Protein Malnourished Rats Research Articles

English

The aim of this study is to evaluate the relationships between metronidazole toxicity and pharmacokinetics in protein malnourished rats. The study was carried out on two sets of rats, normally-fed set and protein malnourished set. Each set was divided into a control group and three-treated groups that received metronidazole daily in one dose of 200, 400 or 800 mg/kg for 30 days. Liver enzymes as well as testosterone and gonadotropin hormones levels were estimated. The pharmacokinetic profiles were carried out over a period of 48 h. Metronidazole plasma levels were determined by a validated high-performance liquid chromatography (HPLC) method. Metronidazole has dose dependent effects on hematological profile, liver enzymes, testosterone and gonadotropins which potentiated in protein malnourished rats. Protein malnutrition elevated metronidazole Cmax, AUC0-48, and AUC0-inf, prolonged t1/2 and MRT, and decreased elimination rate constant. The clinical significance of this work should be stressed in case of protein malnourished individuals due to the risk of accumulation of the drug after repeated doses.   Keywords: Protein malnutrition, rats, Metronidazole, pharmacokinetics, pharmacodynamic, Carbamazepine.

Combination of BCAAs and glutamine enhances dermal collagen protein synthesis in protein-malnourished rats

Skin collagen decreases in protein-malnourished states. Amino acids regulate protein metabolism, glutamine stimulates collagen synthesis through the conversion process to proline and provides 75 % of the intracellular free proline in fibroblasts. However, the impact of these amino acids on collagen synthesis under malnutrition has not been examined. We investigated the effect of amino acids on dermal tropocollagen synthesis in protein-malnourished rats. The fractional synthesis rate (FSR, %/h) of dermal tropocollagen was evaluated by the incorporation of L-[ring-(2)H(5)]-phenylalanine after 4 h infusion of each amino acid and the stable isotope. None of the infused 12 single amino acids (glutamine, proline, alanine, arginine, glutamate, glycine, aspartate, serine, histidine, lysine, phenylalanine and threonine) significantly increased the FSR (P = 0.343, one-way ANOVA). In contrast, amino acid mixtures of essential amino acids + glutamine + arginine (EAARQ) and branched-chain amino acids + glutamine (BCAAQ) significantly increased the FSR compared to saline, but the branched-chain amino acids (BCAAs) and amino acid mixture of collagen protein (AAC) did not alter the FSR (saline, 0.96 ± 0.24 %/h; EAARQ, 1.76 ± 0.89 %/h; BCAAQ 1.71 ± 0.36 %/h; BCAAs, 1.08 ± 0.20 %/h and AAC 1.39 ± 0.35 %/h, P < 0.05, Tukey's test). Proline conversion from glutamine represented only 3.9 % of the free proline in skin, as evaluated by the primed-constant infusion of L-d7-proline and L-α-15N-glutamine in rats. These results suggested that the combination of BCAAQ is a key factor for the enhancement of skin collagen synthesis in protein-malnourished rats. The contribution of extracellular free glutamine on de novo proline synthesis and collagen synthesis is very low in vivo compared to the contribution in vitro.

Decreased insulin secretion in islets from protein malnourished rats is associated with impaired glutamate dehydrogenase function: effect of leucine supplementation

