Brain-derived neurotrophic factor ameliorates hepatic insulin resistance in Zucker fatty rats

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophins, has been reported to ameliorate hyperglycemia in obese diabetic animal models. To elucidate the mechanism of BDNF on glucose metabolism, we determined the glucose turnover under basal and euglycemic hyperinsulinemic (insulin infusion rate, 54 pmol. kg(-1). min(-1)) clamp conditions in obese insulin-resistant rats, male Zucker fatty rats, which had been acutely administered a subcutaneous injection of BDNF (20 mg/kg) (n = 9, BDNF) or vehicle (n = 8, vehicle). Under the basal condition, acute administration of BDNF did not affect the blood glucose level, plasma insulin level, rate of glucose disappearance (Rd), and endogenous glucose production (EGP). Under the clamp condition, the glucose infusion rate (GIR) was significantly higher in BDNF than in vehicle (mean +/- SD, 61.4 +/- 19.1 v 41.4 +/- 4.9 micromol. kg(-1). min(-1), P <.05). There was no significant difference in Rd and EGP between the 2 groups under the clamp condition, but the insulin-mediated suppression ratio of endogenous glucose production in BDNF was significantly greater than in vehicle (48.9 +/- 22.2 v 22.4% +/- 20.6%, P <.05). In BDNF, mRNA expressions of hepatic phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) were comparable to those of vehicle, while hepatic glucokinase (GK) mRNA expression was significantly higher (1.57 +/- 0.33 v 1.03 +/- 0.17, P <.05). We conclude that BDNF mainly improves hepatic insulin resistance in obese insulin-resistant rats, probably by affecting the hepatic GK flux. Copyright 2003, Elsevier Science (USA). All rights reserved.     

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.     

Dynamics of insulin hypersecretion by obese Zucker rats

The relationship between obesity and hypersecretion of insulin by the pancreas was studied. We found that pancreata from obese Zucker rats secrete significantly more insulin than do pancreata from lean Zucker rats. At a glucose stimulation of 300 mg/dL, the overall dynamic biphasic insulin secretory profiles from obese and lean rats were similar. Further studies to investigate the glucose-insulin dose response relationship in obese and lean rat pancreata demonstrated insulin hypersecretion by pancreata from obese rats which was particularly pronounced at normoglycemic and hypoglycemic levels (by factors as much as 14-fold). This hypersecretion is so striking as to suggest that in the intact state the obese animal may lack the ability to readily "shut off" its insulin secretion under normoglycemic conditions, whereas lean animals possess such an ability. Under hypoglycemic conditions (75 mg/dL), the hypersecretion is transient and insulin secretion returns to normal basal levels after 30 minutes of perfusion. Thus the degree to which this hypersecretory phenomenon may occur in vivo remains to be established.     

Effect of diazoxide on brain capillary insulin receptor binding and food intake in hyperphagic obese Zucker rats.

Insulin is believed to act as a central adiposity signal by binding to hypothalamic and other brain insulin receptors. Entry of circulating insulin into the brain is accomplished by a saturable receptor-mediated transendothelial transport system and is believed to be impaired in hyperinsulinemic, insulin-resistant, and hyperphagic obese Zucker rats. Theoretically, if hyperinsulinemia is decreased simultaneously while brain capillary insulin binding is increased, uptake of insulin into the brain can be enhanced leading to reduced food intake. To test this hypothesis, we administered diazoxide (DZ, 150 mg/kg/day), an inhibitor of glucose-mediated insulin secretion, or vehicle (control) to 7-week-old female obese and lean Zucker rats for 4 weeks (n = 24-28/subgroup-strain). Animals were assigned to either fasted (FD) or free-fed (FF) protocol for determination of plasma and cerebrospinal fluid (CSF) insulin and brain capillary insulin binding at the end of 4 weeks. DZ obese consumed fewer calories (P<0.01) and gained less weight than control obese (P<0.01), whereas DZ lean had similar amounts of caloric intake and weight gain compared with lean controls. DZ obese had lower fasting and random plasma glucose than control obese (P<0.05). FD and FF DZ-treated obese and lean rats had lower plasma insulin than their respective obese (P<0.01) and lean (P<0.01) controls. FD and FF DZ-treated obese rats demonstrated higher CSF insulin (P<0.05) and CSF/ plasma insulin ratio (P<0.01) than their controls, while only FF DZ lean animals showed higher CSF/plasma insulin ratio (P<0.01) than their controls (P<0.05). This was associated with enhanced brain capillary insulin binding in FD and FF DZ-treated obese (P<0.01) and lean (P<0.05) animals compared with their respective controls. It was concluded that DZ treatment in obese Zucker rats caused a decrease in insulin secretion and partially reversed impaired insulin binding to brain capillaries, leading to enhanced brain insulin uptake, and resulted in reduced food intake and weight gain observed in these animals.     

