Fenugreek potent activity against nitrate-induced diabetes in young and adult male rats

Nitrate has described as an endocrine disruptor that promotes onset of diabetes. This study was undertaken to evaluate diabetic effect of high nitrate intake in young and adult male rats and its amelioration by fenugreek administration. The study revealed significant increase in serum glucose and blood glycosylated hemoglobin (HbA1c%), while serum insulin and liver glycogen were decreased among nitrate exposed animals, in particular the young group. A significant reduction in the body weight gain and serum thyroid hormones (T4 & T3) was also recorded. Further reduction in serum levels of urea and creatinine, as well as total protein in serum, liver and pancreas was demonstrated, with elevation in their levels in the urine of all nitrate exposed groups. Meanwhile, the activity of serum transaminases (ALT and AST) was increased, with decline in their activity in the liver tissue. In addition, an elevation in serum total bilirubin, tissues (liver and pancreas) nitric oxide and lipid profile, as well as liver activity of glucose-6-phosphatase was recorded. Fenugreek administration to nitrate exposed rats was found to be effective in alleviating hyperglycemia and other biochemical changes characterizing nitrate-induced diabetes. So, fenugreek can be considered to possess potent activity against onset of nitrate induced-diabetes.     

Protein malnutrition during fetal programming induces fatty liver in adult male offspring rats

We evaluated the effects of protein malnutrition on liver morphology and physiology in rats subjected to different malnutrition schemes. Pregnant rats were fed with a control diet or a low protein diet (LPD). Male offspring rats received a LPD during gestation, lactation, and until they were 60 days old (MM group), a late LPD that began after weaning (CM), or a LPD administrated only during the gestation-lactation period followed by a control diet (MC). On day 60, blood was collected and the liver was dissected out. We found a decrease in MM rats’ total body (p < 0.001) and liver (p < 0.05) weight. These and CM rats showed obvious liver dysfunction reflected by the increase in serum glutamic pyruvic transaminase (SGOT) (MM p < 0.001) and serum glutamic pyruvic transaminase (SGPT) (MM and CM p < 0.001) enzymes, and liver content of cholesterol (MM and CM p < 0.001) and triglycerides (MM p < 0.01; CM p < 0.001), in addition to what we saw by histology. Liver dysfunction was also shown by the increase in gamma glutamyl transferase (GGT) (MM, MC, and CM p < 0.001) and GST-pi1 (MM and CM p < 0.001, MC p < 0.05) expression levels. MC rats showed the lowest increment in GST-pi1 expression (MC vs. MM; p < 0.001, MC vs. CM; p < 0.01). ROS production (MM, CM, and MC: p < 0.001), lipid peroxidation (MM, CM, and MC p < 0.001), content of carbonyl groups in liver proteins (MM and CM p < 0.001, MC p < 0.01), and total antioxidant capacity (MM, CM, and MC p < 0.001) were increased in the liver of all groups of malnourished animals. However, MM rats showed the highest increment. We found higher TNF-α (MM and CM p < 0.001), and IL-6 (MM and CM p < 0.001) serum levels and TGF-β liver content (MM p < 0.01; CM p < 0.05), in MM and CM groups, while MC rats reverted the values to normal levels. Pro-survival signaling pathways mediated by tyrosine or serine/threonine kinases (pAKT) (MM and CM p < 0.001; MC p < 0.01) and extrasellular signal-regulated kinase (pERKs) (MM p < 0.01; CM p < 0.05) appeared to be activated in the liver of all groups of malnourished rats, suggesting the presence of cells resistant to apoptosis which would become cancerous. In conclusion, a LPD induced liver damage whose magnitude was related to the developmental stage at which malnutrition occurs and to its length.     

Prolactin response to stress in neonatally estrogenized male rats

Male Wistar rats were injected either 500 micrograms of estradiol benzoate or olive oil on their first day of life. Blood samples were obtained from the adult by decapitation, by decapitation after 15 min of restraint, by decapitation 10 min after a 5 min period of ether exposure or by jugular venipuncture after 60 s of ether exposure. Prolactin (PRL) plasma levels were measured by RIA. The PRL levels in samples obtained by decapitation were similar to control and estrogenized groups. A similar response to restraint was also found in both groups. Sixty s of exposure to ether stress stimulates PRL secretion only in the estrogenized males, this effect being blocked by treatment with Normifensine (5 mg/kg two hours prior to blood sampling). These results suggested that estrogenized male rats show greater sensitivity to acute ether stress than the controls, and that changes in the dopaminergic system could be involved in this response.     

