How obesity causes diabetes in Zucker diabetic fatty rats.

The mechanisms of adipogenic diabetes in Zucker diabetic fatty (ZDF) rats, a model of obesity complicated by diabetes, are reviewed. In ZDF rats, a mutation in the leptin receptor, OB-R, is associated with leptin resistance, obesity, and increased fat content of islets. Exaggerated nitric oxide (NO) generation, attributed to high intracellular levels of long-chain fatty acids, impairs beta-cell function and prevents their compensation for obesity-induced diabetes. The resulting diabetic hyperglycemia can be completely prevented by agents that inhibit NO production.     

Cardiac carbohydrate metabolism in Zucker diabetic fatty rats

OBJECTIVE: The aim of this study was to test the hypothesis that, shortly after the development of Type-2 diabetes, alterations in cardiac carbohydrate metabolism precede the onset of abnormalities in systolic function. METHODS: Hearts from 11-week-old Zucker diabetic fatty (ZDF) rats and age matched controls were perfused in the isovolumic Langendorff mode with 13C-labeled glucose, lactate and pyruvate and unlabeled fatty acids. 13C-Nuclear magnetic resonance glutamate isotopomer analysis was carried out to determine the contributions of substrates to energy production. RESULTS: The ZDF group was hyperglycemic and the relative flux through pyruvate dehydrogenase (PDH) was significantly depressed compared to lean controls. In the lean group, lactate, pyruvate and glucose contributed 64+/-3, 24+/-3 and 11+/-1%, respectively, to total pyruvate oxidation. In the ZDF group, the contribution of glucose both to total pyruvate oxidation and to tissue lactate and alanine formation was significantly depressed. Cardiac function assessed by the rate-pressure product was similar in both groups. The fraction of active PDH was decreased in the ZDF group compared to controls (p<0.025). CONCLUSIONS: These results highlight significant changes in cardiac carbohydrate metabolism shortly after the development of hyperglycemia in a model of Type 2 diabetes in the absence of overt changes in systolic function.     

Beneficial effects of melatonin on obesity and lipid profile in young Zucker diabetic fatty rats

Abstract: The study objective was to investigate the effects of melatonin on obesity and obesity‐associated systolic hypertension and dyslipidemia in young male Zucker diabetic fatty (ZDF) rats, an experimental model of the metabolic syndrome. ZDF rats (n = 30) and lean littermates (ZL) (n = 30) were used. At 6 wk of age, both lean and fatty animals were subdivided into three groups (n = 10): naive (N), vehicle‐treated (V), and melatonin‐treated (M) (10 mg/kg/day) for 6 wk. Vehicle and melatonin were added to the drinking water. Melatonin reduced mean weight gain (51 ± 2/100 g BW) versus N‐ZDF group (58 ± 3, P < 0.05) without food intake differences. M‐ZDF rats showed an apparent reduction in systolic hypertension that proved not to be statistically significant, and a significant improvement in dyslipidemia, with a reduction in hypertriglyceridemia from 580 ± 40 to 420.6 ± 40.9 mg/dL (P < 0.01). Melatonin raised high‐density‐lipoprotein (HDL) cholesterol in ZDF (from 81.6 ± 4.9 to 103.1 ± 4.5 mg/dL, P < 0.01) and ZL rats (from 62.8 ± 4.8 to 73.5 ± 4.8 mg/dL, P < 0.05) and significantly reduced low‐density‐lipoprotein (LDL) cholesterol in ZDF rats from 5.20 ± 0.4 to 4.14 ± 0.3 mg/dL (P < 0.05) but had no effect on total cholesterol levels. To our knowledge, this is the first evidence of a positive effect of melatonin on overweight and lipid pattern of obese Zucker diabetic rats, supporting the proposition that melatonin administration may ameliorate overweight and lipid metabolism in humans. Because these benefits occurred in youth, before advanced metabolic and vascular complications, melatonin might help to prevent cardiovascular disease associated with obesity and dyslipidemia.     

