Leptin resistance before and after obesity: evidence that tissue glucose uptake underlies adipocyte enlargement and liver steatosis/steatohepatitis in Zucker rats from early-life stages.

BACKGROUND: Leptin resistance occurs in obese patients, but its independent contribution to adiposity and the accompanying metabolic diseases, i.e., diabetes, liver steatosis, and steatohepatitis, remains to be established. This study was conducted in an extreme model of leptin resistance to investigate mechanisms initiating diabetes, fat expansion, liver steatosis, and inflammatory disease, focusing on the involvement of glucose intolerance and organ-specific glucose uptake in brown and subcutaneous adipose tissues (BAT, SAT) and in the liver. METHODS: We studied preobese and adult Zucker rats (fa/fa, fa/+ ) during fasting or glucose loading to assess glucose tolerance. Relevant pancreatic and intestinal hormonal levels were measured by Milliplex. Imaging of 18F-fluorodeoxyglucose by positron emission tomography was used to quantify glucose uptake in SAT, BAT, and liver, and evaluate its relationship with adipocyte size and biopsy-proven nonalcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH). RESULTS: Preobese fa/fa pups showed impaired glucose tolerance, adipocyte enlargement, hepatic microsteatosis, and lobular inflammation, with elevated hepatic post-glucose load glucose uptake and production. Adult fa/fa rats had more severe glucose intolerance, fasting hyperglycemia, hormonal abnormalities, elevated glucose uptake in SAT and BAT, and more markedly in the liver, together with macrosteatosis, and highly prevalent hepatic inflammation. Organ glucose uptake was proportional to the degree of fat accumulation and tissue inflammation and was able to dissect healthy from NAFLD and NAFLD/NASH livers. Most severe NASH livers showed a decline in glucose uptake and liver enzymes. CONCLUSIONS: In fa/fa Zucker rats, leptin resistance leads to glucose intolerance, mainly due to hepatic glucose overproduction, preceding obesity, and explaining pancreatic and intestinal hormonal changes and fat accumulation in adipocytes and hepatocytes. Our data support the involvement of liver glucose uptake in the pathogenesis of liver inflammatory disease. Its potential as more generalized biomarker or diagnostic approach remains to be established outside of our leptin-receptor-deficient rat model.

Evaluation of the Zucker diabetic fatty (ZDF) rat as a model for human disease based on urinary peptidomic profiles.

Representative animal models for diabetes-associated vascular complications are extremely relevant in assessing potential therapeutic drugs. While several rodent models for type 2 diabetes (T2D) are available, their relevance in recapitulating renal and cardiovascular features of diabetes in man is not entirely clear. Here we evaluate at the molecular level the similarity between Zucker diabetic fatty (ZDF) rats, as a model of T2D-associated vascular complications, and human disease by urinary proteome analysis. Urine analysis of ZDF rats at early and late stages of disease compared to age- matched LEAN rats identified 180 peptides as potentially associated with diabetes complications. Overlaps with human chronic kidney disease (CKD) and cardiovascular disease (CVD) biomarkers were observed, corresponding to proteins marking kidney damage (eg albumin, alpha-1 antitrypsin) or related to disease development (collagen). Concordance in regulation of these peptides in rats versus humans was more pronounced in the CVD compared to the CKD panels. In addition, disease-associated predicted protease activities in ZDF rats showed higher similarities to the predicted activities in human CVD. Based on urinary peptidomic analysis, the ZDF rat model displays similarity to human CVD but might not be the most appropriate model to display human CKD on a molecular level.

Effects of a diabetes-specific enteral nutrition on nutritional and immune status of diabetic, obese, and endotoxemic rats: interest of a graded arginine supply.