Chronic hyperglucagonemia in rats: effects on insulin, substrates, and hepatic enzymes of intermediary metabolism

The effects of chronic exogenous hyperglucagonemia were evaluated in 120–140 g rats given 0.8 or 0.4 mg glucagon per day for seven days by constant infusion through the use of subcutaneously implanted, osmotically driven minipumps. Untreated animals, with and without dietary restriction, served as controls. Glucagon infusion resulted in plasma glucagon concentrations ranging from 1000 to more than 4000 pg/mL. The experimental animals had decreased food intake resulting in poor weight gain. Plasma insulin, glucose, and β-hydroxybutyrate concentrations were unchanged in the experimental animals. Glucagon administration was associated with generalized hypoaminoacidemia (75% reduction from the control mean total amino acid concentration). Hepatic glycogen content decreased to the level seen in the dietary-restricted animals, which were matched for weight gain. In vitro activities of total hepatic glycogen phosphorylase, pyruvate kinase (measured at saturating PEP concentrations), acetyl-CoA carboxylase and fatty acid synthase were decreased in the experimental animals. Total glycogen synthase and PEP carboxykinase activities increased. These results are not consistent with previously described effects of glucagon mediated through enzyme phosphorylation and are interpreted as suggesting changes in total enzyme content. From these studies we conclude that chronic hyperglucagonemia of 1 week's duration in young rats has persistent metabolic effects. It does not, however, result in persistent changes in plasma glucose or β-hydroxybutyrate concentrations when insulin secretion is preserved. The observed hypoaminoacidemia is analogous to that seen in the glucagonoma syndrome. Long-term effects of glucagon on in vitro hepatic enzyme activities are possibly due to changes in enzyme protein content and not protein phosphorylation.     

Physiologic significance of glucocorticoids and insulin in the regulation of hepatic gluconeogenesis during starvation in rats

The physiologic significance of glucocorticoids and insulin in the regulation of hepatic gluconeogenesis was investigated during a 48-hr starvation period by studying the factors presumed to control the rate of glucose synthesis in the final gluconeogenetic pathway. Rats were used, in which glucorticoids were removed by adrenalectomy before starvation, and in which serum insulin was kept constant before and after food withdrawal by pre-feeding with a proteinfree diet. It was found that adrenalectomized rats at constantly low serum insulin levels responded to starvation as rapidly, and to the same degree, as intact control subjects (1) by a significant increase in plasma glucagon and, consequently, in hepatic cAMP concentration; (2) by a coordinate elevation of the activities of hepatic pyruvate carboxylase, P-enolpyruvate carboxykinase, and fructose-1,6-diphosphatase; (3) by systematic alterations in the concentration of effectors of gluconeogenetic key enzymes; (4) by a shifting of the cytoplasmic NAD system towards the reduced state; (5) by a decrease in the intrahepatic concentration of glycogenic precursor substrates. These results suggest that the hepatic gluconeogenic response to starvation with respect to the regulatory factors 1-5 occurs independently from changes in the concentration of plasma glucocorticoids and insulin. The crossing over of the gluconeogenetic intermediates between pyruvate and P-enolpyruvate (PEP), which was observed in intact but not in adrenalectomized rats, supports the assumption that during starvation, glucocorticoids enhance the rate of glucose production by the liver predominantly by permitting hepatic cAMP to stimulate the yet undefined mechanism, which has been demonstrated in the isolated perfused rat liver to control the substrate flow between pyruvate and PEP.     

高原环境对大鼠肝脏糖异生的影响

目的研究海拔4300 m高原环境对大鼠肝脏糖异生的影响及机制。方法健康成年大鼠36只,随机分为平原对照组(C组)、高原暴露1、3、7、15、30 d组(分别为H1、H3、H7、H15、H30组)。应用RT-PCR方法测定肝组织中葡萄糖-6-磷酸酶(G6Pase)、转录因子叉头框蛋白O1(Fox O1)的mRNA表达水平以及应用Western Blot法检测其蛋白表达水平,应用分光光度法测定肝糖原含量,使用自动生化分析仪检测血糖水平。多组间比较采用方差分析,进一步两两比较采用Tukey检验。结果与C组比较,(1)H1、H3和H7组肝组织中G6Pase的表达水平以及肝糖原含量显著增高(P值均0.05);(2)Fox O1的表达水平在H3、H7、H15及H30组中明显降低(P值均0.05);(3)各高原组血糖水平无明显变化。结论高原暴露急性期肝脏糖异生及糖原合成活跃,可能是高原习服的机制之一;腺苷酸活化蛋白激酶(AMPK)和Fox O1对高原暴露大鼠肝脏糖异生具有调节作用;糖原含量增加可能抑制了AMPK的活性。     

Role of glucagon suppression on gluconeogenesis during insulin treatment of the conscious diabetic dog

