Iracema Senna de Andrade

MONOSODIUM GLUTAMATE (MSG)-OBESE RATS DEVELOP GLUCOSE INTOLERANCE AND INSULIN RESISTANCE TO PERIPHERAL GLUCOSE UPTAKE

Different levels of insulin sensitivity have been described in several animal models of obesity as well as in humans. Monosodium glutamate (MSG)-obese mice were considered not to be insulin resistant from data obtained in oral glucose tolerance tests. To reevaluate insulin resistance by the intravenous glucose tolerance test (IVGTT) and by the clamp technique, newborn male Wistar rats (N = 20) were injected 5 times, every other day, with 4 g/kg MSG (N = 10) or saline (control; N = 10) during the first 10 days of age. At 3 months, the IVGTT was performed by injecting glucose (0.75 g/kg) through the jugular vein into freely moving rats. During euglycemic clamping plasma insulin levels were increased by infusing 3 mU.kg-1.min-1 of regular insulin until a steady-state plateau was achieved. The basal blood glucose concentration did not differ between the two experimental groups. After the glucose load, increased values of glycemia (P < 0.001) in MSG-obese rats occurred at minute 4 and from minute 16 to minute 32. These results indicate impaired glucose tolerance. Basal plasma insulin levels were 39.9 +/- 4 microU/ml in control and 66.4 +/- 5.3 microU/ml in MSG-obese rats. The mean post-glucose area increase of insulin was 111% higher in MSG-obese than in control rats. When insulinemia was clamped at 102 or 133 microU/ml in control and MSG rats, respectively, the corresponding glucose infusion rate necessary to maintain euglycemia was 17.3 +/- 0.8 mg.kg-1.min-1 for control rats while 2.1 +/- 0.3 mg.kg-1.min-1 was sufficient for MSG-obese rats. The 2-h integrated area for total glucose metabolized, in mg.min.dl-1, was 13.7 +/- 2.3 vs 3.3 +/- 0.5 for control and MSG rats, respectively. These data demonstrate that MSG-obese rats develop insulin resistance to peripheral glucose uptake.

Neonatal Treatment with Monosodium Glutamate Increases Plasma Corticosterone in the Rat

Monosodium glutamate (MSG) has been shown to alter several neuroendocrine functions in neonatally treated rats. To evaluate for possible alterations in the hypothalamic-pituitary-adrenal axis, we injected rats during the neonatal period with MSG or saline (controls). An increase in basal plasma corticosterone levels associated with a blunted circadian variation was observed. Ether exposure produced a significant elevation in plasma corticosterone concentration in both groups of animals. However, while the increase in controls was 181.3% for male and 193.9% for female rats, in the MSG-treated rats it was only 60.7 and 31.6%, respectively. The intraperitoneal administration of high dexamethasone doses blocked corticosterone secretion in both groups. However, whereas the lowest dose (0.10 microgram/kg) suppressed corticosterone secretion in control animals, it was ineffective in MSG-treated rats. The morphological study of adrenals revealed signs of a hyperfunctional state in MSG-treated rats. These data suggest that the central lesions produced by MSG treatment disrupt the regulation of the hypothalamic-pituitary-adrenal axis.

HORMONAL AND METABOLIC ADAPTATIONS TO FASTING IN MONOSODIUM GLUTAMATE-OBESE RATS

Abstract The effect of fasting on hormonal and metabolic variables was evaluated in normal rats and in rats with obesity induced by neonatal treatment with monosodium glutamate (MSG). The hyperinsulinemia of the fed obese rats was reversed by fasting. Plasma corticosterone was also high in the fed obese and decreased to levels similar to fed controls, while it increased in the latter group during fasting. In contrast, thyroid hormone levels decreased in controls but increased in the obese rats in response to fasting. The fed obese group had lower carcass protein and higher carcass lipid contents than controls. In response to fasting, the decrements of the initial amount of both protein and fat were lower in MSG than in controls. Fasting induced a sustained increase in plasma free fatty acids only in the obese rats, although a single 100 μmol · l−1 dose of norepinephrine stimulated in vitro glycerol release more pronouncedly in epididymal adipocytes from control than obese rats. The results indicate that MSG-obese rats were able to mobilize fat stores during prolonged fasting. The high availability of lipid fuels and the sharp and sustained decrease in circulating corticosterone in the MSG group were probably important in diminishing body protein consumption during fasting.