OBJECTIVE: Obese and type 2 diabetic patients present metabolic disturbance-related alterations in nonspecific immunity, to which the decrease in their plasma arginine contributes. Although diabetes-specific formulas have been developed, they have never been tested in the context of an acute infectious situation as can be seen in intensive care unit patients. Our aim was to investigate the effects of a diabetes-specific diet enriched or not with arginine in a model of infectious stress in a diabetes and obesity situation. As a large intake of arginine may be deleterious, this amino acid was given in graded fashion. DESIGN: Randomized, controlled experimental study. SETTING: University research laboratory. SUBJECTS: Zucker diabetic fatty rats. INTERVENTIONS: Gastrostomized Zucker diabetic fatty rats were submitted to intraperitoneal lipopolysaccharide administration and fed for 7 days with either a diabetes-specific enteral nutrition without (G group, n=7) or with graded arginine supply (1-5 g/kg/day) (GA group, n=7) or a standard enteral nutrition (HP group, n=10). MEASUREMENTS AND MAIN RESULTS: Survival rate was better in G and GA groups than in the HP group. On day 7, plasma insulin to glucose ratio tended to be lower in the same G and GA groups. Macrophage tumor necrosis factor-α (G: 5.0±1.1 ng/2×106 cells·hr⁻¹; GA: 3.7±0.8 ng/2×106 cells·hr⁻¹; and HP: 1.7±0.6 ng/2×106 cells·hr⁻¹; p<.05 G vs. HP) and nitric oxide (G: 4.5±1.1 ng/2×106 cells·hr⁻¹; GA: 5.1±1.0 ng/2×106 cells·hr⁻¹; and HP: 1.0±0.5 nmol/2×106 cells·hr⁻¹; p<.05 G and GA vs. HP) productions were higher in the G and GA groups compared to the HP group. Macrophages from the G and GA groups exhibited increased arginine consumption. CONCLUSIONS: In diabetic obese and endotoxemic rats, a diabetes-specific formula leads to a lower mortality, a decreased insulin resistance, and an improvement in peritoneal macrophage function. Arginine supplementation has no additional effect. These data support the use of such disease-specific diets in critically ill diabetic and obese patients.

Bromocriptine administration reduces hyperphagia and adiposity and differentially affects dopamine D2 receptor and transporter binding in leptin-receptor-deficient Zucker rats and rats with diet-induced obesity.

BACKGROUND: The dopamine (DA) D(2) receptor (D2R) agonist bromocriptine (BC) decreases body fat in animal and human models and increases lean muscle mass, improves glucose intolerance and insulin resistance, and reduces triglycerides and free fatty acids. We have previously shown a negative correlation between D2R and body weight in obese individuals and in rodents, and that chronic food restriction increases D2R binding in genetically obese rats. The purpose of this study was to assess whether the antiobesity and metabolic effects of BC are related to changes in midbrain DA and D2R activity by measuring D2R and DA transporter (DAT) binding in a genetic (leptin-receptor-deficient) and environmental (diet-induced) rodent obesity model. METHODS: Obese (fa/fa) (leptin-receptor-deficient), lean (FA/FA) Zucker rats and rats with diet-induced obesity (DIO) were treated with 10 mg/kg BC for 4 weeks. Body weight, food intake, locomotor activity and blood glucose levels were measured along with D2R- and DAT-binding levels using in vitro receptor autoradiography. RESULTS: BC decreased food intake and body fat and increased locomotor activity in both the (fa/fa) and DIO rats. Furthermore, BC increased D2R binding in (fa/fa) but not in DIO rats. Finally, BC increased DAT binding in DIO rats but not in the (fa/fa) rats. CONCLUSION: These observations are all consistent with the existence of unique leptin-DA interactions and the hypothesis that there is hyposensitivity of the DA system in obesity.

Differences in the pharmacokinetics of peroxisome proliferator-activated receptor agonists in genetically obese Zucker and sprague-dawley rats: implications of decreased glucuronidation in obese Zucker rats.