We herein studied the role of glutamate dehydrogenase (GDH), in response to leucine (LEU) supplementation, upon insulin secretion of malnourished rats. Weaned male Wistar rats were fed normal-protein (17%) or low-protein diet (6%, LP) for 8 weeks. Half of the rats of each group were supplemented with LEU (1.5%) in the drinking water for the following 4 weeks. Gene and protein expressions, static insulin secretion, and cytoplasmic Ca2+ oscillations were measured. Glutamate dehydrogenase messenger RNA was 58% lower in LP islets, and LEU supplementation augmented it in 28%. The LP islets secreted less insulin when exposed to 20 mmol/L LEU, 20 mmol/L LEU + 2 mmol/L glutamine (with or without 5 mmol/L aminooxyacetic acid, a branched chain aminotransferase inhibitor, or 20 μmol/L epigallocatechin gallate, a GDH inhibitor), 20 mmol/L α-ketoisocaproate, glutamine + 20 mmol/L β-2-aminobicyclo[2.2.1]heptane-2-carboxylic acid (a GDH activator), and 22.2 mmol/L glucose. Leucine supplementation augmented insulin secretion to levels found in normal-protein islets in all the above conditions, an effect that was blunted when islets were incubated with epigallocatechin gallate. The glutamine + β-2-aminobicyclo[2.2.1]heptane-2-carboxylic acid–induced increased [Ca2+]i and oscillations were higher than those for LP islets. Leucine supplementation normalized these parameters in LP islets. Impaired GDH function was associated with lower insulin release in LP islets, and LEU supplementation normalized insulin secretion via restoration of GDH function. In addition, GDH may contribute to insulin secretion through ameliorations of Ca2+ handling in LP islets.

Lithium/nutrition interaction in the brain: A single lithium administration impairs spreading depression in malnourished, but not in well-nourished rats

Lithium salts exert electrophysiological and behavioral effects in animals and humans and have been used clinically in the treatment of bipolar disorders. Little is known about the lithium/nutrition interaction in the developing brain. This work aimed to determine, in adult rats, whether treatment with a single dose of lithium chloride (LiCl) would influence the propagation of the brain excitability-related phenomenon known as cortical spreading depression (CSD). Male well-nourished (W; fed a lab chow diet with 22% protein; n = 22) and previously protein-malnourished rats (M; fed a low-quality 8% protein diet; proteins mostly from vegetable source; n = 20) were treated at 75–80 days of age with a single intraperitoneal injection of either 50 mg/kg LiCl (n = 9 W and 10 M rats) or saline (n = 13 W and 10 M rats). When the pups were 90–110 days, CSD was elicited at the frontal cortex and recorded during 4 hours at two cortical parietal points. In malnourished, but not in well-nourished rats, lithium treatment lowered CSD velocities (P < 0.05), in comparison with saline-injected animals. In a third group (n = 23), in which the low-protein diet was quantitatively corrected to 22%, the lithium effect disappeared (n = 12), compared to saline (n = 11). Our results demonstrate a facilitating effect of malnutrition on the CSD-impairing action of a single lithium administration, suggesting a lithium/nutrition interaction.

Taurine supplementation restored the changes in pancreatic islet mitochondria in the fetal protein-malnourished rat

Intra-uterine growth retardation has been linked to the development of type 2 diabetes in later life. Mitochondrial changes have been suggested as a link between fetal malnutrition and adult insulin resistance. Taurine has been implicated in this process. We investigated whether protein malnutrition in early life alters mitochondria of the pancreatic islets in adulthood, and whether taurine supplementation restores these changes. Male offspring of rats fed a control diet, a low-protein diet or a low-protein diet supplemented with taurine during pregnancy and lactation were weaned onto the control diet. In each group, at 20 weeks of age, intravenous glucose tolerance tests, euglycaemic-hyperinsulinaemic clamp studies, morphometric analysis of the pancreatic islets and ultra-structural analysis of the mitochondria of the β-cells were performed. The expressions of cytochrome c oxidase (COX) I and mitochondrial respiratory chain complex II were also measured. Fetal protein-malnourished rats showed decreased pancreatic islet mass and reduced insulin-secretory responses to a glucose load. These rats also showed reduced mitochondrial DNA-encoded COX I gene expression in the islets. Electron microscopic examination showed abnormal mitochondrial shapes in the β-cells of fetal protein-malnourished rats. Taurine supplementation to the low-protein diet restored all these changes. Our findings indicate that a maternal protein-restriction diet causes long-lasting mitochondrial changes that may contribute to the development of type 2 diabetes later in life. The lack of taurine may be a key causative factor for these dysfunctional mitochondrial changes.

