Response of rat exocrine pancreas to high-fat and high-carbohydrate diets

Intake of diets with high fat content is a risk factor for acute pancreatitis and pancreatic cancer. The underlying mechanisms leading to the development of these diseases due to high fat intake are currently unknown. The current study was designed in rats to determine the physiologic and pathological consequences of a highfat diet that contained excess amounts of cottonseed oil or a high-carbohydrate diet that contained high amounts of sucrose on the exocrine pancreas. Rats were maintained on the diets for 4 weeks, and a cannula was inserted into the right jugular vein and one into the pancreatic duct for collection of pancreatic juice. Volume of the pancreatic juice and concentrations of amylase, lipase, and trypsinogen in the pancreatic juice were measured before and after infusions of CCK-8. Results showed that basal and CCK-stimulated pancreatic outputs of volume, amylase and lipase but not trypsinogen, were significantly elevated in intact rats given a high-fat diet when compared with rats given a high-carbohydrate diet. Forty-eight hours later, rats were sacrificed, and parts of the pancreas were removed for isolation of pancreatic acinar cells and for histopathologic studies. Pancreatic acini isolated from rats on a high-fat diet showed significantly lower basal and CCK-stimulated amylase release when compared with those on a high-carbohydrate diet. Histology of the pancreas of rats on a high-carbohydrate diet appeared normal; however, the pancreas of rats on high-fat diet showed significant alterations in exocrine pancreas. These results showed abnormalities in the exocrine pancreas of rats on a high-fat diet, that were not found in rats on a high-carbohydrate diet; further, they support the contention that a high-fat diet has a deleterious effect on the pancreas.     

Effects of long-term feeding of high-protein or high-fat diets on the response to exercise in the rat

The aim of this work was to find by which mechanisms an increased availability of plasma free fatty acids (FFA) reduced carbohydrate utilization during exercise. Rats were fed high-protein medium-chain triglycerides (MCT), high-protein long-chain triglycerides (LCT), carbohydrate (CHO) or high-protein low-fat (HP) diets for 5 weeks, and liver and muscle glycogen, gluconeogenesis and FFA oxidation were studied in rested and trained runner rats. In the rested state the hepatic glycogen store was decreased by fat and protein feeding, whereas soleus muscle glycogen concentration was only affected by high-protein diets. The percentage decrease in liver and muscle glycogen stores, after running, was similar in fat-fed, high-protein and CHO-fed rats. The fact that plasma glucose did not drastically change during exercise could be explained by a stimulation of hepatic gluconeogenesis: the activity of phosphoenolpyruvate carboxykinase (PEPCK) and liver phosphoenolpyruvate (PEP) concentration increased as well as cyclic adenosine monophosphate (AMPc) while liver fructose 2,6-bisphosphate decreased and plasma FFA rose. In contrast, the stimulation of gluconeogenesis in rested HP-, MCT- and LCT-fed rats appears to be independent of cyclic AMP.     

Increased response to insulin of glucose metabolism in the 6-day unloaded rat soleus muscle

Hind leg muscles of female rats (85-99 g) were unloaded by tail cast suspension for 6 days. In the fresh-frozen unloaded soleus, the significantly greater concentration of glycogen correlated with a lower activity ratio of glycogen phosphorylase (p less than 0.02). The activity ratio of glycogen synthase also was lower (p less than 0.001), possibly due to the higher concentration of glycogen. In isolated unloaded soleus, insulin (0.1 milliunit/ml) increased the oxidation of D-[U-14C]glucose, release of lactate and pyruvate, incorporation of D-[U-14C]glucose into glycogen, and the concentration of glucose 6-phosphate more (p less than 0.05) than in the weight-bearing soleus. At physiological doses of insulin, the percent of maximal uptake of 2-deoxy-D-[1,2-3H]glucose/muscle also was greater in the unloaded soleus. Unloading of the soleus increased by 50% the concentration of insulin receptors, due to no decrease in total receptor number during muscle atrophy. This increase may account for the greater response of glucose metabolism to insulin in this muscle. The extensor digitorum longus, which generally shows little response to unloading, displayed no differential response of glucose metabolism to insulin.     

