Evidence that L‐glutamine is better than L‐alanine as gluconeogenic substrate in perfused liver of weaned fasted rats submitted to short‐term insulin‐induced hypoglycaemia

Gluconeogenesis in livers from overnight fasted weaned rats submitted to short‐term insulin‐induced hypoglycaemia (IIH) was investigated. For this purpose, a condition of hyperinsulinemia/hypoglycaemia was obtained with an intraperitoneal (ip) injection of regular insulin (1.0 U kg^?1). Control group (COG group) received ip saline. The studies were performed 30 min after insulin (IIH group) or saline (COG group) injection. The livers from IIH and COG rats were perfused with L‐alanine (5 mM), L‐lactate (2 mM), L‐glutamine (10 mM) or glycerol (2 mM). Hepatic glucose, L‐lactate and pyruvate production from L‐alanine was not affected by IIH. In agreement with this result, the hepatic ability in producing glucose from L‐lactate or glycerol remained unchanged (IIH group vs. COG group). However, livers from IIH rats showed higher glucose production from L‐glutamine than livers from COG rats and, in the IIH rats, the production of glucose from L‐glutamine was higher than that from L‐alanine. The higher glucose production in livers from the IIH group, when compared with the COG group was due to its entrance further on gluconeogenic pathway. Taken together, the results suggest that L‐glutamine is better than L‐alanine, as gluconeogenic substrate in livers of hypoglyceaemic weaned rats. Copyright © 2008 John Wiley & Sons, Ltd.     

Gluconeogenesis and ketogenesis in perfused liver of rats submitted to short-term insulin-induced hypoglycaemia

Gluconeogenesis and ketogenesis of in situ rat perfused liver submitted to short-term insulin-induced hypoglycaemia (IIH) were investigated. For this purpose, 24-h fasted rats that received intraperitoneal (ip) regular insulin (1.0 U kg(-1)) or saline were compared. The studies were performed 30 min after insulin (IIH group) or saline (COG group) injection. For gluconeogenesis studies, livers from the IIH and COG groups were perfused with increasing concentrations (from basal blood concentrations until saturating concentration) of glycerol, L-lactate (Lac) or pyruvate (Pyr). Livers of the IIH group showed maintained efficiency to produce glucose from glycerol and higher efficiency to produce glucose from Lac and Pyr. In agreement with these results the oral administration of glycerol (100 mg kg(-1)), Lac (100 mg kg(-1)), Pyr (100 mg kg(-1)) or glycerol (100 mg kg(-1)) + Lac (100 mg kg(-1)) + Pyr (100 mg kg(-1)) promoted glycaemia recovery. It can be inferred that the increased portal availability of Lac, Pyr and glycerol could help glycaemia recovery by a mechanism mediated, partly at least, by a maintained (glycerol) or increased (Lac and Pyr) hepatic efficiency to produce glucose. Moreover, in spite of the fact that insulin inhibits ketogenesis, the capacity of the liver to produce ketone bodies from octanoate during IIH was maintained.     

Acute effects of isolated and combined L-alanine and L-glutamine on hepatic gluconeogenesis, ureagenesis and glycaemic recovery in experimental short-term insulin induced hypoglycaemia

The acute effects of isolated and combined L-alanine (L-Ala) and L-glutamine (L-Gln) on liver gluconeogenesis, ureagenesis and glycaemic recovery during short-term insulin-induced hypoglycaemia (IIH) were investigated. For this purpose, 24-h fasted rats that received intraperitoneal injection of regular insulin (1.0 U/Kg) were investigated. The control group (COG group) were represented by rats which received saline. The studies were performed 30 min after insulin (IIH group) or saline (COG group) injection. Livers from IIH and COG groups were perfused with basal or saturating levels of L-Ala, L-Gln or L-Gln + L-Ala (L-G + L-A). The production of glucose, urea, L-lactate and pyruvate in livers from IIH and COG group were markedly increased (p < 0.001) when perfused with saturating levels of L-Ala, L-Gln or L-G + L-A compared with basal levels of the same substrates. In addition, livers from IIH rats showed greater ability in producing glucose and urea from saturating levels of L-Ala compared with L-Gln or L-G + L-A. In agreement with these results, the oral administration of L-Ala (100 mg/kg) promoted better glycaemic recovery than L-Gln (100 mg/kg) or the combination of L-G (50 mg/kg) + L-A (50 mg/kg). It can be concluded that L-Ala, but not L-Gln or L-G + L-A could help glycaemic recovery by a mechanism mediated, partly at least, by the increased gluconeogenic and ureagenic efficiency of L-Ala.     