Biomechanical properties of femoral shafts and bone tissue in protein-malnourished rats from weaning to adulthood

The present study was designed to estimate the bone biomechanical repercussion of protein restriction imposed to rats between the 26th and the 135th days of age. Female rats were placed on either a 4% (P4) or 20% (P20) protein diets. Body weight at the end of the study was 350.5 g (SD 31.0 g) in P20 rats and 87.5 g (19.4 g) in P4 ones. Femur weight and length were significantly reduced. The diaphyseal cross-sectional area and the cross-sectional moment of inertia were also diminished. Femoral calcium concentrations in ashes were similar in both groups. However, the femoral mass (calcium mass) was significantly lower in P4 than in P20 rats. When the femoral bone mass was normalized per 100-g femur, no significant difference between groups was found. The right femur of each animal was subjected to a three-point bending test in order to obtain the load/deformation curve. A significant weakening of bone beams, shown by a correlative impairment of diaphyseal ultimate strength and stiffness, was observed in P4 rats. However, when their values were normalized for body weight, the differences disappeared. The bone material quality indicators, or pre-yield bending stiffness (elastic modulus) and yielding stress were similar in P4 and P20 rats. In summary, we have described a number of alterations in bone morphometrical and biomechanical variables in rat femur shafts resulting from protein restriction from weaning to early adulthood. The clear differences in strength and stiffness of bone beams seemed to be the result of an induced subnormal gain in bone structural properties as a consequence of a correlative subnormal gain in bone growth and mass, yet not in bone material properties.

Sympathoinhibition to Bezold–Jarisch reflex is attenuated in protein malnourished rats

Malnutrition affects cardiovascular reflexes, including chemoreflex and baroreflex. In this study we assessed the hypothesis that malnourishment changes the responses in mean arterial pressure (MAP), heart rate (HR) and renal sympathetic nerve activity (RSNA) evoked from Bezold–Jarisch reflex (BJR). Fischer rats were fed diets containing either (6% malnourished or 14% control) protein for 35 days after weaning. There were no differences in baseline MAP (102±4 vs. 95±3mmHg) whereas higher baseline HR (478±18 vs. 360±11bpm; P<0.05,) and reduced sympathoinhibition (ΔRSNA=−54±9 vs. −84±7%; P=0.0208) to BJR activation were found in malnourished rats. We conclude that malnutrition affects the sympathetic control of BJR.

PTH effects on bone are attenuated in protein malnourished rats

PTH effects on bone are attenuated in protein malnourished rats

Effects of metronidazole and probiotics oligosaccharide on bacterial translocation in protein malnutrition

The present study aims to evaluate the effects of metronidazole, probiotics oligosaccharide on indigenous microflora and bacterial translocation (BT) in protein malnourished rats. Thirty male Wistar rats were divided into three groups: protein malnourished rats PM (group1, n = 10) were fed with maize only, protein malnourished rats (group 2, n = 10) were received metronidazole and protein malnourished rats (group 3, n = 10) were received both metronidazole and probiotics-oligosaccharide for fifteen days. Metronidazole (1000 mg/kg/day) was given via an orogastric feeding tube to the second and third groups. Lyophilized probiotics-oligosaccharide (0.5 mg/g body weight/day) was given in two doses via the same route to the third group. All animals were sacrificed after fifteen days of protein malnutrition and cultures of the mesenteric lymph nodes (MLNs), liver, spleen and cecal contents were done. The incidence of bacterial translocation (BT) was 30% (3/10) in protein malnourished group 1,60% (06/10) in group 2 where protein malnutrition was associated with metronidazole and 25% (2.5/10) in group 3 whose animals were subjected to protein malnutrition associated with metronidazole and probiotics oligosaccharide. A significant increase in the BT incidence was found in group 2 (P < 0.05), while a significant decrease was found in group 3 when compared to group 1. The total bacterial count of cecal flora was significantly low in group 3 than in group 1 (P < 0.01). These results suggest that the incidence of BT in protein malnutrition is increased by using an antibiotic while probioticsoligosaccharide decreases this incidence in protein malnutrition induced by antibiotic. Thus, we conclude that probiotics-oligosaccharide can effectively protect the intestinal mucosa and prevent BT in protein malnourished infants.