Stimulation of microperoxisomal beta-oxidation in rat heart by high-fat diets

1. Heart microperoxisomal beta-oxidation activity, measured as cyanide-insensitive palmitoyl-CoA-dependent NAD+-reduction, was detected in a microperoxisome-enriched fraction from rat myocardium. The effect on this microperoxisomal beta-oxidation of the fatty acid composition of the dietary oils was investigated. 2. Feeding 15% (w/w) high erucic acid rapeseed oil or partially hydrogenated marine oil for 3 weeks increased the microperoxisomal beta-oxidation in the heart 4-5-fold, compared to a soybean oil diet. Increasing amounts (5-30%, w/w) of partially hydrogenated marine oil in the diet led to a 3-fold increase in the microperoxisomal beta-oxidation capacity at 20% or more of this oil in the diet. 3. The activity of the microperoxisomal marker enzyme catalase followed closely the cyanide-insensitive palmitoyl-CoA-dependent NAD+-reduction, except when feeding more than 20% (w/w) partially hydrogenated marine oil where a significant decrease in the catalase activity was observed. 4. In rapeseed oil-fed animals the extent of increase of microperoxisomal beta-oxidation was directly correlated to the amount of erucic acid (22:1, n-9 cis) in the diet. 5. Feeding partially hydrogenated rapeseed oil or partially hydrogenated soybean oil resulted in activities of microperoxisomal beta-oxidation significantly lower than in the corresponding unhydrogenated oils. No significant difference could be detected between diets containing hydrogenated or unhydrogenated marine oil. 6. Addition of 5% soybean oil to the essential fatty acid-deficient, partially hydrogenated marine oil diet did not change the effect on the microperoxisomal beta-oxidation activity. 7. Clofibrate feeding increased the heart microperoxisomal beta-oxidation capacity 2.5-fold, as compared to a standard pelleted diet. 8. These findings are discussed in relation to the transient nature of the cardiac lipidosis observed with animals fed on diets rich in C22:1 fatty acids. It is concluded that the heart plays an important part in the adaptation process.     

Overexpression of manganese superoxide dismutase ameliorates high-fat diet-induced insulin resistance in rat skeletal muscle

Elevated mitochondrial reactive oxygen species have been suggested to play a causative role in some forms of muscle insulin resistance. However, the extent of their involvement in the development of diet-induced insulin resistance remains unclear. To investigate, manganese superoxide dismutase (MnSOD), a key mitochondrial-specific enzyme with antioxidant modality, was overexpressed, and the effect on in vivo muscle insulin resistance induced by a high-fat (HF) diet in rats was evaluated. Male Wistar rats were maintained on chow or HF diet. After 3 wk, in vivo electroporation (IVE) of MnSOD expression and empty vectors was undertaken in right and left tibialis cranialis (TC) muscles, respectively. After one more week, insulin action was evaluated using hyperinsulinemic euglycemic clamp, and tissues were subsequently analyzed for antioxidant enzyme capacity and markers of oxidative stress. MnSOD mRNA was overexpressed 4.5-fold, and protein levels were increased by 70%, with protein detected primarily in the mitochondrial fraction of muscle fibers. This was associated with elevated MnSOD and glutathione peroxidase activity, indicating that the overexpressed MnSOD was functionally active. The HF diet significantly reduced whole body and TC muscle insulin action, whereas overexpression of MnSOD in HF diet animals ameliorated this reduction in TC muscle glucose uptake by 50% (P < 0.05). Decreased protein carbonylation was seen in MnSOD overexpressing TC muscle in HF-treated animals (20% vs. contralateral control leg, P < 0.05), suggesting that this effect was mediated through an altered redox state. Thus interventions causing elevation of mitochondrial antioxidant activity may offer protection against diet-induced insulin resistance in skeletal muscle.     

Exercise-induced adaptations of rat soleus muscle grafts

In female Wistar rats (n = 316) under pentobarbital sodium anesthesia, the soleus muscle was autografted with its nerve reimplanted. One purpose was to characterize the chronological development of graft innervation and recruitment during locomotion. Furthermore, we tested hypotheses regarding the efficacy of run conditioning of different intensities, durations, and postgrafting initiation times to alter mass and pyruvate-malate oxidation capacity of grafts. Choline acetyltransferase activity of grafts increased from 10% of control value at 7 days postgrafting to 55 and 100% at days 28 and 56, respectively. Running-induced glycogen depletion occurred in grafts; this is consistent with graft recruitment during locomotion. There was a threshold of conditioning intensity below which no improvements occurred and above which there were improvements. Spring (50 m/min) and endurance (30 m/min) conditioning of a duration of at least 28 days that was initiated at 28 or 56 days postgrafting increased mass of grafts by 30% compared with grafts from nonconditioned rats. Easy conditioning (15 m/min) had no effect on graft mass. Changes in graft total protein content paralleled those of mass. Oxidation capacity of grafts increased significantly with some conditioning protocols, but not to the same extent as mass. The exercise-induced adaptations should improve graft function in the host organism.     