Genetically obese Zucker rats exhibit symptoms similar to those of obese patients with insulin-resistance or Type II diabetes; therefore, they have been used as a genetic model to study obesity, as well as a pharmacological model for the discovery of new drugs for the treatment of Type II diabetes and hyperlipidemia. In the present study, we compared the pharmacokinetics of two novel peroxisome proliferator-activated receptor (PPAR) agonists, MRL-I [(2R)-7-[3-[2-chloro-4-(4-fluorophenoxy)phenoxy]propoxy]-2-ethyl-3,4-dihydro-2H-benzopyran-2-carboxylic acid] and MRL-II [(2R)-7-[3-[2-chloro-4-(2,2,2-trifluoroethoxy)phenoxy]propoxy]-3,4-dihydro-2-methyl-2H-benzopyran-2-carboxylic acid], in obese Zucker and lean Sprague-Dawley rats following a single intravenous administration. The plasma clearance of both MRL-I and MRL-II was significantly lower in obese Zucker rats (4- and 2-fold, respectively) compared with Sprague-Dawley rats, but without any significant change in the volume of distribution, which resulted in a dramatic increase in the half-life (7- and 3-fold, respectively). The reversible in vitro plasma protein binding of [(14)C]MRL-I and [(14)C]MRL-II was comparable in the two strains, approximately 96% bound. The expression levels of uridine diphosphate-glucuronosyltransferases 1A1, 1A6, 2B1, and CYP2C11 and 3A1 mRNA in liver were lower (30-50%) in Zucker compared with Sprague-Dawley rats, as were the liver glutathione S-transferases (70%), quinone reductase (30%), organic anion-transporting protein 2 (80%), and multidrug resistance-associated protein 2 (Mrp2) (50%) mRNA levels. However, Mrp3 mRNA levels were similar in both strains. Consistent with these observations, the intrinsic clearance (CL(int)), calculated from the V(max)/K(m) of glucuronidation of [(14)C]MRL-I and [(14)C]MRL-II in liver microsomes, was approximately 2-fold lower in obese Zucker rats; the K(m) values were comparable in the two strains for both compounds. In conclusion, differences in the pharmacokinetics of two novel PPAR agonists, both cleared, predominantly, by conjugation, were evident in genetically obese Zucker rats compared with Sprague-Dawley rats. These differences were consistent with changes in the mRNA levels of hepatic drug-metabolizing enzymes and transporters. This information should be considered when comparing pharmacokinetic and efficacious doses in the obese Zucker rats, used as a pharmacological model, with those in Sprague-Dawley rats, which are used widely for drug metabolism and toxicology studies.

Leptin-receptor gene transfer into the arcuate nucleus of female Fatty zucker rats using recombinant adeno-associated viral vectors stimulates the hypothalamo-pituitary-gonadal axis.

Fatty fa/fa Zucker rats with a missense mutation in the leptin receptor (OB-R) are obese and infertile with prolonged estrous cycles. To determine whether their reproductive deficits could be corrected by OB-R installation, we employed viral vectors to introduce the OB-R gene into either the arcuate nucleus (ARC) or the paraventricular nucleus (PVN) of the hypothalamus, sites of OB-R expression in wild-type rats. Recombinant adeno-associated viral (rAAV) vectors encoding the human leptin-receptor gene (rAAV-OB-Rb) were microinjected intraparenchymally to produce doxycycline-regulatable OB-R gene expression. Expression of the OB-R gene in the ARC and PVN was verified using reverse transcription-polymerase chain reaction. Expression of OB-R in the ARC, but not in the PVN, resulted in normalization of estrous cycle length, increased ovarian follicular development, and decreased serum progesterone levels. Compared to saline-injected rats, hypothalamic expression of neuropeptide Y (NPY) and pro-opiomelanocortin were decreased in ARC rAAV-OB-Rb-injected rats. Parallel decreases were noted in NPY and beta-endorphin (beta-END) concentrations in the hypothalamus, whereas luteinizing hormone-releasing hormone (LHRH) levels increased. These studies showed that rAAV vectors can be successfully used to install functional OB-R in the hypothalamus for extended periods. The resultant stimulation of the hypothalamo-pituitary-gonadal (HPG) axis in ARC-injected rats was probably brought about by the observed decreases in NPY and beta-END, which inhibit hypothalamic LHRH. Because these changes were seen in ARC-injected, but not in PVN-injected, rats, the results suggest that the ARC may be the primary site where leptin acts to regulate the HPG axis.

Strain variability in Zucker rats affects their response to oral oleoyl-estrone.

