Acute morphological and physiological effects of lead in the neotropical fish Prochilodus lineatus

O presente estudo investigou os efeitos do chumbo na morfologia branquial, nos hematócritos e nas concentrações plasmáticas de sódio, glicose, lipídeos, proteínas e colesterol de Prochilodus lineatus exposto a duas concentrações subletais de chumbo durante 96 h. Inicialmente, testes agudos (96 h) e estáticos determinaram a CL50 (96 h) de chumbo para P. lineatus em 95 mg Pb.L-1. As concentrações de chumbo utilizadas nos testes subletais foram 24 e 71 mg Pb.L-1, que correspondem a 25% e 75%, respectivamente, da CL50 (96 h). As brânquias de P. lineatus expostos a ambas as concentrações de chumbo apresentaram maior incidência de lesões histopatológicas, como elevação epitelial, hiperplasia e aneurisma lamelar. P. lineatus não apresentou alterações significativas no hematócrito durante a exposição a ambas as concentrações de chumbo. Peixes expostos a 71 mg Pb.L-1 apresentaram decréscimo significativo do Na+ plasmático após 48 h, o que pode estar se refletindo na redução das taxas de influxo desse íon. P. lineatus expostos a ambas as concentrações de chumbo apresentaram resposta clássica ao estresse, como verificado pela hiperglicemia associada ao decréscimo dos lipídeos e proteínas plasmáticas. A magnitude da resposta ao estresse foi dose-dependente. A resposta apresentada na concentração mais baixa representa um processo adaptativo, enquanto na maior concentração caracteriza a exaustão.

Acute morphological and physiological effects of lead in the neotropical fish Prochilodus lineatus.

The present study investigated lead effects on gill morphology, hematocrit, blood sodium, glucose, lipids, protein, and cholesterol of Prochilodus lineatus exposed to two sublethal lead concentrations for 96 h. Preliminary series of short-term static toxicity tests were run to determine LC50 (96 h) of lead in P. lineatus, which was 95 mg Pb.L-1. Therefore, lead concentrations tested in the sublethal experiments were 24 and 71 mg Pb.L-1, which correspond to 25% and 75% of the LC50 (96 h), respectively. Gills of P. lineatus exposed to both lead concentrations during 96 h presented a higher occurrence of histopathological lesions such as epithelial lifting, hyperplasia, and lamellar aneurism. P. lineatus did not show significant alterations in hematocrit during exposure to both lead concentrations. Fish exposed to the highest lead concentration showed a significant decrease in Na+ plasma concentration after 48 h, possibly reflecting a sodium influx rate decrease. P. lineatus exposed to both lead concentrations presented a "classical general adaptation syndrome to stress", as hyperglycemia associated with lowered lipids and proteins was reported. Stress-response magnitude was dose-dependent. While the response to the lowest lead concentration might represent adaptation, the highest concentration seems to characterize exhaustion.

Nutrition and brain function: a multidisciplinary virtual symposium.

A few months ago, the Brazilian Society for Neuroscience and Behavior (SBNeC) promoted a "virtual symposium" (by Internet, under the coordination of R.C.A. Guedes) on "Nutrition and Brain Function". The discussions generated during that symposium originated the present text, which analyzes current topics on the theme, based on the multidisciplinary experience of the authors. The way the brain could be non-homogeneously affected by nutritional alterations, as well as questions like early malnutrition and the development of late obesity and hormone abnormalities were discussed. Also, topics like the role of essential fatty acids (EFAs) on brain development, increased seizure susceptibility and changes in different neurotransmitters and in cognitive performance in malnourished animals, as well as differences between overall changes in nutrient intake and excess or deficiency of specific nutrients (e.g. iodine deficiency) were analyzed. It was pointed out that different types of neurons, possibly in distinct brain structures, might be differently affected by nutritional manipulation, including not only lack-but also excess of nutrient intake. Such differences could help in explaining discrepancies between data on humans and in animals and so, could aid in determining the basic mechanisms underlying lesions or changes in brain function and behavior.

Determination of total proteins: a study of reaction between quinones and proteins.

A previous study was undertaken to test the reaction of several quinones (p-benzoquinone; 2,5-dichloro and 2,6-dichloro p-benzoquinone; tetrachloro-p-benzoquinone; tetrachloro-o-benzoquinone; 2,5-dichloro-3,6-dihydroxy-p-benzoquinone; benz[a]anthracene-7,12-dione) with bovine serum albumin (BSA). From this study, we have devised a spectrophotometric method for determination of total proteins. The quinone, tetrachloro-p-benzoquinone (p-chloranil), showed the best result. The product of reaction between proteins and p-chloranil absorbed at 360 nm and Beer's law was followed up to 200 mug ml(-1) of BSA. The product of reaction of BSA/p-chloranil was stable for 30 min, after that the absorbance increased 16% and kept stable for 24 h. The p-chloranil method showed a limit of detection (1.25 mug ml(-1)) lower than the biuret method (52.0 mug ml(-1)) or p-benzoquinone (PBQ) method (2.6-4.0 mug ml(-1)). The method was applied to spectrophotometric determination of total proteins in blood plasma; the results were compared with the biuret method that is widely used in clinical analysis.