Vasopeptidase inhibition prevents nephropathy in Zucker diabetic fatty rats

BACKGROUND: Blocking the renin-angiotensin system is an established therapeutic principle in diabetic nephropathy. We investigated whether inhibition of both neutral endopeptidase and ACE (vasopeptidase inhibition) can prevent functional and morphological features of nephropathy in the Zucker diabetic fatty (ZDF) rat, an animal model of type II diabetes. METHODS: Homozygous (fa/fa) ZDF rats (each n=15) aged 10 weeks were treated with placebo, ramipril (1 mg/kg/day in drinking water), or the vasopeptidase inhibitor AVE7688 (45 mg/kg/day in chow). Metabolic parameters and renal function (metabolic cages) were assessed at baseline (age 10 weeks), and at age 17, 27, and 37 weeks. Twenty heterozygous animals (fa/-) served as lean, nondiabetic controls. At age 37 weeks, the animals were sacrificed and the kidneys analyzed histopathologically. RESULTS: Overt diabetes mellitus (blood glucose >20 mmol/l) was established at age 17 weeks in all homozygous ZDF rats. In the placebo group, urinary protein excretion increased progressively from 8+/-1 (baseline) to 342+/-56 mg/kg/day (week 37) whereas diabetes and proteinuria were absent in the lean control group. Ramipril tended to reduce albuminuria and morphological damage (p=ns) but AVE7688 virtually prevented albuminuria (33+/-12 mg/kg/day, p<0.05 vs. ZDF placebo) and drastically reduced the incidence and severity of glomerulosclerosis and tubulointerstitial damage. CONCLUSIONS: In ZDF rats, development of diabetes mellitus is accompanied by functional and morphological kidney damage that resembles human diabetic nephropathy. Diabetic nephropathy can be prevented by chronic vasopeptidase inhibition.     

Melatonin improves glucose homeostasis in young Zucker diabetic fatty rats

The aim of this study was to investigate the effects of melatonin on glucose homeostasis in young male Zucker diabetic fatty (ZDF) rats, an experimental model of metabolic syndrome and type 2 diabetes mellitus (T2DM). ZDF rats (n=30) and lean littermates (ZL) (n=30) were used. At 6wk of age, both lean and fatty animals were subdivided into three groups, each composed of ten rats: naive (N), vehicle treated (V), and melatonin treated (M) (10mg/kg/day) for 6wk. Vehicle and melatonin were added to the drinking water. ZDF rats developed DM (fasting hyperglycemia, 460±39.8mg/dL; HbA(1) c 8.3±0.5%) with both insulin resistance (HOMA-IR 9.28±0.9 versus 1.2±0.1 in ZL) and decreased β-cell function (HOMA1-%B) by 75%, compared with ZL rats. Melatonin reduced fasting hyperglycemia by 18.6% (P<0.05) and HbA(1) c by 11% (P<0.05) in ZDF rats. Also, melatonin lowered insulinemia by 15.9% (P<0.05) and HOMA-IR by 31% (P<0.01) and increased HOMA1-%B by 14.4% (P<0.05). In addition, melatonin decreased hyperleptinemia by 34% (P<0.001) and raised hypoadiponectinemia by 40% (P<0.001) in ZDF rats. Moreover, melatonin reduced serum free fatty acid levels by 13.5% (P<0.05). These data demonstrate that oral melatonin administration ameliorates glucose homeostasis in young ZDF rats by improving both insulin action and β-cell function. These observations have implications on melatonin's possible use as a new pharmacologic therapy for improving glucose homeostasis and of obesity-related T2DM, in young subjects.     

Profiling of Zucker diabetic fatty rats in their progression to the overt diabetic state

Blood chemistry profiles (glucose, insulin, and triglycerides) and indirect calorimetry were performed on male Zucker diabetic fatty (ZDF) rats in a longitudinal fashion (starting at 7 weeks of age) to assess the nature and timing of specific events in the transition to overt diabetes. Peripheral (skeletal muscle) insulin resistance was clearly present at 7 weeks of age in ZDF rats, yet circulating glucose was only slightly above normal as a result of compensatory hyperinsulinemia. At a crucial stage from 7 to 8 weeks, a reduction in insulin levels instigated several deleterious changes resulting in reduced whole-body carbohydrate utilization and increased glycemia. In subsequent weeks, an inability to sustain peripheral glucose disposal as a consequence of a continuous decline in insulin levels further reduced carbohydrate utilization (increased lipid utilization) and enhanced the overt hyperglycemia. These observations document in a systematic fashion the alterations that define diabetic progression in ZDF rats.     