There is a considerable variability in the responses of Zucker fa/fa rats in metabolic studies, which could not be solely attributed to the leprfa mutation. In order to fathom the extent of this variability, we compared the response to oleoyl-estrone (OE), a powerful lipid-mobilising agent, of two strains of Zucker lean and obese rats: Harlan (H) and Charles River (CR). Rats were given an oral gavage of 10 micromol/day/kg of OE in sunflower oil, and were compared with oil-receiving controls. Body composition, energy and water balances, and plasma parameters were studied after 10 days of treatment. H rats showed a higher water turnover than CR rats; OE treatment reduced water intake, partly compensated by metabolic water, and decreased stool water. H rats accrued more cholesterol than CR animals, which showed higher cholesterolaemia. OE facilitated cholesterol disposal in lean (CR and H) and H obese rats. CR rats had higher body and liver lipids than H animals. No differences in energy balance were found. Insulin decrease following OE treatment was greater in lean CR than in H rats, but this trend was reversed in the obese rats, lacking effective responses to leptin. The red cell glucose compartment was smaller in H than in CR rats; the higher insulin levels in H rats may be partly responsible for that difference. Obese H maintained glycaemia (and liver glycogen) with higher insulin levels than CR animals. The extent to which the leprfa mutation affects the responses of Zucker fa/fa rats could not be singled out unless the metabolic environment of the batch used is known. This variability must be taken into account when developing a metabolic or hormonal study in which this model of obesity is used.

The rat arcuate nucleus integrates peripheral signals provided by leptin, insulin, and a ghrelin mimetic.

The hypothalamic circuits controlling food intake and body weight receive and integrate information from circulating satiety signals such as leptin and insulin and also from ghrelin, the only known circulating hormone that stimulates appetite following systemic injection. Activation of arcuate neurons by ghrelin and ghrelin mimetics (the growth hormone secretagogues) is augmented in 48-h-fasted rats compared with fed rats, as reflected by a greater number of cells expressing Fos protein in response to administration of the same maximally effective dose. Here we sought to determine whether this increased responsiveness in fasting might reflect or be influenced by low levels of circulating satiety factors such as leptin or insulin. Chronic central infusion of insulin or leptin during a 48-h fast suppressed the threefold increase in the Fos response to intravenous injection of a maximally effective dose of growth hormone-releasing peptide (GHRP)-6, a synthetic growth hormone secretagogue. This appears to be a direct central action of insulin and leptin because the marked decrease in plasma levels of insulin, leptin, and glucose during fasting were unaffected by central administration of either hormone. Furthermore, the GHRP-6-induced Fos response was twofold greater in obese leptin- and insulin-resistant Zucker rats compared with lean controls. These data provide evidence that the ghrelin-sensitive circuits in the hypothalamus are dynamically regulated by central insulin and leptin action.

Evidence for increased functional vascular Na+/K+ pump activity in the obese Zucker rat.

Insulin is known to stimulate Na+/K+ ATPase and to relax vascular smooth muscle. We hypothesized that vascular tone in the obese Zucker (fa/fa) rat, a hyperinsulinemic model in which hypertension can develop, may be influenced by insulin's ability to stimulate Na+/K+ ATPase at the vascular level. We studied isometric preparations of tail and femoral arteries from 10-wk-old, male obese Zucker rats, which were hyperinsulinemic but still normotensive vs. lean controls. Sensitivity to potassium-induced relaxations, an index of vascular Na+/K+ ATPase activity was significantly greater in the obese Zucker rat than control. Sensitivity to transmural-nerve-stimulation-induced contractions was decreased in the femoral and tail arteries from obese rats as compared with lean controls. Insulin (50 to 200 mU/ml) mimicked potassium-induced relaxations in the femoral artery, an effect that was significantly greater in the obese group. These data suggest that in the young hyperinsulinemic Zucker rat, insulin has a stimulatory effect on the vascular Na+/K+ pump, which may be associated with a decreased presynaptic adrenergic influence on vascular tone. Development of resistance to these vascular relaxant effects of insulin with advancing age might contribute to the onset of hypertension in this model.

Anti-TNF treatment does not reverse the abnormalities in lipid metabolism of the obese Zucker rat.