Time sequence of the intensification of the liver glucose production induced by high‐fat diet in mice

It is well established that the development of insulin resistance shows a temporal sequence in different organs and tissues. Moreover, considering that the main aspect of insulin resistance in liver is a process of glucose overproduction from gluconeogenesis, we investigated if this metabolic change also shows temporal sequence. For this purpose, a well‐established experimental model of insulin resistance induced by high‐fat diet (HFD) was used. The mice received HFD (HFD group) or standard diet (COG group) for 1, 7, 14 or 56 days. The HFD group showed increased (P < 0.05 versus COG) epididymal, retroperitoneal and inguinal fat weight from days 1 to 56. In agreement with these results, the HFD group also showed higher body weight (P < 0.05 versus COG) from days 7 to 56. Moreover, the changes induced by HFD on liver gluconeogenesis were progressive because the increment (P < 0.05 versus COG) in glucose production from l ‐lactate, glycerol, l ‐alanine and l ‐glutamine occurred 7, 14, 56 and 56 days after the introduction of the HFD schedule, respectively. Furthermore, glycaemia and cholesterolemia increased (P < 0.05 versus COG) 14 days after starting the HFD schedule. Taken together, the results suggest that the intensification of liver gluconeogenesis induced by an HFD is not a synchronous ‘all‐or‐nothing process’ but is specific for each gluconeogenic substrate and is integrated in a temporal manner with the progressive augmentation of fasting glycaemia. Copyright © 2012 John Wiley & Sons, Ltd.     

Effect of exposure to 50 Hz magnetic field with or without insulin on blood–brain barrier permeability in streptozotocin‐induced diabetic rats

We investigated the effect of long‐term exposure to modulation magnetic field (MF), insulin, and their combination on blood–brain barrier (BBB) permeability in a diabetic rat model. Fifty‐three rats were randomly assigned to one of six groups: sham, exposed to no MF; MF, exposed to MF; diabetes mellitus (DM), DM induced with streptozotocin (STZ); DM plus MF (DMMF); DM plus insulin therapy (DMI); and DM plus insulin therapy plus MF (DMIMF). All the rats underwent Evans blue (EB) measurement to evaluate the BBB 30 days after the beginning of experiments. The rats in MF, DMMF, and DMIMF groups were exposed to MF (B = 5 mT) for 165 min every day for 30 days. Mean arterial blood pressure (MABP), body mass, and serum glucose level of the study rats were recorded. The extravasation of brain EB of the MF, DM, DMMF, DMI, and DMIMF groups was higher than that of the sham group and the extravasation of right hemisphere of the DMIMF group was highest (P < 0.05). The post‐procedure body mass of the sham and MF groups were significantly higher than those of the DM and DMMF groups (P < 0.05). In the DM, DMMF, DMI, and DMIMF groups, the baseline glucose was significantly lower than the post‐procedure glucose (P < 0.05). DM and MF increase BBB permeability; in combination, they cause more increase in BBB permeability, and insulin decreases their effect on BBB. Improved glucose metabolism may prevent body mass loss and the hypoglycemic effect of MF. DM increases MABP but MF causes no additional effect. Bioelectromagnetics 31:262–269, 2010. © 2009 Wiley‐Liss, Inc.     