Metabolic imbalance of the insulin-like growth factor-I axis in Zucker diabetic fatty rats

In healthy conditions, insulin-like growth factor-I (IGF-I) acts in a coordinated fashion with insulin to lower glycemia, mainly by increasing insulin sensitivity in peripheral tissues. The aim of this study was to explore the relationship between glucose homeostasis and the endocrine IGF-I axis in Zucker diabetic fatty (ZDF) rats. The plasma levels of glucose, insulin, growth hormone, free IGF-I, total IGF-I (associated to insulin-like growth factor binding proteins plus free), and corticosterone were measured in 13-week-old ZDF rats and in age-matched controls under fasting and postprandial conditions. The plasma IGF-I binding capacity was measured by radioligand binding. In ZDF rats, fasting total and free IGF-I levels were reduced by 22% and 92%, respectively, compared with controls. Postprandial free IGF-I was reduced by 35%, whereas total IGF-I was unaffected. The plasma IGF-I binding capacity in ZDF rats was reduced by 24% after fasting and by 13% under postprandial conditions. A clear correlation between free IGF-I and insulin was observed in postprandial controls but not in ZDF rats. A principal component analysis clearly separated ZDF and control rats into 2 main components under both fasting and postprandial conditions. The first component was determined equally by total IGF-I, bound IGF-I, the free to total IGF-I ratio, and the IGF-I binding capacity. The second component was determined mostly by glucose and insulin. Our results show a marked alteration of the plasma IGF-I levels and of the capacity of plasma to bind IGF-I, and a disturbed relationship between IGF-I and postprandial insulinemia in a rat model of type 2 diabetes mellitus.     

Treatment of Zucker diabetic fatty rats with AVE7688 improves vascular and neural dysfunction

Aim:  Vasopeptidase inhibitors are drugs that inhibit angiotensin-converting enzyme and neutral endopeptidase (NEP). The latter is a protease that degrades vasoactive peptides and is increased in diabetes. We have previously shown that treating streptozotocin-induced diabetic rats, an animal model of type 1 diabetes, with AVE7688, a vasopeptidase inhibitor, improves neurovascular and neural function. In this study, we determined the effect of treating Zucker diabetic fatty (ZDF) rats, an animal model of type 2 diabetes, with AVE7688 on vascular and neural function.
Methods:  ZDF rats at 12 weeks of age were treated for 12 weeks with AVE7688 (500 mg/kg diet). Afterwards, vascular reactivity of epineurial arterioles of the sciatic nerve and nerve conduction velocity and blood flow was determined.
Results:  Vascular and neural function was significantly impaired in ZDF rats compared with age-matched lean (control) rats. Treating ZDF rats with AVE7688 improved vascular relaxation to acetylcholine and calcitonin gene-related peptide in epineurial arterioles. Motor and sensory nerve conduction velocity, endoneurial blood flow and thermal nociception end-points were also improved by treatment compared with untreated ZDF rats. Superoxide and expression of NEP were increased in epineurial arterioles from ZDF rats and attenuated by treatment with AVE7688.
Conclusions:  AVE7688 is an effective treatment for microvascular and neural disease in ZDF rats. Thus, vasopeptidase inhibitors may be an effective treatment for diabetic microvascular and neural complication in type 2 diabetes.     