Spectrophotometric determination of cysteine and/or carbocysteine in a mixture of amino acids, shampoo, and pharmaceutical products using p-benzoquinone.

A simple, sensitive, and selective method has been developed for determination of cysteine (Cys) or carbocysteine (carboCys) in pharmaceutical products, shampoos and a mixture of amino acids. The results showed the reaction between p-benzoquinone (PBQ) and Cys occurs through the sulfhydryl group. Previous derivatization or extraction is not necessary before the assay is carried out. The method is based on the fact that the product of reaction between PBQ and Cys absorbs at 352 and 500 nm or PBQ and carboCys absorbs at 500 nm. Beer's law is followed in the range 0-40 mug/ml for Cys and 0-150 mug/ml for carboCys. The product of reaction PBQ-Cys is stable for 2 h with absorption bands at 352 and 500 nm. In the presence of amino acids, PBQ is highly selective to Cys. Several substances such as amino acids, urea, salts, and dipeptide did not interfere with the proposed method. A recovery of about 100% is observed for both Cys and carboCys, when the method is applied to determine Cys in a mixture of amino acids resembling blood plasma, shampoo, and pill food as well as carboCys in pharmaceutical products.

Effect of chemical stimulation of the dorsomedial hypothalamic nucleus on blood plasma glucose, triglycerides and free fatty acids in rats.

The effects of chemical stimulation of the dorsomedial hypothalamic nucleus (DMH) on blood plasma concentration of glucose, triglycerides, insulin, and free fatty acids (FFA) were investigated in anesthetized adult Wistar rats. Microinjection of 12.5 nmol of norepinephrine into the DMH increased blood plasma concentration of glucose and FFA, decreased triglycerides, and did not change plasma insulin within 5 min; after 20 min, blood glucose and FFA reached control values. Microinjection of epinephrine (12.5 nmol) into the DMH also increased blood plasma glucose concentration and decreased triglycerides after 5 min. These effects are probably mediated by beta-adrenergic mechanisms, because they were prevented by beta-adrenergic antagonist propranolol, but not by alpha-adrenergic antagonist prazosin. Microinjection into the DMH of glutamate, dopamine, or acetylcholine failed to cause any change in those metabolic parameters, corroborating the hypothesis that the DMH is part of a beta-adrenergic pathway involved in short-term modulation of the availability of glucose and FFA.

Inhibition of protein phosphatase 1 stimulates secretion of Alzheimer amyloid precursor protein.

BACKGROUND: Aberrant metabolism of the Alzheimer amyloid precursor protein (APP) or its amyloidogenic A beta fragment is thought to be centrally involved in Alzheimer's disease. Nonamyloidogenic processing of APP involves its cleavage within the A beta domain by a protease, termed alpha-secretase, and release of the large extracellular domain, termed APPS. Secretion of APPS can be stimulated by phorbol esters, activators of protein kinase C, with concurrent inhibition of A beta production. While the role of protein kinases of APP metabolism has been investigated, considerably less effort has been devoted to elucidating the role played by protein phosphatases. Okadaic acid, a protein phosphatase inhibitor, has been shown to stimulate secretion of APPS, but the identity of the phosphatase involved has not been investigated. MATERIALS AND METHODS: The secretion of APPS from COS-1 cells was measured in the absence or presence of various doses of serine/threonine-specific phosphatase inhibitors. Quantitation of the derived IC50 values was used to determine the identity of the phosphatase involved in the control of APP secretion. RESULTS: The availability of protein phosphatase inhibitors with different relative potencies against the different types of serine/threonine-specific protein phosphatase allowed us to examine which of the four known types of protein phosphatase might be involved in the regulation of APP secretion. Both okadaic acid and calyculin A stimulated the secretion of APP from COS-1 cells in a dose-dependent manner. The half-maximal dose for stimulation of APP secretion was approximately 100-fold higher with okadaic acid than with calyculin A. CONCLUSIONS: The nearly 100-fold difference in the observed IC50 values for okadaic acid and calyculin A implicates a type 1 protein phosphatase in the control of APPS production. Protein phosphatase 1 (PP1) is known to be highly expressed in adult mammalian brain, both in neurons and glia. The identification of a specific phosphatase type in the control of APP secretion opens new avenues to the development of rational therapeutic intervention strategies aimed at the prevention and/or treatment of Alzheimer's Disease.

Intracerebroventricular injection of dibutyryl cyclic adenosine 3',5'-monophosphate increases hypothalamic levels of neuropeptide Y.

This investigation examined in vivo the relationship between the nucleotide cAMP and hypothalamic levels of two peptides, neuropeptide Y (NPY) and galanin (GAL), which are known to potentiate feeding behavior. In brain-cannulated rats, third ventricular injections of N6,2'-O-dibutyryl cyclic adenosine 3',5'-monophosphate ((Bu)2cAMP, 25 micrograms), compared to saline, caused a significant increase in NPY levels in the arcuate nucleus (ARC) and medial parvocellular portion of the paraventricular nucleus (mPVN), while having no impact in other hypothalamic areas. These site-specific changes in NPY occurred in the absence of any alteration in circulating levels of insulin, corticosterone, aldosterone or glucose, or of changes in hypothalamic levels of GAL. These findings implicate cAMP as having regulatory functions within specific hypothalamic NPY-synthesizing neurons, projecting from the ARC to the mPVN, that are believed to be involved in energy homeostasis.