Daily Oral Oleoyl‐Estrone Gavage Induces a Dose‐Dependent Loss of Fat in Wistar Rats

Objective: To establish whether single daily oral doses of oleoyl‐estrone result in dose‐dependent slimming effects on normal weight rats, and to determine the changes in energy parameters induced by this treatment. Research Methods and Procedures: The effects of a daily oral gavage of oleoyl‐estrone (0, 0.2, 0.5, 1, 2, 5, 10, and 20 μmol/kg per day) in 0.2 ml of sunflower oil given over a 10‐day period were studied in groups, each of which contained six adult female Wistar rats initially weighing 190 to 230 g. A group of intact control rats receiving no gavage was included for comparison. Body weight and food intake were measured daily. Rats were killed on day 10 of treatment, and body composition (protein nitrogen, lipids, and water), liver lipids, and plasma parameters (glucose, triacylglycerols, total cholesterol, free fatty acids, 3‐hydroxybutyrate, urea, aspartate, alanine transaminases, insulin, leptin, and free and acyl‐estrone) were measured. Results: The administration of oleoyl‐estrone resulted in a dose‐dependent loss of body fat, because of a partly maintained energy expenditure combined with decreased food intake. The differences in the energy budget were met by internal fat pools. The changes recorded did not affect the levels of the main plasma energy homeostasis indicators: unaltered glucose, triacylglycerols, free fatty acids, 3hydroxybutyrate, and urea. Protein was accrued even under conditions of severe lipid store drainage. There were no changes in transaminases. No lipid accumulation was recorded in the liver. Plasma insulin and leptin levels decreased with increased oleoyl‐estrone doses, whereas the levels of free and esterified estrone increased with treatment, although not in proportion to the dose received. Discussion: Oral treatment with oleoyl‐estrone resulted in the specific dose‐related loss of fat reserves with little change to other metabolic parameters. These results agree with the postulated role of oleoyl‐estrone as a ponderostat signal.     

Impaired sensitivity to insulin of rat livers perfused with blood of diminished haematocrit.

1. In livers from fed rats perfused with homologous whole blood of a haematocrit value of 37%, insulin decreased the perfusate concentrations of glucose and amino acids, production of ketone bodies (3-hydroxybutyrate + acetoacetate) and increased bile flow. 2. Perfusion with blood diluted with buffer to a haematocrit value of 17% decreased hepatic O2 consumption by 40-50%. Perfusate concentrations of glucose and lactate, the rate of ketogenesis and the ratios [lactate]/[pyruvate] and [3-hydroxybutyrate]/[acetoacetate] were all increased. 3. In livers perfused with blood of diminished haematocrit, effects of insulin on perfusate glucose an amino acids, ketogenesis and bile flow were abolished.     

Comparative acute effects of leptin and insulin on gluconeogenesis and ketogenesis in perfused rat liver

The acute effects of physiological levels of leptin (10 ng ml(-1)) and insulin (20 microU ml(-1)) on hepatic gluconeogenesis and ketogenesis were compared. Leptin or insulin alone decreased (p<0.05) the activation of hepatic glucose, L-lactate and urea production from L-alanine. However, the hepatic glucose production was not modified if leptin was combined with insulin. These results indicated that both, i.e. leptin and insulin, could promote a non-additive reduction in the rate of catabolism of L-alanine. However, in contrast with insulin (p<0.05), leptin did not inhibit the activation of hepatic glucose production from pyruvate or glycerol. On the other hand, activation of hepatic production of acetoacetate and beta-hydroxybutyrate from octanoate was not affected by leptin or insulin. Thus, our data demonstrate that the acute effect of leptin on hepatic metabolism was partially similar to insulin (activation of glucose production from L-alanine and activation of acetoacetate or beta-hydroxybutyrate production from octanoate) and partially different from insulin (activation of glucose production from pyruvate or glycerol).     