Prenatal stress induces long-term effects in cell turnover in the hippocampus-hypothalamus-pituitary axis in adult male rats

Subchronic gestational stress leads to permanent modifications in the hippocampus-hypothalamus-pituitary-adrenal axis of offspring probably due to the increase in circulating glucocorticoids known to affect prenatal programming. The aim of this study was to investigate whether cell turnover is affected in the hippocampus-hypothalamus-pituitary axis by subchronic prenatal stress and the intracellular mechanisms involved. Restraint stress was performed in pregnant rats during the last week of gestation (45 minutes; 3 times/day). Only male offspring were used for this study and were sacrificed at 6 months of age. In prenatally stressed adults a decrease in markers of cell death and proliferation was observed in the hippocampus, hypothalamus and pituitary. This was associated with an increase in insulin-like growth factor-I mRNA levels, phosphorylation of CREB and calpastatin levels and inhibition of calpain -2 and caspase -8 activation. Levels of the anti-apoptotic protein Bcl-2 were increased and levels of the pro-apoptotic factor p53 were reduced. In conclusion, prenatal restraint stress induces a long-term decrease in cell turnover in the hippocampus-hypothalamus-pituitary axis that might be at least partly mediated by an autocrine-paracrine IGF-I effect. These changes could condition the response of this axis to future physiological and pathophysiological situations.     

Neonatal administration of N-acetyl-L-aspartyl-L-glutamate induces early neurodegeneration in hippocampus and alters behaviour in young adult rats

N-acetyl-L-aspartyl-L-glutamate (NAAG) is a dipeptide that could be considered a sequestered form of L-glutamate. As much as 25% of L-glutamate in brain may be present in the form of NAAG. NAAG is also one of the most abundant neuroactive small molecules in the CNS: it is an agonist at Group II metabotropic glutamate receptors (mGluR II) and, at higher concentrations, at the N-methyl-D-aspartate (NMDA) type of ionotropic glutamate receptors. As such, NAAG can be either neuroprotective or neurotoxic and, in fact, both characteristics have been discussed and described in the literature. In the present studies, 250 nmol NAAG was infused into each lateral cerebral ventricle of 12-day-old rat pups and, using Nissl-stained sections, neurodegeneration in the hippocampus was evaluated 24 or 96 h after the infusion. In several experiments, the neuronal death was also visualised by Fluoro-Jade B staining and studied by TUNEL technique. Some of the NAAG-treated animals were allowed to survive until 50 days post partum and subjected to behavioural (open field) tests. The administration of NAAG to 12-day-old rats resulted in extensive death of neurons particularly in the dentate gyrus of the hippocampus. The neurodegeneration was, in part, prevented by administration of an NMDA receptor antagonist MK-801 (0.1 mg/kg). The nuclear DNA-fragmentation demonstrated by TUNEL technique pointed to the presence of non-specific single-strand DNA cleavage. The NAAG-associated neonatal neuronal damage may have perturbed development of synaptic circuitry during adolescence as indicated by an altered performance of the experimental animals in the open field testing (changes in grooming activity) at postnatal day 50. The results underscore the potential neurotoxicity of NAAG in neonatal rat brain and implicate neonatally induced, NMDA receptor-mediated neuronal loss in the development of abnormal behaviour in young adult rats.     

Bisphenol A exposure at an environmentally relevant dose induces meiotic abnormalities in adult male rats

Whether environmental exposure to bisphenol A (BPA) may induce reproductive disorders is still controversial but certain studies have reported that BPA may cause meiotic abnormalities in C. elegans and female mice. However, little is known about the effect of BPA on meiosis in adult males. To determine whether BPA exposure at an environmentally relevant dose could induce meiotic abnormalities in adult male rats, we exposed 9-week-old male Wistar rats to BPA by gavage at 20 μg/kg body weight (bw)/day for 60 consecutive days. We found that BPA significantly increased the proportion of stage VII seminiferous epithelium and decreased the proportion of stage VIII. Consequently, spermiation was inhibited and spermatogenesis was disrupted. Further investigation revealed that BPA exposure delayed meiosis initiation in the early meiotic stage and induced the accumulation of chromosomal abnormalities and meiotic DNA double-strand breaks (DSBs) in the late meiotic stage. The latter event subsequently activated the phosphatidylinositol 3-kinase-related protein kinase (ATM). Our results suggest that long-term exposure to BPA may lead to continuous meiotic abnormalities and ultimately put mammalian reproductive health at risk.     