Summary In seven insulin-deficient (<3 mU/l) pancreatectomised dogs, the direct and glucagon-related indirect effects of intraportal insulin infusion (350 U/kg-min; 12±1 mU/l) on glucose production were determined. Insulin was infused for 300 min during which time the plasma glucagon concentration was allowed to fall (314±94 to 180±63 ng/l) for 150 min before being replaced by an infusion intraportally at 2.6ng/kg-min (323±61 ng/l) for the remaining 150 min. Glucose production and gluconeogenesis were determined using arterio-venous difference and tracer techniques. Insulin infusion shut off net hepatic glucose output and caused the plasma glucose, blood glycerol and plasma non-esterified fatty acid levels to fall. It caused the hepatic fractional extraction of alanine (0.41 ±0.10 to 0.21±0.06) and lactate (0.32 ±0.09 to 0.04 ±0.03) to fall which increased their concentrations. When glucagon was replaced, all of these changes were fully or partly reversed with the exception of the changes in glycerol and nonesterified fatty acids. Indeed, 70% of the fall in hepatic glucose production and virtually 100% of the changes in lactate and alanine metabolism produced by basal insulin infusion were mediated by a fall in glucagon. However, the fall in hepatic uptake of glycerol was unaffected by changes in glucagon and thus gluconeogenesis from this substrate was inhibited by insulin per se probably as a result of reduced lipolysis. The latter effect of insulin may explain the incomplete restoration of hepatic glucose production when hyperglucagonaemia was re-established during insulin infusion.     

Selective impairment of insulin signalling in the hypothalamus of obese Zucker rats

Aim/hypothesis By acting in the brain, insulin suppresses food intake. However, little is known with regard to insulin signalling in the hypothalamus in insulin-resistant states.Methods Western blotting, immunohistochemistry and polymerase chain reaction assays were combined to compare in vivo hypothalamic insulin signalling through the PI3-kinase and MAP kinase pathways between lean and obese Zucker rats.Results Intracerebroventricular insulin infusion reduced food intake in lean rats to a greater extent than that observed in obese rats, and pre-treatment with PI3-kinase inhibitors prevented insulin-induced anorexia. The relative abundance of IRS-2 was considerably higher than that of IRS-1 in hypothalamus of both lean and obese rats. Insulin-stimulated phosphorylation of IR, IRS-1/2, the associations of PI 3-kinase to IRS-1/2 and phosphorylation of Akt in hypothalamus were decreased in obese rats compared to lean rats. These effects seem to be mediated by increased phosphoserine content of IR, IRS-1/2 and decreased protein levels of IRS-1/2 in obese rats. In contrast, insulin stimulated the phosphorylation of MAP kinase equally in lean and obese rats.Conclusion/interpretation This study provides direct measurements of insulin signalling in hypothalamus, and documents selective resistance to insulin signalling in hypothalamus of Zucker rats. These findings provide support for the hypothesis that insulin could have anti-obesity actions mediated by the PI3-kinase pathway, and that impaired insulin signalling in hypothalamus could play a role in the development of obesity in this animal model of insulin-resistance.Abbreviations ERK extracellular signal-regulated kinase - Grb2 growth factor receptor binding protein 2 - IR insulin receptor - IRS insulin receptor substrate - MAPK mitogen-activated protein kinase - PI 3-kinase phosphatidylinositol 3-kinase - PKC Protein kinase C - Shc Src-homology and collagen homology - SHP2 Src-homology phosphatase 2 - TNF- Tumor-necrosis factor-     

Exenatide对肥胖大鼠肝脏糖异生基因表达的观察

目的 探讨Exenatide对肥胖大鼠肝脏糖异生基因表达的作用。方法将雄性SD大鼠随机分为高脂饮食（HF）组、高脂用药（HF＋Exe）组与正常对照（Nc）组。测定各组肝脏叉头蛋白转录因子O1（FoxO1）、磷酸烯醇式丙酮酸羧化酸（PEPCK）、葡萄糖-6-磷酸酶（G6Pase）mRNA,以及Fox01蛋白水平。结果HF组Fins、TG、FFA升高,Fox01、PEPCK、G6PasemRNA表达上调,Fox01蛋白水平增加（P〈0．05）;Exenatide用药8周后,HF＋Exe组Fins、TG、TC、FFA降低,Fox01、PEPCK、G6PasemRNA表达下调,FoxO1蛋白水平降低（P〈0．05）。结论Exenatide具有改善肥胖大鼠糖脂代谢的作用,其机制可能为抑制肝脏糖异生基因的表达。     

Serum cholesterol, lecithin-cholesterol acyltransferase, and hepatic hydroxymethylglutaryl coenzyme A reductase activities of lean and obese Zucker rats