Preliminary report: Leucine supplementation enhances glutamate dehydrogenase expression and restores glucose-induced insulin secretion in protein-malnourished rats

Low-protein diet impairs insulin secretion in response to nutrients and may induce several metabolic disorders including diabetes, obesity, and cardiovascular disease. In the present study, the influence of leucine supplementation on glutamate dehydrogenase (GDH) expression and glucose-induced insulin secretion (GIIS) was investigated in malnourished rats. Four groups were fed with different diets for 12 weeks: a normal-protein diet (17%) without or with leucine supplementation or a low (6%)-protein diet without (LP) or with leucine supplementation (LPL). Leucine (1.5%) was supplied in the drinking water. Western blotting analysis revealed reduced GDH expression in LP, whereas LPL displayed improved GDH expression, similar to control. The GIIS and leucine-induced insulin release were also enhanced in LPL compared with LP and similar to those observed in rats fed a normal-protein diet without leucine supplementation. In addition, GDH allosteric activators produced an increased insulin secretion in LPL. These findings indicate that leucine supplementation was able to increase GDH expression leading to GIIS restoration, probably by improved leucine metabolic pathways.

Dietary Glutamic Acid Increased Postprandial Leucine Availability in Protein Malnourished Rats

Glutamic acid (Glu), the most abundant amino acid in food proteins, is major intestinal energy source. Recently we have found that Glu removal from foods reduced dietary leucine (Leu) availability by reducing its oxidation. The present study investigated roles of dietary Glu on Leu metabolism in details using protein malnourished rats.Fischer strain male rats were fed normal amino acid (AA) defined diet containing 4% Glu or low AA diet (50% reduction) for 1 week. Then, the rats in each dietary group were divided into two groups and given daily amount of experimental diets with or without Glu in which all the Leu was replaced with [1‐13C]Leu. After 24h collection of expired air samples, blood and tissue samples were harvested. Breath 13CO2 and [1‐13C]Leu incorporation in tissue proteins were analyzed by IRMS and GCMS, respectively.Feeding of low AA diet reduced dietary Leu oxidation markedly. Glu removal from low AA diet significantly enhanced dietary Leu oxidation and reduced its incorporation in liver protein and plasma albumin but not kidney or muscle proteins, indicating decrease in dietary Leu availability for hepatic protein synthesis in these animals presumably due to increase in its intestinal oxidation. Besides, the effects of Glu removal from normal AA diet were not apparent. These results suggest Leu‐sparing effects of dietary Glu by reducing postprandial Leu oxidation especially in protein malnutrition.

Effects of tactile stimulation and underwater trauma on the behavior of protein-malnourished rats in the elevated plus-maze test

Effects of tactile stimulation and underwater trauma on the behavior of protein-malnourished rats in the elevated plus-maze test

Effects of tactile stimulation and underwater trauma on the behavior of protein-malnourished rats in the elevated plus-maze test

The objective of the present study was to evaluate the interaction of two kinds of external stimuli (long-term tactile stimulation and underwater trauma) on the exploratory behavior of malnourished rats in an elevated plus maze (EPM). The results showed that tactile stimulation partially recovered body weight deficits produced by malnutrition and decreased EPM exploration. Malnutrition increased EPM exploration while underwater trauma decreased it, thus reducing differences between well nourished and malnourished animals. These data show that only one of the two external stimuli (underwater trauma) interacted with diet condition, suggesting that the stress caused by trauma was efficient in decreasing higher EPM exploration, equalizing the response of malnourished animals to that of non-traumatized well-nourished animals.

Effects of tactile stimulation and underwater trauma on the behavior of protein-malnourished rats in the elevated plus-maze test.

Effects of tactile stimulation and underwater trauma on the behavior of protein-malnourished rats in the elevated plus-maze test.