Prolactin increases the density of striatal dopamine receptors in normal and hypophysectomized male rats

Administration of prolactin to adult male rats, by s.c. injection, significantly increases the density of the striatal dopamine (DA) receptors, without altering the apparent affinity of the receptors for [³H]spiroperidol. Larger doses of prolactin are required to increase the density of the striatal DA receptors in hypophysectomized rates compared to normal rats. These results suggest that prolactin might be the common mediator of the increase in striatal DA receptor density produced by either estrogen or haloperidol administration. Monitoring and/or altering prolactin levels might be informative in neurologic or psychiatric disorders involving striatal DA neurotransmission.     

Corticosteroids and lung surfactant levels in adult male rats

Following lung instillation in adult male rats of 3.4 mumol hexavalent chromium (K2Cr2O7) dissolved in 0.5 ml of 0.9% NaCl, increased levels of lung surfactant could be detected after 48 h. The blood serum concentration of corticosterone was elevated in these animals. Blood serum thyroxine and triiodothyronine showed an initial increase after lung instillation of hexavalent chromium followed by a decline. Metabolism of testosterone by the alveolar macrophages to 17 beta-hydroxy-5 alpha-androstane-3-one and 5 alpha-androstane-3 alpha, 17 beta-diol was reduced 6 and 12 h after the K2Cr2O7 instillation, which was also associated with damage of lung cell function and decreased uptake by the alveolar macrophages of Candida albicans particles. As early as 12 h after s.c. administration of 400 micrograms dexamethasone/100 g body wt, increased levels of lung surfactant could be measured. At this time the lungs showed no signs of cellular damage, and metabolism of testosterone as well as uptake of Candida albicans particles by the alveolar macrophages were normal. Lower s.c. doses of dexamethasone did not result in raising the levels of lung surfactant in 12 h. Within 12 h after s.c. administration of large doses of testosterone, dihydrotestosterone or dehydroepiandrosterone no measurable effects on the levels of lung surfactant could be measured. Since animals treated with dexamethasone (200 micrograms/100 g body wt) or long-acting synthetic ACTH (100 micrograms i.m. Synacthen Depot/100 g body wt) for 5 days after lung instillation of K2Cr2O7 had extremely high levels of lung surfactant, it is concluded that the corticosteroids in adult rats may help to create augmented surfactant levels following lung intoxication. This could proceed via stimulation of surfactant production and reduction of surfactant removal. Different aspects of lung surfactant metabolism are discussed.     

Testosterone potentiation of the effectiveness of ACTH1-24 on the induction of the stretch-yawning syndrome (SYS) in male guinea pigs

Ventricular administration of ACTH^1–24 stimulated stretch-yawning in castrated male guinea pigs in a dose-related manner. Daily subcutaneous treatment with testosterone propionate (TP) facilitated the effects of ACTH^1–24 on this response. TP given without ACTH^1–24 stimulated yawning, but not stretching or combined stretch-yawning. Unlike most other species, guinea pigs did not display auto-grooming, scratching, or wet-dog shaking in response to intracranial ACTH^1–24. The results suggest that testosterone may alter the sensitivity of neural mechanisms which are responsive to ACTH^1–24.     

Prolactin stimulates the L-type calcium channel-mediated transepithelial calcium transport in the duodenum of male rats