Diurnal rhythm of galanin-like immunoreactivity in the paraventricular and suprachiasmatic nuclei and other hypothalamic areas.

The peptide galanin (GAL), when injected into the rat hypothalamus, is known to stimulate feeding behavior and affect the secretion of various hormones, including insulin and the adrenal steroid, corticosterone. To determine whether endogenous peptide levels shift in relation to natural rhythms of feeding and circulating hormone levels, rats were sacrificed at different times of the light/dark cycle, and their GAL levels were measured, via radioimmunoassay, in medial hypothalamic dissections and micropunched hypothalamic areas. The results suggest the existence of two distinct diurnal rhythms for hypothalamic GAL. One rhythm, detected exclusively in the area of the SCN, is characterized by bimodal peaks of GAL, threefold higher than basal peptide levels, around the onset of the dark and light periods. The second rhythm shows a single peak of GAL towards the middle of the nocturnal feeding cycle, specifically between the third and sixth hour. This latter rhythm is evident in the dorsal region of the medial hypothalamus, localized specifically to the lateral portion of the PVN. Moreover, it is inversely related to circulating insulin but unrelated to the adrenal steroids, suggesting a possible association between this pancreatic hormone and GAL in the PVN.

Neuropeptide Y in the arcuate nucleus is modulated by alterations in glucose utilization.

This study examined the response of hypothalamic neuropeptide Y (NPY) to specific metabolic challenges. After intraperitoneal administration of 2-deoxy-D-glucose, which blocks glucose utilization, NPY levels measured via radioimmunoassay were significantly potentiated in the arcuate (ARC) and suprachiasmatic nuclei of the rat hypothalamus. The antimetabolite mercaptoacetate, in contrast, which blocks fatty acid oxidation, produced no significant change and actually tended to reduce NPY levels in the ARC. It is concluded that glucose utilization, in particular, may constitute an important signal, either direct or indirect, in the modulation of NPY production in the hypothalamus.

Spectrophotometric determination of total proteins in blood plasma with p-benzoquinone.

1. In the present study we have documented the use of the reagent, p-benzoquinone (PBQ) for the spectrophotometric determination of total proteins in blood plasma. 2. Since the products of reaction are stable for several hours at room temperature after the 20-min boiling step, the time at which absorbance is measured is not a critical factor. 3. Common anticoagulants such as EDTA, citrate, or heparin do not interfere with the PBQ method at concentrations used in clinical laboratories. 4. The products of the reaction between PBQ and either plasma (specific absorbance 2.33 x 10(-3) +/- 0.20 x 10(-3) micrograms cm-2) or purified proteins (specific absorbance 2.61 x 10(-3) +/- 0.31 x 10(-3) micrograms cm-2) show an absorption band at 350 nm, which follows Beer's law, and therefore can be used for analytical purposes. 5. The PBQ method has a lower limit of detection (4 micrograms/ml) than that of the biuret method (45 micrograms/ml) for a final reaction mixture of 5.0 and 4.2 ml, respectively.

Spectrophotometric determination of total proteins in blood plasma with p-benzoquinone

In the present study we have documented the use of the reagent p-benzoquinone (PBQ) for the spectrophotometric determination of total protein in blood plasma. Since the products of reaction are stable for several hours at room temperature after the 20-min boiling step, the time at which absorbance is measured is not a critical factor. Common anticoagulants such as EDTYA, citrate, or heparin do not interfere with the PBQ method at concentrations used in clinical laboratories. The products of the reaction between PBQ and either plasma (specific absorbance 2.33 x 10-3 ñ 0.20 x 10-3 ug cm -2) or purified proteins (specific absorbance 2.61 x 10-3 ñ 0.31 x 10-3 ug cm-2) show an absorption band at 350 nm, which follows Beer's law, and therefore can be used for analytical purposes. The PBQ method has a lower limit of detection (4 ug/ml) than that of biuret method (45 yg/ml) for a final reaction mixture of 5.0 and 4.2 ml, respectively

Glycemic response to stress stimulation by ether exposure in adrenalectomized rats.

Stress was induced by short-term ether exposure (2 min) and tail vein puncture in normal (SO), adrenodemedullated (ADM), and adrenalectomized (ADR) rats. In ADM and SO rats stress provoked a significant hyperglycemic response with no change in plasma insulin levels. In ADR rats, on the other hand, the hyperglycemic response was not present. Actually, a significant rapid decrease in blood glucose, plasma insulin and hepatic glycogen content was observed. When the hypoglycemic effect of stress was prevented by glucose injection into ADR rats the decrease in plasma insulin and hepatic glycogen was not observed. The data suggest that the fall in plasma insulin and hepatic glycogen content observed in ADR animals result from an activation of the sympathetic nervous system induced by the decrease in blood glucose.