Pharmacologic action of L-carnitine on hypertriglyceridemia in obese Zucker rats

Administration of pharmacologic amounts of L-carnitine was studied in the hypertriglyceridemic Zucker rat. When administered subcutaneously, doses from 250 to 2,000 mg/kg/d significantly decreased plasma triglycerides in obese rats over eight to 12 weeks, with no effect on plasma triglycerides in lean rats. Oral doses at the same high levels were not effective in decreasing plasma triglycerides. Triglyceride secretion rate was reduced from 367 micrograms/min to 168 micrograms/min in treated obese rats. Concurrently, liver lipid was increased twofold in obese treated rats, and the livers of these rats showed significant fatty infiltration. The mechanism of action of carnitine in decreasing plasma triglycerides appeared to be via decreased secretion of triglycerides by the liver of obese rats. There was no effect of L-carnitine in lean or obese rats on the following variables: carnitine palmitoyltransferase-A kinetics or malonyl CoA inhibition, mitochondrial or peroxisomal oxidative capacity, lipoprotein lipase in heart, muscle, and adipose, or fecal lipids. The effect of pharmacologic L-carnitine thus appears to be an inhibition of triglyceride synthesis and/or secretion by the liver.     

Endothelin antagonism improves hepatic insulin sensitivity associated with insulin signaling in Zucker fatty rats

In the present study, we investigated the effects of long-term treatment with the endothelin (ET) antagonist atrasentan, an ET(A)-selective antagonist, on whole body glucose metabolism and insulin signaling in a commonly used model of insulin resistance, the Zucker fatty rat. Zucker lean and fatty rats were maintained for 6 weeks on either control or atrasentan-treated water. Euglycemic-hyperinsulinemic clamps (4 mU/kg per minute) were performed at the end of the 6-week treatment on a subset of rats (n=10/treatment). In another subset (n=5/treatment), an insulin tolerance test was performed; liver and muscle tissues were harvested 10 minutes following the challenge for further analysis. Results of the clamps demonstrated that long-term atrasentan treatment significantly increased whole body glucose metabolism in fatty rats compared with vehicle control subjects. Insulin-induced insulin receptor substrate 1 tyrosine and protein kinase B serine phosphorylation were significantly reduced in the liver and muscle of fatty animals compared with their lean littermates. This reduction was overcome with atrasentan treatment in the liver but not in the muscle. There was no difference between lean and fatty animals, however, in insulin receptor substrate 1 and protein kinase B protein expression in the liver and muscle and no effect by atrasentan. In contrast, expression of the regulatory subunit of PI-3 kinase (p85alpha) was significantly increased in the liver but not in the muscle of fatty animals compared with their lean littermates and this was normalized to levels of lean animals with atrasentan treatment. These findings indicate that long-standing ET antagonism improves whole body glucose metabolism in Zucker fatty rats through improvements in insulin signaling in the liver. These results indicate that therapeutic ET antagonism may assist in correcting the insulin-resistant state.     

Arterial stiffness and the autonomic nervous system during the development of Zucker diabetic fatty rats

AimThis study aimed to investigate the role played by sympathovagal balance in arterial stiffness, a common feature of insulin resistance and type 2 diabetes.MethodsWe investigated the relationship between autonomic nervous system activity and arterial stiffness in Zucker diabetic fatty rats (ZDF: Gmi-fa/fa) and their age-matched controls (lean: ?/fa). Using simultaneous catheterization of the proximal and distal aorta, we measured intra-arterial blood pressure (BP), heart rate (HR), their variability (spectral analysis) and aortic pulse wave velocity (PWV) in a series of at least six conscious rats aged 6, 12, 18 and 24 weeks.ResultsBP and PWV increased with age (P < 0.001) in both strains with no differences between strains, despite the insulin resistance already present at 6 weeks in ZDF rats. HR was significantly lower (P < 0.001) in ZDF than in lean rats. In ZDF compared with lean rats, the low-frequency (LF) component of the systolic BP variations and the LF/high-frequency (HF) component of the pulse interval (PI) variation ratio were reduced (P < 0.01 and P < 0.05, respectively), while the HF component of the PI (HF-PI) variation was raised (P < 0.05). PWV was negatively correlated with HF-PI (r = −0.37, P < 0.01), but not with biochemical parameters. HF-PI was an independent variable explaining the variation in PWV.ConclusionDuring the development of disease of ZDF rats, sympathovagal balance might account for the lack of increase in PWV.     