Impact of Aluminum on Protein Malnourished Rat Brain

Available reports suggesting that aluminum and protein deficiency both are causing deficits in the neurobehavioral development learning. It has been aimed to study the impact of protein malnutrition on the general responses of brain to aluminum exposure. Male albino rats of Wistar strain weighing 80-100g were injected intraperitoneally, at a dose of 4.2 mg / Kg body weight (15 % of LD50 ) / day for 4 weeks while they were maintained on either 18% protein diet or 6% protein diet. Rat brain aluminum content was significantly increased in the aluminum-exposed rats of the normal protein diet group but this increase was insignificant in the low protein diet group. The changes in brain weight, acid and alkaline phosphoesterases, aspartate and alanine aminotransferases, reduced glutathione, thiobarbituric acid reactive substances, DNA, RNA and protein contents suggested a nonspecific impact of dietary protein status on the response of brain to aluminum exposure. Name of the department and Institution where the work was done: Department of Physiology, University of Calcutta. The source of any support received: University Grants Commission, New Delhi. India.

Increased rewarding properties of morphine in perinatally protein-malnourished rats

In the current research, we assessed the influence of a protein malnutrition schedule from the 14th day of gestation up to 40 days of age (D-rats) on the rewarding properties of morphine in adult rats by means of the conditioned place preference paradigm. Well-nourished animals (C-rats) administered with different doses of morphine (0.75, 1.5, 3, 6, 12 or 24 mg/kg i.p.) exhibited a conditioning place preference with doses of 3 and 6 mg/kg, whereas in D-rats such a conditioning effect was observed with doses of 1.5 and 3 mg/kg. No adverse effects were observed in either C- or D-rats for the higher doses of morphine. In addition, when animals of both groups were pretreated twice a day for 3 days with increasing doses of morphine (5, 10 and 20 mg/kg s.c.), only D-rats elicited sensitization to the conditioning effect with the lowest dose of morphine (0.75 mg/kg i.p.). Furthermore, sensitized D-rats showed a selective and significant increase in FosB expression in the nucleus accumbens (core and shell), basolateral amygdala and medial prefrontal cortex, brain areas that are functionally related to the rewarding neural circuit. These results demonstrate that a deficient nutritional status during the perinatal period results in adult subjects having neural alterations, leading to an increased responsiveness to morphine and/or enhanced reinforcement effects, which correlates with an overexpression of FosB in selective brain areas related to the rewarding network.

Prenatal malnutrition and sleep states in adult rats: Effects of restraint stress

Independently, prenatal malnutrition and psychological/physical stress have been shown to affect sleep architecture in adult rats. As malnutrition and stress commonly co-exist in malnourished human populations, the objective of the present study was to ascertain the combined effects of these two insults by examining sleep–wake parameters following a brief restraint stress in prenatally protein malnourished rats. The male offspring of rats provided with a protein deficient diet (6% casein) for 5 weeks prior to mating and throughout pregnancy were implanted with recording electrodes beginning at postnatal day 90. Polygraph recordings were obtained to quantify sleep states during the first 4 h of the dark phase of the cycle on 2 consecutive days. The first followed a 24-h habituation session to the recording chamber (baseline). The second occurred at the same time of day but followed 20 min of restraint stress in a Plexiglas tube. During baseline, prenatally malnourished rats spent more time in rapid eye movement sleep (REMS) in the first 2 h after “lights off” (block 1), and greater amounts of wakefulness (W) with a corresponding reduction in slow wave sleep (SWS) in the second two hours (block 2), as compared with controls. Following stress, the sleep architecture of both groups of rats remained unaltered in block 1 relative to their baseline day. In block 2, both groups exhibited significant reductions in SWS and REMS with significantly greater reductions being expressed in the prenatally malnourished group (most dramatically, REMS was completely eliminated). These findings suggest that sleep disturbances may be more severe in those malnourished human populations subjected to acutely stressful experiences.