Impairment of the serotonergic control of feeding in adult female rats exposed to intra-uterine malnutrition.

We have previously shown that adult female rats exposed to intra-uterine malnutrition were normophagic, although obese and resistant to insulin-induced hypophagia. The present study aimed at examining aspects of another important catabolic component of energy homeostasis control, the hypothalamic serotonergic function, which inhibits feeding and stimulates energy expenditure. Pregnant dams were fed ad libitum or were restricted to 50 % of ad libitum intake during the first 2 weeks of pregnancy. Control and restricted 4-month-old progeny were studied. The restricted rats had increased body adiposity with normal daily food intake but failed to respond with hypophagia to an intracerebroventricular injection of serotonin (5-hydroxytryptamine; 5-HT). Stimulation, by food ingestion, of extracellular levels of serotonin in medial hypothalamus microdialysates was more pronounced and lasted longer in the restricted than in the control rats. In the restricted group, hypothalamic levels of 5-HT 2C receptor protein tended to be reduced (P = 0.07) while the levels of 5-HT1B receptor and serotonin transporter proteins were significantly elevated (36 and 79 %, respectively). In conclusion, female rats undernourished in utero had normophagic obesity as adults but had an absence of serotonin-induced hypophagia and low hypothalamic levels of the 5-HT 2C receptor. Compensatory adaptations for the functional serotonergic impairment were evidenced, such as an enhanced release of serotonin in response to a meal allied to up-regulated hypothalamic 5-HT1B and transporter expression. Whether these compensations will persist in later life warrants further investigation. Moreover, it cannot be ruled out that the serotonergic component of energy expenditure was already impaired, thus contributing to the observed body-fat phenotype.

Deficit in hippocampal long-term potentiation in monosodium glutamate-treated rats

Rats subjected to monosodium glutamate (MSG) administration during the neonatal period present chronic neuroendocrine dysfunction associated with marked cognitive deficits. Long-term potentiation (LTP) in the hippocampus provides a model suited for the study of mammalian brain plasticity and memory formation. In the present work, we used the LTP protocol to investigate the synaptic plasticity in the hippocampal CA1 area of adult rats subjected to MSG treatment during the first 10 days of life. Synaptic transmission in CA1 area was analyzed using extracellular field recordings in response to Schaffers collateral fiber stimulation in hippocampal slices. Animals injected with MSG exhibited a dramatic decrement of LTP field excitatory postsynaptic potentials (fEPSPs) compared to control group. Analysis of percent enhancement of fEPSP slope at 2 min after high frequency stimulation (HFS) increased by 189.3 +/- 33.2% in slices from control rats and 129.45 +/- 18.5% (p < 0.01) in slices from MSG-treated rats. Additionally, MSG-treated animals failed to maintain or consolidate LTP as revealed by a significant reduction in fEPSP slope enhancement over time after HFS. The mean fEPSP slope, 60 min after HFS, was 154.28 +/- 21% of the average baseline slope in control slices versus only 124.4 +/- 15% in MSG-treated rats (p < 0.01). At 90 min after HFS, slices from controls reached a potentiation of 44.5 +/- 2.9%, whereas the MSG group displayed an overall response enhancement of 17.65 +/- 2.7% of basal levels (p < 0.01). These findings indicate that MSG-treated rats display a chronic impairment of CA1 synaptic plasticity.