Induction of peroxisomal beta-oxidation in rat liver by high-fat diets.

Liver peroxisomes were prepared by using a Percoll gradient in a vertical rotor. beta-Oxidation was measured in peroxisomes isolated from livers of rats fed on either high-(15% by wt.) or low- (5% by wt.) fat diets. The feeding of high-fat diets gave a 1.4-2.4-fold increase in total liver peroxisomal beta-oxidation, and a similar increase in specific activity. A 1.5-4.5-fold increase was seen in the specific activity of purified peroxisomal preparations. The reasons for these increases are discussed.     

Chronic leptin treatment enhances insulin-stimulated glucose disposal in skeletal muscle of high-fat fed rodents

The aim of this investigation was to evaluate if chronic leptin administration corrects high fat diet-induced skeletal muscle insulin resistance, in part, by enhancing rates of glucose disposal and if the improvements are accounted for by alterations in components of the insulin-signaling cascade. Sprague-Dawley rats consumed normal (CON) or high fat diets for three months. After the dietary lead in, the high fat diet group was further subdivided into high fat (HF) and high fat, leptin treated (HF-LEP) animals. HF-LEP animals were injected twice daily with leptin (5 mg/100 g body weight) for 10 days, while the CON and HF animals were injected with vehicle. Following the treatment periods, all animals were prepared for and subjected to hind limb perfusion. The high fat diet decreased rates of insulin-stimulated skeletal muscle glucose uptake and glycogen synthesis in the red gastrocnemius (RG), but did not affect glycogen synthase activity, rates of glucose oxidation or nonoxidative disposal of glucose. Of interest, IRS-1-associated PI3-K activity and total GLUT4 protein concentration were reduced in the RG of the high fat-fed animals. Leptin treatment increased rates of insulin-stimulated glucose uptake and glucose oxidation, and normalized rates of glycogen synthesis. Leptin appeared to mediate these effects by normalizing insulin-stimulated PI3-K activation and GLUT4 protein concentration in the RG. Collectively, these data suggest that chronic leptin treatment reverses the effects of a high fat diet thereby allowing the insulin signaling cascade and glucose transport effector system to be fully activated which in turn affects the amount of glucose that is transported across the plasma membrane and made available for glycogen synthesis.     

Effects of oxygen deprivation on incubated rat soleus muscle

Isolated soleus muscle deprived of oxygen produces more lactate and alanine than oxygen-supplied muscle. Oxygenated muscle synthesized glutamine, while anoxic muscle used this amino acid. Oxygen deprivation decreased adenine nucleotides leading to the efflux of nucleosides. Protein synthesis and degradation responded differently to anoxia. Synthesis almost completely ceased, while proteolysis increased. Therefore, protein degradation in soleus muscle is enhanced when energy supplies and oxygen tension are low.     

The effect of high-fat diets on microsomal lauric acid hydroxylation in rat liver

A diet with 20% (w/w) fish oil or partially hydrogenated fish oil has been shown to stimulate omega-oxidation of lauric acid 2.5-fold with rat liver microsomal preparations after 1 week of feeding. A diet containing either 20% (w/w) soybean oil, partially hydrogenated soybean oil or rapeseed oil had no effect. The omega-oxidation was also stimulated by fasting (3.7-fold) and by clofibrate (13-fold). The stimulation of omega-oxidation with partially hydrogenated fish oil was at its highest level after 3 days of feeding, and was dose dependent in the dietary oil of range 5-25% (w/w). With various high-fat diets, a high correlation was found (r = 0.81) between peroxisomal beta-oxidation of palmitoyl-CoA and microsomal omega-oxidation of lauric acid.     