Because obesity, insulin resistance, and hyperlipidemia are often associated, and recent evidence suggests that the cytokine tumor necrosis factor-alpha (TNF) may influence the activity of insulin in various target tissues, the present study was designed to see whether TNF was also associated with the changes in lipid metabolism that lead to hyperlipidemia in the obese model of the Zucker rat. A polyclonal goat anti-rat TNF antibody was subcutaneously administered to Zucker rats for 4 days to block TNF actions. The results indicate that none of the alterations in lipid metabolism seen in the obese animals were reversed by the anti-TNF treatment. This was the case for the lipogenic rate in liver and adipose tissue, the disposal of an exogenous [14C]triolein load, adipose tissue lipoprotein lipase activity, and the hypertriglyceridemia. Measurements of lipolysis in adipose tissue slices from the anti-TNF-treated animals also did not show any significant effect of the treatment. In conclusion, TNF does not seem to be involved in the abnormalities of lipid metabolism observed in the obese Zucker rat.

Differential regulation of Na(+)-K(+)-ATPase in the obese Zucker rat.

Expression of Na(+)-K(+)-adenosinetriphosphatase (ATPase) in tissues from obese and lean Zucker rats was monitored. The phosphatase activity of the sodium pump was increased in liver and intestinal mucosa from obese animals but was unaltered in skeletal muscle, brown adipose tissue, kidney, and heart. Induction of Na(+)-K(+)-ATPase activity was correlated with increased alpha 1-subunit protein amounts in liver and intestinal mucosa, although alpha 1-subunit mRNA levels were increased only in liver tissue. Neither protein nor mRNA amounts for both subunits were significantly altered in the other tissues analyzed. The only exception was a decrease in the amount of beta 1-protein in kidney from obese rats. alpha 2-Subunit protein and alpha 2- and beta 2-mRNA levels were not altered in brown adipose tissue, heart, and soleus. In summary, this study shows that in obese Zucker rats the expression of the sodium pump is enhanced in tissues that are directly involved in nutrient uptake and processing. This adaptation may be related to the ongoing hyperphagia and to tissue hypertrophia but develops in a different manner in each tissue, suggesting differential regulation of alpha 1-subunit expression.

Evidence for regulated coupling of A1 adenosine receptors by phosphorylation in Zucker rats.

Studies were designed to find the molecular basis for previous observations that lipolysis is less active and A1 adenosine receptor signaling is more active in adipocytes from obese than from lean Zucker rats. With quantitative immunoblot procedures for detection, Gi alpha 1 and Gs alpha 45 levels were found anomalously low in obese compared with lean membranes (50 and 30%, respectively), but other G alpha subunit levels were normal. However, the sensitivity of the receptor-Gi protein to GTP was about 5- to 10-fold higher in obese compared with lean membranes when assessed from 1) the ability of GTP to inhibit forskolin-stimulated adenylyl cyclase in the presence of an adenosine receptor agonist and 2) the ability of a nonhydrolyzable guanine nucleotide analogue to alter A1 adenosine receptor agonist binding. Alkaline phosphatase treatment of isolated adipocyte membranes from obese but not lean animals decreased guanine nucleotide sensitivity of agonist binding. Surprisingly, solubilized adipocyte A1 adenosine receptors from all animals exhibited the same high sensitivity to guanine nucleotides as that of intact obese membranes, and this high sensitivity could be decreased 20-fold by treatment with alkaline phosphatase. These data suggest that protein phosphorylation may regulate coupling of the A1 adenosine receptor in rat adipocyte membranes.

Muscle bioenergetics in obese Zucker rats.

The purpose of this study was to investigate the energetic metabolism in obese Zucker rats, using phosphorus nuclear magnetic resonance spectroscopy at rest and during a 2-Hz muscle stimulation and subsequent recovery. Animals were anesthetized with ketamine (150 mg/kg ip). Fed obese rats and 2-day-fasted obese rats were compared with their normally fed and 2-day-fasted lean litter mates. No differences were found between the two groups for ATP, total creatine, phosphocreatine (PCr), and intracellular pH. Starvation in lean rats resulted in a significant fall in inorganic phosphate (Pi), increased resting ADP level, and decreased PCr and ADP recovery after stimulation. The obese rats exhibited a decreased PCr/Pi and increased ADP at rest and a decreased PCr resynthesis and ADP metabolization rate after stimulation. Muscle stimulation in fasted obese rats induced higher PCr depletion and more pronounced acidosis. These results suggest an in vivo mitochondrial metabolism dysfunction in fasted lean as well as in fed and fasted obese rats.