Elevated plasma levels of prolactin (PRL) have been reported in several physiological and pathological conditions, such as lactation, prolactinoma, and dopaminergic antipsychotic drug uses. Although PRL is a calcium-regulating hormone that stimulates intestinal calcium absorption in lactating rats, whether PRL is capable of stimulating calcium absorption in male rats has been elusive. Herein, the transepithelial calcium transport and electrical characteristics were determined in ex vivo duodenal tissues of male rats by Ussing chamber technique. We found that PRL receptors were abundantly present in the basolateral membrane of the duodenal epithelial cells. PRL (200–800 ng/mL) markedly increased the active duodenal calcium transport in a dose-dependent fashion without effect on the transepithelial resistance. The PRL-enhanced active duodenal calcium transport was completely abolished by L-type calcium channel blocker (nifedipine) as well as inhibitors of the major basolateral calcium transporters, namely plasma membrane Ca²⁺-ATPase and Na⁺/Ca²⁺ exchanger. Several intracellular mediators, such as JAK2, MEK, PI3K and Src kinase, were involved in the PRL-enhanced transcellular calcium transport. Moreover, PRL also stimulated the paracellular calcium transport in the duodenum of male rats in a PI3K-dependent manner. In conclusion, PRL appeared to be a calcium-regulating hormone in male rats by enhancing the L-type calcium channel-mediated transcellular and the paracellular passive duodenal calcium transport. This phenomenon could help restrict or alleviate negative calcium balance and osteoporosis that often accompany hyperprolactinemia in male patients.     

Faecal corticosterone and immunoglobulin A in young adult rats

Quantitative analyses of relevant molecules in faeces may have potential as future non-invasive measures of stress. This study examined levels of faecal corticosterone and immunoglubulin A (IgA) in young adult rats and how these levels varied according to age, gender and time of day. Faecal samples were collected from 40 young adult rats (7 weeks old, n = 20 and 10 weeks old, n = 20) of both sexes from two time windows: day and night. The concentrations of corticosterone and IgA were measured by ELISAs following organic solvent extraction and aqueous extraction, respectively, of the molecules from faecal pellets. The production of faeces per time unit was higher in males than in females, and linear correlations were found between the faecal concentrations of corticosterone and IgA and total amounts of the respective molecules excreted in faeces per kg body weight per hour. In all further analyses the levels of the two molecules were calculated as amounts secreted per kg of body weight per hour. There was no gender difference between females and males in the production of corticosterone and IgA, but 7-week-old animals excreted significantly higher amounts of both molecules than did 10-week-old rats. The levels of IgA excreted by female rats were higher in the evening than in the morning, and male rats excreted higher concentrations of corticosterone in the morning than in the evening.     

Glutamate-induced differential mitochondrial response in young and adult rats

Excitatory amino acid glutamate is involved in neurotransmission in the nervous system but it becomes a potent neurotoxin under variety of conditions. However, the molecular mechanism of excitotoxicity is not known completely. We have studied the influence of glutamate on intracellular calcium and mitochondrial functions in cortical slices from young and adult rats. The slices from both the age groups exhibited comparable intracellular calcium changes upon glutamate stimulation. Glutamate treatment caused a decrease in adenosine 5'-diphosphate/adenosine 5'-triphosphate (ADP/ATP) and an increase in nicotinamide adenine dinucleotide/nicotinamide adenine dinucleotide reduced form (NAD/NADH) ratio in both the age groups but the magnitude and the nature of temporal change was different. Glutamate-induced decrease in ATP/ADP and increase in NAD/NADH ratio was significantly higher in slices from the adult as compared to the young rats. The slices from young rats elicited slightly higher mitochondrial depolarization than adult rats. However, the formation of reactive oxygen species (ROS) and lactate dehydrogenase (LDH) release were significantly higher in adult rats as compared to young rats. Glutamate-induced mitochondrial depolarization, ROS formation and LDH release were highly dependent on the presence of Ca(2+) in the extracellular medium. The treatment of slices with mitochondrial inhibitors rotenone and oligomycin inhibited ROS formation and LDH release substantially. Our results suggest that the glutamate-induced increase in intracellular calcium is not the only factor responsible for neuronal cell death but the mitochondrial functions could be crucial in excitotoxicity.     