Effects of protein malnutrition on oxidative status in rat brain

Objectives This study evaluated the effects of protein malnutrition on oxidative status in rat brain areas. Methods We investigated various parameters of oxidative status, free radical content (dichlorofluorescein formation), indexes of damage to lipid (thiobarbituric acid-reactive substances assay), and protein damage (tryptophan and tyrosine content) in addition to total antioxidant reactivity levels and antioxidant enzyme activities of superoxide dismutase, glutathione peroxidase, and catalase in different cerebral regions (cortex, hippocampus, and cerebellum) from rats subjected to prenatal and postnatal protein malnutrition (control 25% casein and protein malnutrition 7% casein). Results Protein malnutrition altered various parameters of oxidative stress, especially damage to macromolecules. Free radical content was unchanged by protein malnutrition. There was an increase in levels of thiobarbituric acid-reactive substances, the index of lipid peroxidation, in the cerebellum and cerebral cortex ( P < 0.05) from protein-malnourished rats. Moreover, significant decreases in tryptophan and tyrosine in all tested brain structures ( P < 0.05) were observed. Catalase activity was significantly decreased in the cerebellum ( P < 0.05). In addition, a significant decrease in total antioxidant reactivity levels ( P < 0.05) was observed in the cerebral cortex from protein-malnourished rats. Conclusions The present data indicated that protein malnutrition increased oxidative damage to lipids and proteins from the studied brain areas. These results may be an indication of an important mechanism for changes in brain development that are caused by protein malnutrition.

The interaction of housing condition and acute immobilization stress on the elevated plus-maze behaviors of protein-malnourished rats.

Protein malnutrition induces structural, neurochemical and functional alterations in the central nervous system, leading to behavioral alterations. In the present study, we used the elevated plus-maze (EPM) as a measure of anxiety to evaluate the interaction between acute immobilization and housing conditions on the behavior of malnourished rats. Pups (6 males and 2 females) were fed by Wistar lactating dams receiving a 6% (undernourished) or 16% (well-nourished) protein diet. After weaning, the animals continued to receive the same diets ad libitum until 49 days of age when they started to receive a regular lab chow diet. From weaning to the end of the tests on day 70, the animals were housed under two different conditions, i.e., individual or in groups of three. On the 69th day, half of the animals were submitted to immobilization for 2 h, while the other half were undisturbed, and both groups were tested 24 h later for 5 min in the EPM. Independent of other factors, protein malnutrition increased, while immobilization and social isolation per se decreased, EPM exploration. Analysis of the interaction of diet vs immobilization vs housing conditions showed that the increased EPM exploration presented by the malnourished group was reversed by acute immobilization in animals reared in groups but not in animals reared individually. The interaction between immobilization and housing conditions suggests that living for a long time in social isolation is sufficiently stressful to reduce the responses to another anxiogenic procedure (immobilization), while living in groups prompts the animals to react to acute stress. Thus, it is suggested that housing condition can modulate the effects of an anxiogenic procedure on behavioral responses of malnourished rats in the EPM.

Spatial Learning Deficits Induced by Muscimol and CL218,872: Lack of Effect of Prenatal Malnutrition

The sensitivity of prenatal protein malnourished rats to the amnestic properties of the direct GABA<PRE>A</PRE> receptor agonist muscimol and the selective benzodiazepine (BZ) receptor agonist, CL218,872, was studied in the male offspring of rats provided with a protein deficient diet (6% casein) for 5 weeks prior to mating and throughout pregnancy. At postnatal day 90, rats were tested during acquisition of the submerged platform version of the Morris water maze task using four systemic doses of muscimol (0.1, 0.3, 1.0 and 1.8 mg/kg i.p.) or three systemic doses of CL218,872 (1.0, 3.2, and 5.6 mg/kg i.p.). In a dose dependent manner both drugs impaired acquisition of the task and impaired accuracy of the search pattern on the probe trial (platform removed). However, neither drug dissociated the performance of the two nutritional groups. These data are important in light of previous findings of differential behavioral effects of the non-specific BZ agonist, chlordiazepoxide (CDP), on spatial learning and on drug discrimination in prenatally malnourished rats and in the context of previous findings of reduced sensitivity to the anxiolytic effects of non-specific BZ receptor agonists across a wide variety of models of malnutrition. The present findings also support the concept that prenatal malnutrition does not affect the global functioning of the GABA<PRE>A</PRE> receptor, but fundamentally alters the way in which a subset of GABA<PRE>A</PRE> receptors (i.e. those containing the 2, 3 and/or the 5 but not the 1 subunit) is modulated by BZs.