Analysis of gene expression profiles in insulin-sensitive tissues from pre-diabetic and diabetic Zucker diabetic fatty rats

Insulin resistance occurs early in the disease process, preceding the development of type 2 diabetes. Therefore, the identification of molecules that contribute to insulin resistance and leading up to type 2 diabetes is important to elucidate the molecular pathogenesis of the disease. To this end, we characterized gene expression profiles from insulin-sensitive tissues, including adipose tissue, skeletal muscle, and liver tissue of Zucker diabetic fatty (ZDF) rats, a well characterized type 2 diabetes animal model. Gene expression profiles from ZDF rats at 6 weeks (pre-diabetes), 12 weeks (diabetes), and 20 weeks (late-stage diabetes) were compared with age- and sex-matched Zucker lean control (ZLC) rats using 5000 cDNA chips. Differentially regulated genes demonstrating > 1.3-fold change at age were identified and categorized through hierarchical clustering analysis. Our results showed that while expression of lipolytic genes was elevated in adipose tissue of diabetic ZDF rats at 12 weeks of age, expression of lipogenic genes was decreased in liver but increased in skeletal muscle of 12 week old diabetic ZDF rats. These results suggest that impairment of hepatic lipogenesis accompanied with the reduced lipogenesis of adipose tissue may contribute to development of diabetes in ZDF rats by increasing lipogenesis in skeletal muscle. Moreover, expression of antioxidant defense genes was decreased in the liver of 12-week old diabetic ZDF rats as well as in the adipose tissue of ZDF rats both at 6 and 12 weeks of age. Cytochrome P450 (CYP) genes were also significantly reduced in 12 week old diabetic liver of ZDF rats. Genes involved in glucose utilization were downregulated in skeletal muscle of diabetic ZDF rats, and the hepatic gluconeogenic gene was upregulated in diabetic ZDF rats. Genes commonly expressed in all three tissue types were also observed. These profilings might provide better fundamental understanding of insulin resistance and development of type 2 diabetes.     

Melatonin induces browning of inguinal white adipose tissue in Zucker diabetic fatty rats

Melatonin limits obesity in rodents without affecting food intake and activity, suggesting a thermogenic effect. Identification of brown fat (beige/brite) in white adipose tissue (WAT) prompted us to investigate whether melatonin is a brown‐fat inducer. We used Zücker diabetic fatty (ZDF) rats, a model of obesity‐related type 2 diabetes and a strain in which melatonin reduces obesity and improves their metabolic profiles. At 5 wk of age, ZDF rats and lean littermates (ZL) were subdivided into two groups, each composed of four rats: control and those treated with oral melatonin in the drinking water (10 mg/kg/day) for 6 wk. Melatonin induced browning of inguinal WAT in both ZDF and ZL rats. Hematoxylin–eosin staining showed patches of brown‐like adipocytes in inguinal WAT in ZDF rats and also increased the amounts in ZL animals. Inguinal skin temperature was similar in untreated lean and obese rats. Melatonin increased inguinal temperature by 1.36 ± 0.02°C in ZL and by 0.55 ± 0.04°C in ZDF rats and sensitized the thermogenic effect of acute cold exposure in both groups. Melatonin increased the amounts of thermogenic proteins, uncoupling protein 1 (UCP1) (by ~2‐fold, P < 0.01) and PGC‐1α (by 25%, P < 0.05) in extracts from beige inguinal areas in ZL rats. Melatonin also induced measurable amounts of UCP1 and stimulated by ~2‐fold the levels of PGC‐1α in ZDF animals. Locomotor activity and circulating irisin levels were not affected by melatonin. These results demonstrate that chronic oral melatonin drives WAT into a brown‐fat‐like function in ZDF rats. This may contribute to melatonin′s control of body weight and its metabolic benefits.     

