Thomas J. Lauterio

Development of Hypertension in a Rat Model of Diet-Induced Obesity

Although obesity is a risk factor for hypertension, the relationship between these 2 conditions is not well understood. Therefore, we examined some parameters of hypertension and cardiovascular disease in a dietary model of obesity. Male Sprague-Dawley rats were provided either a control diet (C) or a diet containing 32% kcal as fat (similar to a Western diet) for 1, 3, or 10 weeks. Rats in the latter group diverged based on body weight gain into obesity-prone (OP) and obesity-resistant (OR) groups. Systolic blood pressure in OP rats was significantly higher after 10 weeks of the diet (149+/-4. 8 mm Hg) compared with both OR and C groups (131+/-3.7 and 129+/-4.5 mm Hg, respectively). The aortic wall area of OP rats was significantly increased, indicating arterial hypertrophy, and a 2-fold increase in plasma renin activity was found in OP rats compared with OR and C rats. The lipid profile showed a significant increase in plasma and VLDL triglycerides of OP versus OR and C groups as early as 3 weeks on the diet. Plasma and LDL-cholesterol levels were increased in the OP group versus the OR and C groups after 3 weeks of the diet, but the difference was blunted after 10 weeks. Lipid peroxidation (thiobarbituric acid-reactive substances) in OP rats was increased 2-fold in LDL and 1.5-fold in aortic wall compared with OR rats, suggesting an increased oxidative stress in these animals. Periodic acid-Schiff staining of the kidney showed mesangial expansion and focal sclerosis that were more prominent in OP rats than in OR rats. The results suggest that hypercholesterolemia, but not hypertriglyceridemia, is linked to the diet; that hypertension and renin-angiotensin system activation are associated with obesity; and that lipid peroxidation and renal damage are the results of both factors.

Oxidative Stress in a Rat Model of Obesity-Induced Hypertension

The mechanisms underlying the development of hypertension in obesity are not yet fully understood. We recently reported the development of hypertension in a rat model of diet-induced obesity. When Sprague-Dawley rats (n=60) are fed a moderately high fat diet (32 kcal% fat) for 10 to 16 weeks, approximately half of them develop obesity (obesity-prone [OP] group) and mild hypertension (158±3.4 mm Hg systolic pressure), whereas the other half (obesity-resistant [OR] group) maintains a body weight equivalent to that of a low fat control group and is normotensive (135.8±3.8 mm Hg). We examined the potential role of oxidative stress in the development of hypertension in this model. Lipid peroxides measured as thiobarbituric acid–reactive substances showed a significant increase in the LDL fraction of OP rats (2.8±0.32 nmol malondialdehyde/mg protein) compared with OR and control rats (0.9±0.3 nmol malondialdehyde/mg protein). Also, aortic and kidney thiobarbituric acid–reactive substances showed a significant (3- and 5- fold) increase in OP rats after 16 weeks of diet. In addition, superoxide generation by aortic rings, measured by lucigenin luminescence, showed a 2-fold increase in the OP group compared with both the OR and control groups. In addition, free isoprostane excretion and nitrotyrosine in the kidney showed an increase in OP rats only. The urine and plasma nitrate/nitrite measured by the LDH method showed a 1.8-fold decrease in OP rats compared with OR rats. However, endothelial NO synthase expression in the kidney cortex and medulla assessed by reverse transcriptase–polymerase chain reaction showed a strong increase in the OP rats versus OR and control rats (endothelial NO synthase/&bgr;-actin ratio 1.3±0.04 in OP rats versus 0.44±0.02 in OR rats), suggesting a possible shift toward superoxide production by the enzyme. Collectively, the data show a decreased NO bioavailability in OP animals that is due in part to the increased oxidative stress.

β-endorphin and dynorphin abnormalities in rats subjected to exercise and restricted feeding : relationship to anorexia nervosa ?

Exercise and the endogenous opioids have been linked to anorexia nervosa. This investigation determined the effects of the weight-loss syndrome induced by voluntary exercise (22.5 h/day) in food-restricted rats (1.5 h/day food access) on the endogenous opioids. The animals were tested under resting-fed and 2-deoxy-D-glucose (2DG) stimulated conditions. Weight-matched, freely fed exercised and ad libitum fed unexercised groups served as controls. Specific opioid abnormalities were found in the syndrome. These included a basal elevation in plasma beta-endorphin, which was abnormally suppressed by 2DG, and 2DG-induced elevations in arcuate hypothalamic beta-endorphin content and supraoptic hypothalamic dynorphin-A content. None of these changes occurred in controls. Finally, it was found that short-term moderate exercise itself chronically reduced adenohypophysial beta-endorphin content and elevated supraoptic dynorphin-A content. The relationship of the syndromes hyperendorphinism to the hypothalamo-pituitary-adrenal axis and the auto-addiction hypothesis of anorexia nervosa was considered, as was the significance of the supraoptic dynorphin-A abnormality to the hypothalamo-neurohypophysial system. The differential sensitivity of the supraoptic dynorphin-A system compared to the arcuate hypothalamic beta-endorphin system to moderate exercise was also discussed.