Time-dependent regulation of muscle caveolin activation and insulin signalling in response to high-fat diet

We studied the effect of high-fat diet on the expression and activation of the three caveolins in rat skeletal muscle and their association with the insulin signalling cascade. Initial response was characterized by increased signalling through Cav-1 and Cav-3 phosphorylation, suggesting that both participate in an initial acute response to the calorie surplus. Afterwards, Cav-1 signalling was slightly reduced, whereas Cav-3 remained active. Late chronic phase signalling through both proteins was impaired inducing a prediabetic state. Summarizing, caveolins seem to mediate a time-dependent regulation of insulin cascade in response to high-fat diet in muscle.     

Twitch potentiation and caffeine contractures in isolated rat soleus muscle

1. Electrically-evoked twitch and tetanic tension were measured in isolated rat soleus muscle after exposure to caffeine. 2. Between 0.01 and 2.5 mM caffeine twitch tension was potentiated, reaching a peak of 150% of Resting Tension at 0.5 mM. 3. Biphasic Tension development with relaxation was observed at 2.5 mM caffeine with maximal contractures (110% tetanic tension) occurring at 20 mM. 4. Creatine phosphate and ATP stores were maintained throughout the period of tension development and relaxation. 5. In contrast with amphibian muscle, the isolated soleus is very sensitive to low doses of caffeine and produces biphasic caffeine contractures which relax in the presence of caffeine.     

Activity-induced recovery of excitability in K(+)-depressed rat soleus muscle

Increased extracellular K(+) concentration ([K(+)](o)) can reduce excitability and force in skeletal muscle. Here we examine the effects of muscle activation on compound muscle action potentials (M waves), resting membrane potential, and contractility in isolated rat soleus muscles. In muscles incubated for 60 min at 10 mM K(+), tetanic force and M wave area decreased to 23 and 24%, respectively, of the control value. Subsequently, short (1.5 s) tetanic stimulations given at 1-min intervals induced recovery of force and M wave area to 81 and 90% of control levels, respectively, within 15 min (P < 0.001). The recovery of force and M wave was associated with a partial repolarization of the muscle fibers. Experiments with tubocurarine suggest that the force recovery was related to activation of muscle Na(+)-K(+) pumps caused by the release of some compound from sensory nerves in response to muscle activity. In conclusion, activity produces marked recovery of excitability in K(+)-depressed muscle, and this may protect muscles against fatigue caused by increased [K(+)](o) during exercise.     

GSK3 involvement in amylin signaling in isolated rat soleus muscle

Amylin can evoke insulin resistance by antagonizing insulin in a non-competitive manner. Here, we investigated the glycogenolytic effect of amylin in isolated skeletal muscle and compared it to the effects of a calcitonin gene-related peptide (CGRP). Amylin alone had no statistically significant effect on glucose transport. However, amylin decreased insulin-stimulated glucose transport by about 30%. The involvement of cAMP could not be detected at the concentrations shown to promote glycogenolysis. Previously, it has been shown that increased glycogen synthase kinase 3 (GSK3) activity plays a role in insulin resistance. Here, the ratio of GSK3 :β isoforms in rat soleus was found to be 1.2:1. We found that amylin increased GSK3 activity, which in turn led to increased phosphorylation of glycogen synthase and decreased glycogen synthesis de novo.     

History-dependent mechanical properties of permeabilized rat soleus muscle fibers.

Permeabilized rat soleus muscle fibers were subjected to repeated triangular length changes (paired ramp stretches/releases, 0.03 l(0), +/- 0.1 l(0) s(-1) imposed under sarcomere length control) to investigate whether the rate of stiffness recovery after movement increased with the level of Ca(2+) activation. Actively contracting fibers exhibited a characteristic tension response to stretch: tension rose sharply during the initial phase of the movement before dropping slightly to a plateau, which was maintained during the remainder of the stretch. When the fibers were stretched twice, the initial phase of the response was reduced by an amount that depended on both the level of Ca(2+) activation and the elapsed time since the first movement. Detailed analysis revealed three new and important findings. 1) The rates of stiffness and tension recovery and 2) the relative height of the tension plateau each increased with the level of Ca(2+) activation. 3) The tension plateau developed more quickly during the second stretch at high free Ca(2+) concentrations than at low. These findings are consistent with a cross-bridge mechanism but suggest that the rate of the force-generating power-stroke increases with the intracellular Ca(2+) concentration and cross-bridge strain.