Blood amino acids concentration during insulin induced hypoglycemia in rats: the role of alanine and glutamine in glucose recovery

Summary. Our purpose was to determine the blood amino acid concentration during insulin induced hypoglycemia (IIH) and examine if the administration of alanine or glutamine could help glycemia recovery in fasted rats. IIH was obtained by an intraperitoneal injection of regular insulin (1.0 U/kg). The blood levels of the majority of amino acids, including alanine and glutamine were decreased (P < 0.05) during IIH and this change correlates well with the duration than the intensity of hypoglycemia. On the other hand, the oral and intraperitoneal administration of alanine (100 mg/kg) or glutamine (100 mg/kg) accelerates glucose recovery. This effect was partly at least consequence of the increased capacity of the livers from IIH group to produce glucose from alanine and glutamine. It was concluded that the blood amino acids availability during IIH, particularly alanine and glutamine, play a pivotal role in recovery from hypoglycemia.     

Antioxidant icariside II combined with insulin restores erectile function in streptozotocin‐induced type 1 diabetic rats

Erectile dysfunction (ED) worsens in patients with diabetes mellitus (DM) despite good control of blood glucose level with insulin. Recent studies imply that diabetic vascular stresses (e.g. oxidative stress) persist in spite of glucose normalization, which is defined as metabolic memory. Studies suggest that the interaction between advanced glycation end products (AGEs) and their receptor (RAGE) mediates the development of metabolic memory. To investigate the effects of the antioxidant icariside II plus insulin on erectile function in streptozotocin (STZ)‐ induced type 1 diabetic rats. Fifty 8‐week‐old Sprague‐Dawley rats were randomly distributed into five groups: normal control, diabetic, insulin‐treated diabetic, icariside II‐treated diabetic, and insulin plus icariside II‐treated diabetic. Diabetes was induced by a single intraperitoneal injection of STZ. Eight weeks after induction of diabetes, icariside II was administered by gastric lavage once a day (5 mg/kg) for 6 weeks; and 2–6 units of intermediate‐acting insulin were given to maintain normal glycemia for 6 weeks. The main outcome measures were the ratio of intracavernous pressure (ICP) to mean arterial pressure (MAP); histology of penile endothelial cells and smooth muscle cells; neural nitric oxide synthase, AGEs and RAGE expression; malondialdehyde concentration; superoxide dismutase activity; and apoptosis index. Diabetic rats demonstrated a significantly lower ICP/MAP ratio, reduced penile endothelial cells, reduced smooth muscle cells, increased AGEs and RAGE, and increased apoptosis. Insulin and icariside II monotherapy partially restored erectile function and histological changes. However, the combination therapy group showed significantly better erectile parameters, cytological components and biochemistry, similar to those in the normal control group. These results suggest that, although insulin can effectively control glycemic levels, it does not completely alter the pathological changes in erectile tissues. Better efficacy could be expected with tight glycemic control plus the antioxidant icariside II. The proposed combination therapy might have the potential to eliminate metabolic memory by down‐regulating the AGEs‐RAGE‐oxidative stress axis.     

Contribution of hepatic glycogenolysis and gluconeogenesis in the defense against short-term insulin induced hypoglycemia in rats

In this study, the contribution of liver glycogenolysis and gluconeogenesis in the defense against short-term insulin induced hypoglycemia (IIH) was investigated. For this purpose, we used an experimental model in which IIH was obtained by administering an IP injection of a pharmacological dose (1 U/kg) of regular insulin to rats that had been deprived of food for a period of six hours. This experimental model is suitable to study the simultaneous participation of glycogen breakdown and gluconeogenesis in the defense against IIH. The livers of IIH rats showed insignificant changes in the glycogen concentration, total phosphorylase, active phosphorylase, and percent of active phosphorylase. Our results also indicated that the livers of IIH rats that received the concentration of L-alanine, L-glutamine, L-lactate, or glycerol found in the blood during IIH (basal values) showed negligible glucose production. Nonetheless, glucose, urea, and pyruvate production increased (P<0.05) if the livers were perfused with a saturating concentration of gluconeogenic precursors. In agreement with these results, IIH rats that received intragastric L-alanine, L-glutamine, or L-lactate showed increased (P<0.05) glycemia 30 min after the administration of these substances. However, when using glycerol, higher glycemia (P<0.05) was observed at 2 and 5 min, but not 30 min after the administration of this hepatic gluconeogenic precursor. Thus, we can conclude that the oral availability of gluconeogenic precursors could allow for their use as important antidote in the defense against IIH.     