Insulin-induced translocation of GLUT 4 in skeletal muscle of insulin-resistant Zucker rats.

The genetically obese Zucker rat (fa/fa) is an animal model with severe insulin resistance of the skeletal muscle. We investigated whether a defect of insulin-dependent glucose transporter (GLUT 4) translocation might contribute to the pathogenesis of the insulin-resistant state. fa/fa rats, lean controls (Fa/Fa) as well as normal Wistar rats were injected intraperitoneally with insulin and were killed after 2 or 20 min, respectively. Subcellular fractions were prepared from hind-limb skeletal muscle and were characterized by determination of marker-enzyme activities and immunoblotting applying antibodies against alpha 1 Na+/K+ ATPase. The relative amounts of GLUT 1 and GLUT 4 were determined in the fractions by immunoblotting with the respective antibodies. Insulin induced an approximately two-fold increase of GLUT 4 in a plasma membrane and transverse tubule enriched fraction and a decrease in the low density enriched membrane fraction in all three groups of rats. There was a high individual variation in GLUT 4 translocation efficiency within the groups. However, no statistically significant difference was noted between the groups. No effect of insulin was detectable on the distribution of GLUT 1 or alpha 1 Na+K+ ATPase. The data suggest that skeletal muscle insulin resistance of obese Zucker rats is not associated with a lack of GLUT 4 translocation.

Galanin in the hypothalamus of fed and fasted lean and obese Zucker rats.

Galanin (GAL), a 29 aminoacid peptide, is widely distributed in the central nervous system and especially in the hypothalamus. It strongly stimulates food intake when it is injected in the paraventricular nucleus (PVN) of normal rats. The obese Zucker rat with a well-established hyperphagia is characterized by a general dysregulation of some important neuropeptides involved in the regulation of feeding behavior e.g. neurotensin, NPY or CCK and the aim of this study was to measure GAL in different microdissected brain areas in lean (Fa/Fa) and obese (fa/fa) male Zucker rats. As feeding status may modulate the central peptide concentrations, it was measured in ad libitum fed rats and in 48-h fasted rats of both genotypes. GAL was measured by a specific radioimmunoassay in the arcuate nuclei (ARC) and parvocellular (PVNp) and magnocellular (PVNm) parts of the PVN as well as in the median eminence (ME), median preoptic area (MPOA), supraoptic (SON) and dorsomedian (DMN) nuclei. Two-way analysis of variance revealed a very significant effect of genotype in the PVNp (P < 0.001), SON (P < 0.001) and in the ME (P < 0.02). No significant variations at all were noted in the ARC, PVNm, MPOA and DMN. GAL concentrations were more than doubled in the PVNp and SON of ad lib obese rats when compared to the ad lib lean rats (P < 0.005). On the other hand, in the ME where GAL concentration was about 4-fold greater than in the other areas, there was a 20 to 30% decrease in GAL concentrations in the obese rat (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolic effects of coconut, safflower, or menhaden oil feeding in lean and obese Zucker rats.

The aim of the present investigation was to study the effects of fish oil feeding in obese Zucker rats to establish its suitability as an animal model of hyperlipidaemia, and to understand the possible mechanism of fish oil-induced perturbations in cell metabolism. Lean and obese Zucker rats were fed on diets containing 180 g coconut, safflower, or menhaden oil/kg for 10 weeks. Body-weights and food intakes of lean coconut (LC), safflower (LS), and menhaden (LM) groups were similar. Obese menhaden (OM) rats had lower food intakes and body-weights compared with obese coconut (OC) and obese safflower (OS) groups, but values for all obese rats were higher than those for lean rats. Liver weights were higher in obese compared with lean rats, but on a percentage body-weight basis menhaden oil rats had higher values within genotype. Serum cholesterol and triacylglycerol levels were lower in the OM group compared with the OC and OS groups, and in the LM group compared with the LC group. Glucose and insulin levels were highest in OS rats followed by OC and OM rats and then the lean rats. Serum triiodothyronine and thyroxine were lower in OM rats compared with OC and OS rats. Liver mitochondrial state 3 rates with glutamate-malate and succinate were lower; mitochondrial beta-oxidation was unaffected and peroxisomal beta-oxidation was higher in menhaden oil rats compared with both coconut and safflower oil rats. In general, consumption of menhaden oil lowered hepatic malic enzyme (EC 1.1.1.38, 1.1.1.40), glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and glutathione peroxidase (EC 1.11.1.9) activities and elevated long-chain fatty acyl-CoA hydrolase (EC 3.1.2.2) activity when compared with the two other diets. It is concluded that obese Zucker rats do respond like human subjects to fish oil feeding but not to vegetable oils. The hypolipidaemic effect of fish oil appears to be mediated through a lowering of lipogenic enzymes, glucose-6-phosphate dehydrogenase and malic enzyme.