Central administration of a nitric oxide precursor abolishes both the hypothalamic serotonin release and the hypophagia induced by interleukin-1β in obese Zucker rats

Serotonin-induced anorexia has long been recognized as an important part of the CNS mechanisms controlling energy balance. More recently, interleukin-1beta and nitric oxide have been suggested to influence this control, possibly through modulation of hypothalamic serotonin. The present work aimed at investigating the interaction of these systems. We addressed whether 5-HT is affected during IL-1beta-induced anorexia in obese Zucker rats and the influence of the central NO system on this IL-1beta/5-HT interaction. Using microdialysis, we observed that an intracerebroventricular injection of 10 ng IL-1beta significantly stimulated 5-HT extracellular levels in the VMH, with a peak variation of 102+/-41% above baseline. IL-1beta also significantly reduced the 4-h feeding by 33% and the 24-h feeding by 42%. Contrarily, these effects were absent when IL-1beta was injected 2 h after the i.c.v. administration of 20 microg of the NO precursor L-arginine. The results suggest that, in obese Zucker rats, activation of the serotonergic system in the medial hypothalamus participates in IL-1beta-induced anorexia. Since L-arginine, probably through NO stimulation, abolished both the anorexia and the serotonergic activation, it can be proposed that the NO system, either directly or indirectly, counteracts IL-1beta anorexia. The hypothalamic serotonergic system is likely to mediate this NO effect.

Evaluation of new leptin fragments on food intake and body weight of normal rats.

Leptin, a protein hormone originating from adipose tissue, circulates in the plasma and affects the energy balance by interacting with the hypothalamus. Leptin plays an important role in the regulation of a variety of physiological functions, including food intake, body temperature and body weight maintenance. Tertiary structure of the leptin molecule reveals the existence of a four-helix bundle that is characteristic of the short-helix cytokines. To identify regions of the leptin molecule responsible for its bioactivity, we have recently synthesized six peptides based on the protein three-dimensional structure. Our results indicated that the fragments Ac-hLEP(92-115)-NH(2) (IV) and Ac-[Ser(117)]-hLEP(116-140)-NH(2) (V) were recognized by leptin receptor present in hp-75 cells validating that this region of the molecule contain the functional epitope of the leptin molecule. In the present study, a new series of decapeptides encompassing the region of fragments IV and V of leptin were synthesized, and their effects on body weight and food intake were assessed when administered into the lateral cerbroventricle of normal rats. Peptides were synthesized by SPPS, purified by RP-HPLC and characterized by LC/ESI-MS. We also performed a conformational study of the peptides by circular dichroism in order to correlate the biological activity and secondary structure of the leptin fragments. Among the fragments tested, we found that Ac-hLEP(110-119)-NH(2) (VI) induce a significant reduction in both body weight and food intake. The use of synthetic leptin-derivate fragments may offer the basis for the development of compounds with potential application in human obesity or to its related metabolic dysfunctions.

Prolonged consumption of soy or fish-oil-enriched diets differentially affects the pattern of hypothalamic neuronal activation induced by refeeding in rats

Abstract We used c-Fos immunoreactivity to estimate neuronal activation in hypothalamic feeding-regulatory areas of 3-month-old rats fed control or oil-enriched diets (soy or fish) since weaning. While no diet effect was observed in c-Fos immunoreactivity of 24-h fasted animals, the acute response to refeeding was modified by both hyperlipidic diets but with different patterns. Upon refeeding, control-diet rats had significantly increased c-Fos immunoreactivity only in the paraventricular hypothalamic nucleus (PVH, 142%). In soy-diet rats, refeeding with the soy diet increased c-Fos immunoreactivity in dorsomedial hypothalamic nucleus (DMH, 271%) and lateral hypothalamic area (LH, 303%). Refeeding fish-diet rats with the fish diet increased c-Fos immunoreactivity in PVH (161%), DMH (177%), VMH (81%), and ARC (127%). Compared to the fish-diet, c-Fos immunoreactivity was increased in LH by the soy-diet while it was decreased in ventromedial hypothalamic nucleus (VMH) and arcuate hypothalamic nucleus (ARC). Based on the known roles of the activated nuclei, it is suggested that, unlike the fish-diet, the soy-diet induced a potentially obesogenic profile, with high LH and low VMH/PVH activation after refeeding.