Liver glycogen metabolism during short-term insulin-induced hypoglycemia in fed rats

The activities of glycogen phosphorylase and synthase during infusions of glucagon, isoproterenol, or cyanide in isolated liver of fed rats submitted to short-term insulin-induced hypoglycemia (IIH) was investigated. A condition of hyperinsulinemia/hypoglycemia was obtained with an intraperitoneal injection of regular insulin (1.0 U kg(-1)). The control group received ip saline. The experiments were carried out 60 min after insulin (IIH group) or saline (COG group) injection. The rats were anesthetized and after laparotomy, blood was collected from the vena cava for glucose and insulin measurements. The liver was then infused with glucagon (1 nM), isoproterenol (2 microM), or cyanide (0.5 mM) during 20 min and a sample of the organ was collected for determination of the activities of glycogen phosphorylase and synthase 5 min after starting and 10 min after stopping the infusions. The infusions of cyanide, glucagons, and isoproterenol did not change the activities of glycogen synthase and glycogen phosphorylase. However, glycogen catabolism was decreased during the infusions of glucagon and isoproterenol in IIH rats, being more intense with isoproterenol (p < 0.05), than glucagon. It was concluded that short-term IIH promoted changes in the liver responsiveness of glycogen degradation induced by glucagon and isoproterenol without a change in the activities of glycogen phosphorylase and synthase.     

Effects of Insulin on Ketogenesis Following Fasting in Lean and Obese Men

The ketone bodies (KBs) D‐3‐hydroxybutyrate (D‐3HB) and acetoacetate (AcAc) play a role in starvation and have been associated with insulin resistance. The dose–response relationship between insulin and KBs was demonstrated to be shifted to the right in type 2 diabetes patients. However, KB levels have also been reported to be decreased in obesity. We investigated the metabolic adaptation to fasting with respect to glucose and KB metabolism in lean and obese men without type 2 diabetes using stable glucose and D‐3HB isotopes in a two‐step pancreatic clamp after 38 h of fasting. We found that D‐3HB fluxes in the basal state were higher in lean compared to obese men: 15.2 (10.7–27.1) vs. 7.0 (3.5–15.1) µmol/kg lean body mass (LBM)·min, respectively, P < 0.01. No differences were found in KB fluxes between lean and obese volunteers during the pancreatic clamp (step 1: 6.9 (1.8–12.0) vs. 7.4 (4.2–17.8) µmol/kg LBM·min, respectively; and step 2: 2.9 (0–7.2) vs. 3.4 (0.85–18.7) µmol/kg LBM·min, respectively), despite similar plasma insulin levels. Meanwhile, peripheral glucose uptake was higher in lean compared to obese men (step 1: 15.2 (12.3–25.6) vs. 14.7 (11.9–22.7) µmol/kg LBM·min, respectively, P ≤ 0.05; and step 2: 12.5 (7.0–17.3) vs. 10.8 (5.2–15.0) µmol/kg LBM·min, respectively, P ≤ 0.01). These data show that obese subjects who display insulin resistance on insulin‐mediated peripheral glucose uptake have the same sensitivity for the insulin‐mediated suppression of ketogenesis. This implies differential insulin sensitivity of intermediary metabolism in obesity.     