Cardiomyopathy associated with noninsulin-dependent diabetes.

Cardiovascular disease represents the major cause of morbidity and mortality in noninsulin-dependent diabetic patients. While it was once thought that atherosclerotic vascular disease was responsible for all of these adverse effects, recent studies support the notion that one of the major adverse complications of diabetes is the development of a diabetic cardiomyopathy characterized by defects in both diastolic and systolic function. Contributing to the development of the cardiomyopathy is a shift in myosin isozyme content in favor of the least active V3 form. Also defective in the noninsulin-dependent diabetic heart is regulation of calcium homeostasis. While transport of calcium by the sarcolemmal and sarcoplasmic reticular calcium pumps are minimally affected by noninsulin-dependent diabetes, significant impairment occurs in sarcolemmal Na(+)-Ca2+ exchanger activity. This defect limits the ability of of the diabetic heart to extrude calcium, contributing to an elevation in [Ca2+]i. Also promoting the accumulation of calcium by the diabetic cell is a decrease in Na+, K+ ATPase activity, which is known to increase [Ca2+]i secondary to a rise in [Na+]i. In addition, calcium influx via the calcium channel is stimulated. Although the molecular mechanisms underlying these defects are presently unknown, the possibility that they may be related to aberrations in glucose or lipid metabolism are considered. The evidence suggests that classical theories of glucose toxicity, such as excessive polyol production or glycosylation, appear to be insignificant factors in heart. Also insignificant are defects in lipid metabolism leading to accumulation of toxic lipid amphiphiles or triacylglycerol. Rather, the major defects involve membrane changes, such as phosphatidylethanolamine N-methylation and protein phosphorylation, which can be attributed to the state of insulin resistance.

Hypersecretion of islet amyloid polypeptide from pancreatic islets of ventromedial hypothalamic-lesioned rats and obese Zucker rats.

To investigate the possible role of islet amyloid polypeptide (IAPP) in the development of type 2 diabetes mellitus, we examined the IAPP content and secretion in pancreatic islets isolated from ventromedial hypothalamic (VMH)-lesioned rats and genetically obese Zucker rats, using a specific RIA for IAPP. Obesity and hyperinsulinemia were observed in rats 21 days after VMH lesioning. IAPP content was increased in the islets of VMH-lesioned rats compared with findings in the sham-operated controls (100.9 +/- 6.6 vs. 72.8 +/- 3.85 fmol/islet; P less than 0.01). Isolated islets of VMH-lesioned rats secreted larger amounts of IAPP in the presence of 2.8 mM and 16.7 mM glucose (2.99 +/- 0.98 and 11.2 +/- 1.29 fmol.islet(-1).3 h-1) than was noted in sham-operated rats (ND and 6.65 +/- 0.78 fmol.islet(-1).3 h-1). In the obese Zucker rats, aged 14 weeks, IAPP concentrations in the islets were elevated compared with lean rats (133.3 +/- 10.6 vs. 84.4 +/- 8.5 fmol/islet; P less than 0.01). The isolated islets secreted larger amounts of IAPP in response to 2.8 mM and 16.7 mM glucose (2.83 +/- 0.88 and 15.81 +/- 1.35 fmol.islet(-1).3 h-1) than did those from lean control rats (0.36 +/- 0.19 and 12.49 +/- 1.20 fmol.islet(-1).3 h-1). These results strongly suggest that overproduction and hypersecretion of IAPP occur in animals with obesity and hyperinsulinemia.