Plasma growth hormone secretion is impaired in obesity-prone rats before onset of diet-induced obesity

Sprague-Dawley rats, which become obese (obesity prone) when fed a moderately high-fat (MHF; 32.5% of kcal as fat) diet, have decreased growth hormone (GH) concentrations compared with obesity-resistant rats fed the same diet. To determine whether plasma GH concentrations are different in obesity-prone rats compared with obesity-resistant rats before diet-induced obesity occurs, total integrated GH concentrations were determined in male Sprague-Dawley rats before exposure to the MHF diet. After initial blood sampling, rats were fed an MHF diet for 15 wk, over which time the animals were separated into two discrete populations based on body weight gain. Analysis of GH in episodic blood samples showed that the obesity-prone group had a GH secretion deficit before the onset of obesity (115.2 +/- 12.9 ng . ml-1 . 200 min-1) compared with obesity-resistant rats (237.2 +/- 47.1 ng . ml-1 . 200 min-1). The GH concentration difference was due to a decrease in mean GH peak height in rats that later became obese (34.8 ng/ml) compared with rats that remained lean (74.2 ng/ml). The results suggest that GH secretion impairment exists before dietary challenge or onset of obesity and may contribute to the susceptibility to obesity observed in these animals.Sprague-Dawley rats, which become obese (obesity prone) when fed a moderately high-fat (MHF; 32.5% of kcal as fat) diet, have decreased growth hormone (GH) concentrations compared with obesity-resistant rats fed the same diet. To determine whether plasma GH concentrations are different in obesity-prone rats compared with obesity-resistant rats before diet-induced obesity occurs, total integrated GH concentrations were determined in male Sprague-Dawley rats before exposure to the MHF diet. After initial blood sampling, rats were fed an MHF diet for 15 wk, over which time the animals were separated into two discrete populations based on body weight gain. Analysis of GH in episodic blood samples showed that the obesity-prone group had a GH secretion deficit before the onset of obesity (115.2 ± 12.9 ng ⋅ ml-1 ⋅ 200 min-1) compared with obesity-resistant rats (237.2 ± 47.1 ng ⋅ ml-1 ⋅ 200 min-1). The GH concentration difference was due to a decrease in mean GH peak height in rats that later became obese (34.8 ng/ml) compared with rats that remained lean (74.2 ng/ml). The results suggest that GH secretion impairment exists before dietary challenge or onset of obesity and may contribute to the susceptibility to obesity observed in these animals.

Fluoxetine induces vasopressin and oxytocin abnormalities in food-restricted rats given voluntary exercise: relationship to anorexia nervosa

Anorexia nervosa is associated with vasopressin, oxytocin and serotonin abnormalities. Because of the relationship between exercise and anorexia nervosa, we explored the weight-loss syndrome produced by wheel running in food-deprived rats. Its effects on regional vasopressin and oxytocin concentrations were determined under basal conditions and following systemic fluoxetine. Weight-matched, exercised and unexercised rats served as controls. Fluoxetine caused abnormalities in suprachiasmatic vasopressin and dynorphin A content and in thymus oxytocin content that did not occur in weight-matched or exercised controls. No syndrome-specific anomalies occurred in the hypothalamo-neurohypophysial system or dorsal vagal complex (DVC). However, weight reduction and fluoxetine increased circulating vasopressin; moderate exercise caused fluoxetine-induced elevations in posterior pituitary vasopressin and oxytocin; and, unlike the other groups, fluoxetine increased DVC oxytocin in freely fed unexercised rats. It was concluded that syndrome-specific vasopressin and oxytocin abnormalities occur that are not secondary to weight loss or moderate exercise; that weight loss or fluoxetine increases circulating vasopressin; that moderate exercise alters neurohypophysial vasopressin and oxytocin content; and that weight loss or exercise inhibits a fluoxetine-stimulated increase in DVC oxytocin. Finally, it was argued that the fluoxetine abnormalities indicate possible serotonin dysfunction in the syndrome.