On the lack of formation of l-(+)-3-hydroxybutyrate by liver

The terminal carbon of palmitic acid, traced with ¹⁴C, is preferentially incorporated into carbon 4 of hydroxybutyrate formed by hepatocytes and perfused livers from 18- to 19-day-old rats and perfused livers from fasted adult rats. However, ¹⁴C from [13-¹⁴C]palmitic acid is incorporated into carbon 1 of the hydroxybutyrate to the same extent as any one of the first 12 carbons of palmitic acid as assessed with [1-¹⁴C]palmitic acid and [6-¹⁴C]palmitic acid. Therefore, the hydroxybutyrate is formed via hydroxymethylglutaryl-CoA, i.e., it is in the d configuration, and hydrolysis of l-hydroxybutyryl-CoA, the intermediate in the β oxidation of the palmitate, does not occur. Further, a negligible amount of ¹⁴C remains in hydroxybutyrate formed from ¹⁴C-labeled palmitic acid by isolated hepatocytes and perfused livers from the young rats, when the hydroxybutyrate is treated with d-(?)-3-hydroxybutyrate dehydrogenase to convert the d isomer to acetoacetate. Thus, l-(+)-3-hydroxybutyrate is not produced by rat liver as assessed using these preparations.     

A possible mechanism for the anti-ketogenic action of alanine in the rat

1. The anti-ketogenic effect of alanine has been studied in normal starved and diabetic rats by infusing l-alanine for 90min in the presence of somatostatin (10μg/kg body wt. per h) to suppress endogenous insulin and glucagon secretion. 2. Infusion of alanine at 3mmol/kg body wt. per h caused a 70±11% decrease in [3-hydroxybutyrate] and a 58±9% decrease in [acetoacetate] in 48h-starved rats. [Glucose] and [lactate] increased, but [non-esterified fatty acid], [glycerol] and [3-hydroxybutyrate]/[acetoacetate] were unchanged. 3. Infusion of alanine at 1mmol/kg body wt. per h caused similar decreases in [ketone body] (3-hydroxybutyrate plus acetoacetate) in 24h-starved normal and diabetic rats, but no change in other blood metabolites. 4. Alanine [3mmol/kg body wt. per h] caused a 72±9% decrease in the rate of production of ketone bodies and a 57±8% decrease in disappearance rate as assessed by [3-¹⁴C]acetoacetate infusion. Metabolic clearance was unchanged, indicating that the primary effect of alanine was inhibition of hepatic ketogenesis. 5. Aspartate infusion at 6mmol/kg body wt. per h had similar effects on blood ketone-body concentrations in 48h-starved rats. 6. Alanine (3mmol/kg body wt. per h) caused marked increases in hepatic glutamate, aspartate, malate, lactate and citrate, phosphoenolpyruvate, 2-phosphoglycerate and glucose concentrations and highly significant decreases in [3-hydroxybutyrate] and [acetoacetate]. Calculated [oxaloacetate] was increased 75%. 7. Similar changes in hepatic [malate], [aspartate] and [ketone bodies] were found after infusion of 6mmol of aspartate/kg body wt. per h. 8. It is suggested that the anti-ketogenic effect of alanine is secondary to an increase in hepatic oxaloacetate and hence citrate formation with decreased availability of acetyl-CoA for ketogenesis. The reciprocal negative-feedback cycle of alanine and ketone bodies forms an important non-hormonal regulatory system.     

Seizure‐induced impairment in neuronal ketogenesis: Role of zinc‐α2‐glycoprotein in mitochondria