Normal perivascular sensory dilator nerve function in arteries of Zucker diabetic fatty rats

BackgroundType II diabetes in humans is associated with pathology of both the cardiovascular and peripheral sensory nervous systems. Because abnormal vasodilator responses have been reported in animals of type II diabetes and perivascular sensory nerves are a source of vasodilator substances, we tested the hypothesis that sensory nerve-dependent relaxation is abnormal in arteries of the Zucker diabetic fatty (ZDF) rat model of type II diabetes.MethodsThe ZDF rats and genetic controls were studied at 26 weeks of age. Tail-cuff systolic blood pressure (BP) was measured, serum was obtained for chemical determinations, and mesenteric branch arteries were isolated for wire myograph analysis and confocal-based measurement of calcitonin gene-related peptide (CGRP) positive nerve density.ResultsNo differences in BP were detected. Serum glucose, triglycerides, and cholesterol were significantly elevated in ZDF. Sensory nerve-dependent vasodilation was assessed by measuring relaxation of phenylephrine preconstricted arterial segments to cumulative addition of divalent calcium ion (Ca²⁺) or capsaicin. Neither Ca²⁺- nor capsaicin-induced relaxation were different in ZDF versus control (maximal ZDF response to Ca²⁺ = 64% ± 2% v 59% ± 4%; ED₅₀ for Ca²⁺ = 3.7 ± 0.5 mmol/L v 3.2 ± 0.5 mmol/L; n = 5, P = not significant [NS]; maximal ZDF response to capsaicin = 68% ± 9% v 74% ± 4%; ZDF ED₅₀ = 3.8 ± 0.5 nmol/L v 9.8 ± 7 nmol/L; n = 5, P = NS). In contrast, the maximal relaxation response to acetylcholine was impaired in ZDF (maximal ZDF response = 83% ± 5% v 94% ± 2%, n = 4, P = .039; ED₅₀ for acetylcholine = 8.1 ± 2.9 nmol/L for ZDF v 33.5 ± 18.2; n = 4 per group, P = .086). The CGRP positive nerve density was not different between groups.ConclusionsBlood pressure, perivascular sensory nerve CGRP content, and dilator function is normal in the ZDF model of type II diabetes, whereas endothelium-dependent relaxation is impaired.     

Basal insulin hypersecretion in insulin-resistant Zucker diabetic and Zucker fatty rats: role of enhanced fuel metabolism.

The biochemical mechanisms responsible for basal hyperinsulinemia in insulin-resistant states have not been fully defined. We therefore studied pancreatic beta-cell function in vitro to characterize the relative importance of fuel metabolism or secretion via a constitutive pathway in the maintenance of high basal insulin secretion in Zucker diabetic fatty (ZDF) and Zucker fatty (ZF) rats. Insulin secretion from ZF (10+/-1.8 v 5+/-0.6 pmol/ng DNA/h) and ZDF (30+/-4 v 7+/-0.8 pmol/ng DNA/h) islets at 2.8 mmol/L glucose was two to four times greater than secretion from islets of lean littermate control rats. In response to a decreasing glucose concentration (from 12 to 0 mmol/L), a paradoxical increase in insulin secretion was observed in perfused ZDF rat pancreas. Insulin secretion at 2.8 mmol/L glucose was suppressed approximately 70% to 80% in islets from ZDF and ZF rats following exposure to diazoxide, a K+-adenosine triphosphate (K(ATP)) channel opener that inhibits membrane depolarization, or rotenone and oligomycin, agents that inhibit ATP production, or by incubation at 23 degrees C. Inhibition of glycolysis with mannoheptulose, 2-deoxyglucose, and iodoacetate or fatty acid oxidation with a carnitine palmitoyltransferase I inhibitor also significantly inhibited basal insulin secretion in islets of ZDF and ZF rats but not their lean littermates. Furthermore, the glycolytic flux at 2.8 mmol/L glucose was significantly higher in ZDF islets versus ZDF lean littermate (ZLC) islets (2.2+/-0.1 v 3.7+/-0.3 pmol/ng DNA/2 h, P < .01) and was suppressed by mannoheptulose. In ZDF and ZF islets, high basal insulin secretion was maintained despite a 50% reduction in the rate of proinsulin/insulin biosynthesis at 2.8 mmol/L glucose. The rate of proinsulin to insulin conversion and the ratio of proinsulin to insulin secretion by islets of ZDF rats were similar to the values in the lean littermates. Thus, basal hypersecretion in these two insulin-resistant models appears to be related to enhanced fuel metabolism rather than the contribution of a constitutive pathway of secretion.