Beneficial effects of Ginkgo biloba extract on insulin signaling cascade, dyslipidemia, and body adiposity of diet-induced obese rats

Ginkgo biloba extract (GbE) has been indicated as an efficient medicine for the treatment of diabetes mellitus type 2. It remains unclear if its effects are due to an improvement of the insulin signaling cascade, especially in obese subjects. The aim of the present study was to evaluate the effect of GbE on insulin tolerance, food intake, body adiposity, lipid profile, fasting insulin, and muscle levels of insulin receptor substrate 1 (IRS-1), protein tyrosine phosphatase 1B (PTP-1B), and protein kinase B (Akt), as well as Akt phosphorylation, in diet-induced obese rats. Rats were fed with a high-fat diet (HFD) or a normal fat diet (NFD) for 8 weeks. After that, the HFD group was divided into two groups: rats gavaged with a saline vehicle (HFD+V), and rats gavaged with 500 mg/kg of GbE diluted in the saline vehicle (HFD+Gb). NFD rats were gavaged with the saline vehicle only. At the end of the treatment, the rats were anesthetized, insulin was injected into the portal vein, and after 90s, the gastrocnemius muscle was removed. The quantification of IRS-1, Akt, and Akt phosphorylation was performed using Western blotting. Serum levels of fasting insulin and glucose, triacylglycerols and total cholesterol, and LDL and HDL fractions were measured. An insulin tolerance test was also performed. Ingestion of a hyperlipidic diet promoted loss of insulin sensitivity and also resulted in a significant increase in body adiposity, plasma triacylglycerol, and glucose levels. In addition, GbE treatment significantly reduced food intake and body adiposity while it protected against hyperglycemia and dyslipidemia in diet-induced obesity rats. It also enhanced insulin sensitivity in comparison to HFD+V rats, while it restored insulin-induced Akt phosphorylation, increased IRS-1, and reduced PTP-1B levels in gastrocnemius muscle. The present findings suggest that G. biloba might be efficient in preventing and treating obesity-induced insulin signaling impairment.

Central but not peripheral glucoprivation is impaired in monosodium glutamate-treated rats

In the present study, newborn male Wistar rats were injected, subcutaneously, five times, every other day, with monosodium glutamate (MSG, 4 g/kg bw) or saline (as control, C), during the neonatal period. MSG animals developed destruction of the arcuate nuclei (ARC) with absence of NPY-immunoreactive cell bodies, which impaired both the food intake (baseline) and the 2-deoxy-D-glucose (2DG) glucoprivic feeding response. Increases in the immunoreactivity of corticotropin-releasing hormone-cell bodies in the paraventricular nuclei might have developed to compensate for the atrophy of the pituitary in MSG-treated rats. After systemic 2DG injection, neither the C nor the MSG rats increased their food intake, but they showed similar hyperglycemic responses, whereas plasma free fatty acids (FFA) increased only in the C group. In other groups, 2DG, norepinephrine (NE), neostigmine (NEO) and saline were intracerebroventricularly (i.c.v.) administered. In this condition, impairment of the hyperglycemic and food intake responses, associated to a lower increase in plasma FFA levels, were observed. As opposed to this, the MSG treatment gives support to NE effects, enhancing food intake, as well as plasma glucose and FFA levels. After NEO, plasma glucose increased only in the MSG group, while plasma FFA levels were elevated in the C rats. Taken together, the results obtained after MSG treatment point to a separate neural control of the hyperglycemic response and of the lipid mobilization when stimulated by central 2DG, NE or NEO administration. It seems likely that the excitatory neural pathway that controls lipid metabolism and is present in C rats was destroyed by the MSG treatment.