The relationship between thromboxane and cytosolic calcium modifiers in rat platelets.

Platelet activity is controlled, in part, by cytosolic free ionized calcium concentration ([Ca++]i). Regulation of platelet thromboxane (TXB2) synthesis may be by regulation of [Ca++]i. Dietary linoleate is a regulator of TXB2 synthesis, therefore, it may act by influencing [Ca++]i. Aspirin is a regulator of TXB2 synthesis by inhibition of cyclooxygenase; ouabain and nifedipine are regulators of [Ca++]i. This study was conducted to determine whether these affectors of TXB2 synthesis and [Ca++]i cause associated responses. Male nonobese Zucker rats were fed diets supplying 30% of energy (en%) as fat. Dietary fat was a mixture of corn oil and beef tallow to provide 3.0, 4.5, 6.0, or 7.5 en% linoleic acid, with cholesterol added to provide equal cholesterol in all diets. Rats were fed for 30 days with 6 rats/diet. Isolated rat platelets were assayed for FA composition; the percentage of linoleic acid in platelet FA rose linearly with increasing dietary linoleate (r = 0.76, P less than 0.0001). Resting and thrombin-stimulated platelet [Ca++]i and TXB2 synthesis were measured in the presence or absence of extracellular calcium and aspirin, ouabain, or nifedipine. Aspirin caused reductions in both parameters; nifedipine blocked [Ca++]i, but did not affect TXB2; ouabain increased both. Changes induced by those modifiers of TXB2 and platelet [Ca++]i caused changes that were in the same direction for both. CaCl2 caused an increase in both and the [Ca++]i was correlated with the square root of the TXB2; without CaCl2 the two were negatively correlated; aspirin, ouabain, and nifedipine treatments resulted in no significant correlations. The results suggest that there is a common modifier of [Ca++]i and TXB2 synthesis.

Dynamic of the GRF-induced GH response in genetically obese Zucker rats: influence of central and peripheral factors.

To determine the time onset of the growth hormone (GH) alteration in the genetically obese rat, we studied the in vivo and in vitro rat growth hormone releasing factor (rGRF(1-29)NH2)-induced GH secretion in 6- and 8-week-old lean and obese male Zucker rats. Under sodium pentobarbital anesthesia, rGRF(1-29)NH2 (GRF) was injected intravenously at two doses: 0.8 and 4.0 micrograms/kg b.w. Basal serum GH concentrations were similar in lean and obese age-matched animals. The GH response to both GRF doses tested was unchanged in 6-week-old obese rats as compared to their lean litter mates. In contrast, a significant decrease of the GH secretion in response to 4.0 micrograms/kg b.w. GRF was observed in the 8-week-old obese rats. The effect of GRF (1.56, 6.25 and 12.5 pM) was further studied in vitro, in a perifusion system of freshly dispersed anterior pituitary cells of lean and obese Zucker rats. Basal GH release was similar in the 6-week-old animal group. In contrast, it was significantly decreased in 8-week-old obese rats as compared to their lean litter mates. Stimulated GH response to 1.56 and 6.25 pM GRF was significantly greater in the 6-week-old obese group than in the age-matched control group. In contrast, the GH response to all GRF concentrations tested was significantly decreased in the 8-week-old obese rats as compared to their respective lean siblings. In 8-week-old obese rats, a decrease of GH pituitary content and an increase of hypothalamic somatostatin (SRIF) concentration were observed. Insulin and free fatty acid serum were significantly increased in 8-week-old obese rats. In contrast, lower insulin-like growth factor I serum levels were observed in the obese animals as compared to their lean litter mates. Finally, to further clarify the role of the periphery in the inhibition of GH secretion observed in the 8-week-old fatty rats, we exposed cultured pituitary cells of 8-week-old lean animals to 17% serum of their obese litter mates. A significant decrease of GRF-stimulated GH secretion of lean rat pituitary cells exposed to the obese serum was noted (P less than 0.05). This study demonstrates that, in the obese Zucker rat, an alteration of the GH response to GRF is evident by the 8th week of life. This defective GH secretion could be related to peripheral and central abnormalities.