Dobutamine stress echocardiography in healthy adult male rats

Medical imaging market consists of several billion tests per year worldwide. Out of these, at least one third are cardiovascular procedures. Keeping in mind that each test represents a cost, often a risk, and a diagnostic hypothesis, we can agree that every unnecessary and unjustifiable test is one test too many. Small individual costs, risks, and wastes multiplied by billions of examinations per year represent an important population, society and environmental burden. Unfortunately, the appropriateness of cardiac imaging is extra-ordinarily low and there is little awareness in patients and physicians of differential costs, radiological doses, and long term risks of different imaging modalities. For a resting cardiac imaging test, being the average cost (not charges) of an echocardiogram equal to 1 (as a cost comparator), the cost of a CT is 3.1x, of a SPECT 3.27x, of a Cardiovascular Magnetic Resonance imaging 5.51x, of a PET 14.03x, and of a right and left heart catheterization 19.96x. For stress cardiac imaging, compared with the treadmill exercise test equal to 1 (as a cost comparator), the cost of stress echocardiography is 2.1x and of a stress SPECT scintigraphy is 5.7x. Biohazards and downstream long-term costs linked to radiation-induced oncogenesis should also be considered. The radiation exposure is absent in echo and magnetic resonance, and corresponds to 500 chest x rays for a sestamibi cardiac stress scan and to 1150 chest x rays for a thallium scan. The corresponding extra-risk in a lifetime of fatal cancer is 1 in 2000 exposed patients for a sestamibi stress and 1 in 1000 for a thallium scan. Increased awareness of economic, biologic, and environmental costs of cardiac imaging will hopefully lead to greater appropriateness, wisdom and prudence from both the prescriber and the practitioner. In this way, the sustainability of cardiac imaging will eventually improve.     

Gonadectomy impairs T-maze acquisition in adult male rats

Recent studies have shown that chronic gonadectomy increases the density of dopaminergic axons in prefrontal but not sensorimotor cortices in adult male rats. Since supranormal prefrontal cortical dopamine stimulation is known to impair rats' performance in T-maze delayed alternation paradigms, we tested whether long-term gonadectomy might also impair T-maze performance. For comparison, sensorimotor functions were also assessed. Adult male rats were gonadectomized and placebo-, estradiol-, or testosterone propionate-treated or were sham operated and placebo-treated. Four weeks after surgery, the subjects were tested using a rotorod apparatus and in the acquisition of a T-maze delayed alternation paradigm. Gonadectomized placebo-treated and estradiol-treated rats took significantly longer to acquire the T-maze rule than controls, and gonadectomized, testosterone-treated rats acquired the task within the same time frame as controls. No group differences were detected in rotorod performance. Thus, chronic gonadectomy induced testosterone-sensitive, estradiol-insensitive acquisition deficits in a spatial learning task but had no demonstrable effects on the sensorimotor functions tested.     

Long-term exposure to a continuous 900 MHz electromagnetic field disrupts cerebellar morphology in young adult male rats

The pathological effects of exposure to an electromagnetic field (EMF) during childhood and adolescence may be greater than those from exposure during adulthood. We investigated possible pathological changes in the cerebellum of adolescent rats exposed to 900 MHz EMF daily for 25 days. We used three groups of six 21-day-old male rats as follows: unexposed control group (Non-EG), sham-exposed group (Sham-EG) and an EMF-exposed group (EMF-EG). EMF-EG rats were exposed to EMF in an EMF cage for 1 h daily from postnatal days 21 through 46. Sham-EG rats were placed in the EMF cage for 1 h daily, but were not subjected to EMF. No procedures were performed on the Non-EG rats. The cerebellums of all animals were removed on postnatal day 47, sectioned and stained with cresyl violet for histopathological and stereological analyses. We found significantly fewer Purkinje cells in the EMF-EG group than in the Non-EG and Sham-EG groups. Histopathological evaluation revealed alteration of normal Purkinje cell arrangement and pathological changes including intense staining of neuron cytoplasm in the EMF-EG group. We found that exposure to continuous 900 MHz EMF for 1 h/day during adolescence can disrupt cerebellar morphology and reduce the number of Purkinje cells in adolescent rats.     

Serum concentrations of reproductive hormones after administration of various anesthetics to immature and young adult male rats

Immature and young adult male rats were either castrated or unoperated. One of seven anesthetic agents (Rompun, Bio-Tal, Thiopental, pentobarbital, ketamine, halothane, or ether) was administered. When the animals were clearly anesthetized, they were decapitated. Control rats were decapitated without anesthesia. Serum concentrations of luteinizing hormone (LH), follicle stimulating hormone (FSH), prolactin, testosterone, and androstenedione were determined by radioimmunoassay. None of the anesthetics was clearly suitable for study of all these hormones. Most would be suitable for acute LH studies. Ketamine and halothane appeared inappropriate for FSH studies in immature rats. Pentobarbital, Rompun, and ether caused increases in serum prolactin. Most of the agents appeared to cause a reduction in serum testosterone in intact rats but an increase in castrated animals, suggesting an inhibition of testicular androgen secretion and a stimulation of adrenal androgen secretion.