Intracerebroventricular injection of insulin or glucose alters insulin-like growth factor II (IGF-II) concentrations in specific hypothalamic nuclei

Peripherally administered insulin has been shown to alter content and gene expression of hypothalamic insulin-like growth factor II (IGF-II) in a region specific manner (Lauterio, TJ. et al., Endocrinology, 126 (1990) 392-398. The objective of this experiment was to determine whether central administration of insulin can modulate hypothalamic IGF-II peptide content. Male Sprague-Dawley rats were implanted with lateral ventricular cannulae and allowed to recover from surgery one week prior to injection. At that point animals were remotely injected with one of the following: (1) synthetic cerebral spinal fluid vehicle (sCSF); (2) 2 mIU porcine insulin (I); (3) glucose (100 mg%) only. Animals were decapitated 30 min after injections and brains were quickly removed, frozen and dissected into specific hypothalamic regions for IGF-II analysis by RIA. Insulin increased IGF-II content in the ventromedial hypothalamic region by 80% (P < 0.001) and paraventricular nucleus by 30% (P < 0.01) compared to sCSF or glucose treatment. Arcuate nucleus and neurointermediary lobe pituitary IGF-II content was decreased with insulin treatment compared to controls (P < 0.01). Insulin had no effect on IGF-II concentrations in the dorsomedial or lateral hypothalamic regions or in the supraoptic and suprachiasmatic nuclei. Peripheral concentrations of glucose, insulin and IGF-II were unaffected by any treatment. Results show that insulin which reaches the brain can alter IGF-II levels in specific regions of the hypothalamus and suggests a possible role for IGF-II in insulin mediated changes in metabolism or hypothalamic hormone secretion.

The Effects of IGF-I and IGF-II on Cell Growth and Differentiation in the Central Nervous System

Both IGF-I and IGF-II peptides have been localized to specific brain regions. The distribution of IGF-I is homogeneous whereas IGF-II appears to be more local. Two species of IGF receptors are found in the CNS. The type II (m6P) is similar to that in the periphery, but the type I has nearly the same affinity for IGF-I and IGF-II. IGF-I has now been shown to provide cell growth and survival as well as stimulate neurite outgrowth. Dorsal root ganglia and sympathetic neurons are sensitive to IGF-II and the action may be additive with NGF. Cells other than neurites, such as oligodendrocytes respond to the IGFs as well as primary and transformed lines. The mechanism of action has not been resolved but IDG-II appears to act via G-protein coupled activation of protein kinase C. Interaction between various growth factors and the IGFs may be due to up or down-regulation of the receptor predicated by the non-homologous peptide.

Regulation and Physiological Function of Insulin-Like Growth Factors in the Central Nervous System

The function of insulin-like growth factors in the central nervous system (CNS) and the physiological regulators of brain and pituitary IGF synthesis has been studied from many perspectives. Until recently, these efforts have not met with great success. However as more laboratories obtain the means to study IGF mRNA and peptide synthesis the multifaceted approach of investigation has provided new insights on growth factor metabolism.

Fluoxetine induced insulin-like growth factor II (IGF-II) changes in hypothalami of normal, exercised and food restricted rats

Hypothalamic and pituitary insulin-like growth factor II (IGF-II) peptide concentrations are differentially regulated by factors associated with metabolism such as insulin and glucoprivation. However, the effects of other metabolic stressors such as food restriction or exercise on hypothalamic IGF-II concentrations remain largely to be explored. In order to assess whether metabolic stress alters central nervous system IGF-II secretion, peptide analysis was conducted in rats exhibiting activity-based anorexia (ABA) compared to exercised-matched, body weight-matched or ad libitum fed controls. Further, the possibility of serotonergic control of IGF-II secretion was examined by determining IGF-II response to fluoxetine (FLX) injections (15 mg/kg body wt., i.p.). While ABA and body weight loss altered peripheral IGF-II concentrations compared to ad libitum fed or exercised controls, these treatments had no effect on hypothalamic or posterior pituitary IGF-II content. However, FLX administration increased IGF-II concentrations in the ventromedial hypothalamus and decreased IGF-II content in the lateral hypothalamus compared to vehicle injected. Anterior pituitary levels of IGF-II were also decreased by FLX. These data suggest that a serotonergic influence on CNS IGF-II exists and that IGF-II secretion may be altered by factors affecting serotonin metabolism or efficacy.

Effects of insulin and 2-deoxy-D-glucose administration on CNS IGF-II content

In order to determine whether effects of insulin on central nervous system (CNS) insulin-like growth factor II (IGF-II) content are direct or whether they are due to insulin-induced glucoprivation, short-term (2 h) time course studies were conducted utilizing the glucoprivic agent 2-deoxy-D-glucose (2-DG). Insulin (2 U/kg b.wt.), 2-DG (500 mg/kg b.wt.) or vehicle were administered to Sprague-Dawley rats (350-450 kg) and groups of animals were killed at time 0 and 15, 30, 45, 60, and 120 min following injection. Specific hypothalamic and hindbrain regions obtained by microdissection, were analyzed for IGF-II content by RIA. Insulin and 2-DG exerted similar effects on IGF-II content in the dorsomedial hypothalamus and the paraventricular nucleus. These data suggest that IGF-II was primarily regulated by glucoprivation in these regions. Only 2-DG altered ventromedial hypothalamic IGF-II content, and opposite responses to the two agents were observed in the arcuate nucleus. This uncoupling of IGF-II response suggests that differences in peripheral vs. central signals for IGF-II secretion may be involved. The vagal complex was responsive only to insulin injection indicating a specific response that may be tied to sensory vagal function.