Ketone bodies (KBs) were known to suppress seizure. Untraditionally, neurons were recently reported to utilize fatty acids and produce KBs, but the effect of seizure on neuronal ketogenesis has not been researched. Zinc‐α2‐glycoprotein (ZAG) was reported to suppress seizure via unclear mechanism. Interestingly, ZAG was involved in fatty acid β‐oxidation and thus may exert anti‐epileptic effect by promoting ketogenesis. However, this promotive effect of ZAG on neuronal ketogenesis has not been clarified. In this study, we performed immunoprecipitation and mass spectrometry to identify potential interaction partners with ZAG. The mechanisms of how ZAG translocated into mitochondria were determined by quantitative coimmunoprecipitation after treatment with apoptozole, a heat shock cognate protein 70 (HSC70) inhibitor. ZAG level was modulated by lentivirus in neurons or adeno‐associated virus in rat brains. Seizure models were induced by magnesium (Mg²⁺)‐free artificial cerebrospinal fluid in neurons or intraperitoneal injection of pentylenetetrazole kindling in rats. Ketogenesis was determined by cyclic thio‐NADH method in supernatant of neurons or brain homogenate. The effect of peroxisome proliferator–activated receptor γ (PPARγ) on ZAG expression was examined by Western blot, quantitative real‐time polymerase chain reaction (qRT‐PCR) and chromatin immunoprecipitation qRT‐PCR. We found that seizure induced ketogenesis deficiency via a ZAG‐dependent mechanism. ZAG entered mitochondria through a HSC70‐dependent mechanism, promoted ketogenesis by binding to four β‐subunits of long‐chain L‐3‐hydroxyacyl‐CoA dehydrogenase (HADHB) and alleviated ketogenesis impairment in a neuronal seizure model and pentylenetetrazole‐kindled epileptic rats. Additionally, PPARγ activation up‐regulated ZAG expression by binding to promoter region of AZGP1 gene and promoted ketogenesis through a ZAG‐dependent mechanism.     

Metabolic Implications of Dietary Trans‐fatty Acids

Dietary trans‐fatty acids are associated with increased risk of cardiovascular disease and have been implicated in the incidence of obesity and type 2 diabetes mellitus (T2DM). It is established that high‐fat saturated diets, relative to low‐fat diets, induce adiposity and whole‐body insulin resistance. Here, we test the hypothesis that markers of an obese, prediabetic state (fatty liver, visceral fat accumulation, insulin resistance) are also worsened with provision of a low‐fat diet containing elaidic acid (18:1t), the predominant trans‐fatty acid isomer found in the human food supply. Male 8‐week‐old Sprague–Dawley rats were fed a 10% trans‐fatty acid enriched (LF‐trans) diet for 8 weeks. At baseline, 3 and 6 weeks, in vivo magnetic resonance spectroscopy (¹H‐MR) assessed intramyocellular lipid (IMCL) and intrahepatic lipid (IHL) content. Euglycemic–hyperinsulinemic clamps (week 8) determined whole‐body and tissue‐specific insulin sensitivity followed by high‐resolution ex vivo ¹H‐NMR to assess tissue biochemistry. Rats fed the LF‐trans diet were in positive energy balance, largely explained by increased energy intake, and showed significantly increased visceral fat and liver lipid accumulation relative to the low‐fat control diet. Net glycogen synthesis was also increased in the LF‐trans group. A reduction in glucose disposal, independent of IMCL accumulation was observed in rats fed the LF‐trans diet, whereas in rats fed a 45% saturated fat (HF‐sat) diet, impaired glucose disposal corresponded to increased IMCL_TA. Neither diet induced an increase in IMCL_soleus. These findings imply that trans‐fatty acids may alter nutrient handling in liver, adipose tissue, and skeletal muscle and that the mechanism by which trans‐fatty acids induce insulin resistance differs from diets enriched with saturated fats.     

Beneficial effects of (−)‐epigallocatechin‐3‐O‐gallate on diabetic peripheral neuropathy in the rat model

