and dose-dependently inhibited endobain E effect. These results indicate that the endogenous Na⁺, K⁺-ATPase inhibitor endobain E, like ouabain, is able to stimulate phosphoinositide turnover transiently during postnatal brain development.     

Different properties of two brain extracts separated in sephadex G-50 that modify synaptosomal ATPase activities

We have previously reported that Na⁺,K⁺-ATPase of nerve ending membranes is stimulated by catecholamines only in the presence of a brain soluble fraction. The filtration of this soluble fraction through Sephadex G-50 permitted the separation of two extracts of maximal UV absorbance (peaks I and II) which showed different effects on ATPases. Peak I stimulated both Na⁺,K⁺-ATPase and Mg²⁺-ATPase activities and peak II inhibited Na⁺,K⁺-ATPase activity. We have now studied the activity of ATPases in the presence of the whole eluate obtained from the Sephadex G-50 column. It was observed that maximal effects on ATPases were obtained with peaks I and II. Peak I and peak II fractions were unable to modify the activity of acetylcholinesterase or 5-nucleotidase present in the synaptosomal membranes. The stimulatory effect of peak I on ATPases was concentration dependent (up to 1100), it was stable at different pHs and it was reverted by catecholamines. The inhibitory effect of peak II on Na⁺,K⁺-ATPase was concentration dependent (up to 150,000), it was stable only at acid pH, and it was partially reverted by catecholamines. These findings indicate that the factors responsible for the effects of peaks I and II have different properties and that their actions on ATPases show enzyme specifity.     

THE EFFECT OF THE CONVULSANT 3-MERCAPTOPROPIONIC ACID ON ENZYMES OF THE γ-AMINO-BUTYRATE SYSTEM IN THE RAT CEREBRAL CORTEX

—The activities of GABA enzymes in rat cerebral cortex were studied after the administration of the convulsant, 3-mercaptopropionic acid (MP). We found that MP markedly inhibited glutamate decarboxylase (EC 4.1.1.15) and activated GABA-aminotransferase (EC 2.6.1.c). The level of GABA appeared to be decreased during convulsions but thereafter returned to normal. The study of the subcellular distribution of GABA enzymes after the administration of MP indicated that the glutamate decarboxylase present in the nerve endings was not affected, while GABA aminotransferase in the mitochondria was activated to a similar extent to that observed in the original homogenate. These results together with the recovery of glutamate decarboxylase activity in cortical homogenates by dialysis suggested a reversible type of inhibition, whereas the effect on GABA-aminotransferase seemed to be more permanent.     

Regulation of (Na+, K+) adenosinetriphosphatase of nerve ending membranes

Norepinephrine added in vitro to nerve ending membranes from rat cerebral cortex stimulates the activity of (Na⁺, K⁺) adenosinetriphosphatase (ATPase) only in the presence of the soluble brain fraction. In its absence norepinephrine inhibits the enzyme. (Mg²⁺)ATPase also showed stimulation by norepinephrine in the presence of the soluble fraction, but of lesser magnitude. The activation of (Na⁺, K⁺)ATPase by norepinephrine is not reproduced by cyclic AMP and is not antagonized by either - or -adrenergic blocking agents. These results suggest that the stimulation caused by norepinephrine is a direct effect on the enzyme and is not mediated by cyclic AMP or adrenergic receptors.