Diabetic neuropathic pain is characterized by spontaneous pain with hyperalgesia and allodynia. We investigated whether (?)‐epigallocatechin‐3‐O‐gallate could improve diabetic neuropathic pain development through hypoglycemic, hypolipidemic, antioxidant, and anti‐inflammatory effects. Diabetes was induced in rats by streptozotocin (55 mg/kg/once) and treated with (?)‐epigallocatechin‐3‐O‐gallate (25 mg/kg/orally/once/daily/5 weeks). Diabetic rats showed an increase in serum levels of glucose, nitric oxide, triglyceride, total cholesterol, and low‐density lipoprotein‐cholesterol with a decrease in high‐density lipoprotein‐cholesterol and body weight. Also, there was an elevation in brain malondialdehyde with a marked reduction in brain levels of glutathione, superoxide dismutase, catalase, glutathione peroxidase, and glutathione‐S‐transferase. Furthermore, diabetic rats showed a clear reduction in plasma levels of insulin and an increase in plasma cytokines (interleukin‐6 and tumor necrosis factor‐α). Moreover, diabetic rats exhibited hyperalgesia as indicated by a hot plate, tail immersion, formalin, and carrageenan‐induced edema tests as well as brain histopathological changes (neuron degeneration, gliosis, astrocytosis, congestion and hemorrhage). (?)‐Epigallocatechin‐3‐O‐gallate treatment ameliorated alterations in body weight, biochemical parameters, pain sensation, and histopathological changes in brain tissue. (?)‐Epigallocatechin‐3‐O‐gallate offers promising hypoglycemic, hypolipidemic, antioxidant and anti‐inflammatory effects, which can prevent the development and progression of diabetic neuropathic pain.     

Enhanced Dopamine D1 and D2 Receptor Gene Expression in the Hippocampus of Hypoglycaemic and Diabetic Rats

Hypoglycaemic coma and brain injury are potential complications of insulin therapy. Hippocampal neurons are particularly vulnerable to hypoglycaemic stress leading to memory impairment. In the present article, we have investigated the dopamine (DA) content, homovanillic acid (HVA)/DA turnover ratio, DA D₁ and DA D₂ receptors in the hippocampus of insulin-induced hypoglycaemic (IIH) and streptozotocin induced diabetic rats where brain functions are impaired. The DA content decreased significantly in hippocampus of diabetic, diabetic +IIH and control +IIH rats compared to control. The HVA/DA turnover ratio also increased significantly in diabetic, diabetic +IIH and control +IIH rats compared to control. Scatchard analysis using [³H] DA in the hippocampus showed a significant increase in DA receptors of diabetic, diabetic +IIH and control +IIH rats with decreased affinity. Gene expression studies using Real-time PCR showed an increased expression of DA D₁ and DA D₂ receptors in the hippocampus of hypoglycaemic and diabetic rats. Our results indicate that the dopaminergic system is impaired in the hippocampus of hypoglycaemic and hyperglycaemic rats impairing DA related functions of hippocampus. We observed a prominent dopaminergic functional disturbance in the hypoglycaemic condition than in hyperglycaemia compared to control. This dopaminergic dysfunction in hippocampus during hypoglycaemia and hyperglycaemia is suggested to contribute to cognitive and memory deficits. This will have clinical significance in the treatment of diabetes.     

The effects of ketone bodies,bicarbonate, and calcium on hepatic mitochondrial ketogenesis

The effect of various factors on hepatic mitochondrial ketogenesis was investigated in the rat. A comparison of three different incubation media revealed that bicarbonate ion inhibited the rate of ketone body production and decreased the ratio of 3-hydroxybutyrate/acetoacetate. The addition of 0.8 mm calcium caused significant inhibition of ketogenesis from both octanoate (40–50%) and palmitate (25–30%) and no change in the ratio of 3-hydroxybutyrate/acetoacetate. In the presence of components of the malate/aspartate shuttle, the inhibition by calcium was 80% or more with both substrates. Experimental alteration of the respiratory state of the mitochondria from state 3 to state 4 was associated with an enhanced rate of ketogenesis. The addition of ketone bodies themselves had marked effects on the rate of ketone body production. Increasing amounts of exogenously added acetoacetate were accompanied by increasing rates of total ketone body production reflecting enhanced 3-hydroxybutyrate synthesis. In the presence of added 3-hydroxybutyrate, there was striking inhibition of ketogenesis. Rotenone, which prevents oxidation of NADH₂ via the electron transport chain, almost completely inhibited ketone body synthesis. This inhibition was partially overcome by the addition of acetoacetate which regenerates NAD⁺ from NADH₂ during conversion to 3-hydroxybutyrate. These observations provide evidence for additional sites of metabolic control over hepatic ketogenesis.