Serum cholesterol concentrations, lecithin-cholesterol acyltransferase (LCAT), and hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activities of lean and obese Zucker rats were compared. The excess serum cholesterol of the female obese rat is found to be mainly free cholesterol associated with very low-density lipoproteins, whereas that of the male obese rat is carried as cholesterol esters associated with high-density lipoproteins. The high level of serum free cholesterol in the female obese rat is not due to a deficiency in lecithin-cholesterol acyltransferase activity. This enzyme activity is found to be elevated in the male obese rat. Hepatic HMG-CoA reductase activity declines as rats mature; this observation is most apparent in obese male rats. Lean rats exhibit the normal diurnal rhythm, but mature obese rats show little diurnal variation in HMG-CoA reductase activity. Obese female rats maintain high reductase activities, but the activities of obese male rats remain low at all times. Starvation suppresses liver HMG-CoA reductase and serum cholesterol in both lean and obese female rats. Thus, an increase in hepatic cholesterol synthesis may contribute to hypercholesterolemia in the obese female Zucker rat. On the other hand, factors such as nonhepatic synthesis or a decreased cholesterol catabolism may play more important roles in maintaining high serum cholesterol in the obese male Zucker rat.     

Mediation of beta-endorphin in exercise-induced improvement in insulin resistance in obese Zucker rats

BACKGROUND: Aerobic exercise including treadmill running has long been used to successfully treat and/or prevent insulin resistance and type-2 diabetes. Increase of plasma beta-endorphin is observed with exercise. The present study was designed to clarify the role of endogenous beta-endorphin in exercise-induced improvement in insulin resistance. METHODS: We used a moderate exercise program consisting of treadmill running at 20 m/min and 0% grade for 1 h/day, 7 days/week, for 8 weeks. Plasma glucose concentration was assessed by the glucose oxidase method. The enzyme-linked immunosorbent assay was performed to quantify the plasma level of beta-endorphin-like immunoreactivity (BER). The glucose disposal rate (GDR) was measured by the hyperinsulinemic euglycemic clamp technique. Changes of the insulin signaling in isolated soleus muscle were then detected by immunoprecipitation and immunoblotting. RESULTS: An increase of plasma BER in parallel with the reduction of plasma glucose was obtained in exercise-trained obese Zucker rats. Different from a marked reduction in sedentary obese rats, the value of insulin-stimulated GDR obtained from the exercised obese rats was reversed to near that of the sedentary lean group, eight weeks after the last period of exercise. This effect of exercise was inhibited by naloxone or naloxonazine at doses sufficient to block opioid micro-receptors. Signaling-related defects in the soleus muscle of sedentary obese Zucker rats, which impaired glucose transporter subtype 4 (GLUT 4), included decreased phosphorylation of insulin receptor substrate (IRS)-1, as well as an attenuated p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3 kinase) and Akt serine phosphorylation. In contrast, exercise training failed to modify the levels of insulin receptor (IR), IRS-1, and IR tyrosine autophosphorylation in obese Zucker rats. CONCLUSION: Enhanced insulin sensitivity via exercise training might be mediated by endogenous beta-endorphin through an increase of postreceptor insulin signaling related to the IRS-1-associated PI3-kinase step that leads to the enhancement of GLUT 4 translocation and improved glucose disposal in obese Zucker rats.     

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.     

Immunoreactive insulin levels are elevated in the cerebrospinal fluid of genetically obese Zucker rats

Immunoreactive insulin (IRI) concentrations were measured in plasma and cerebrospinal fluid (CSF) of four-month old genetically obese Zucker rats, their heterozygote lean littermates, and age-matched normal-weight Wistar rats. Basal plasma IRI was 201 + 35 microU/ml (means +/- SEM) in the obese animals and was significantly elevated compared to both lean Zucker rats (18 +/- 2.4 microU/ml, P less than 0.001) and Wistar rats (12 +/- 2.4 microU/ml, P less than 0.001). The mean CSF IRI concentration of fasted obese Zucker rats was 1.59 +/- 0.19 microU/ml; this was significantly higher than the CSF IRI level of either fasted Zucker lean rats (0.31 +/- 0.08 microU/ml, P less than 0.001) or Wistar rats (0.34 +/- 0.12 microU/ml, P less than 0.001). Plasma and CSF IRI concentrations were increased in free-feeding as compared with fasted animals. These data provide evidence that endogenous CSF insulin is derived from circulating plasma insulin in the rat and suggest that the hyperphagia and obesity of the Zucker fatty rat are not due to an inability of circulating insulin to gain access to the CSF.     

Insulin-dependent changes in subcellular distribution of liver insulin receptors in obese Zucker rats

Summary Acute hyperinsulinaemias induced by insulin and stimulants of insulin secretion have been shown to cause a translocation of liver insulin receptors from the cell surface to the intracellular compartment, with little or no change in total receptor number. To determine whether a similar phenomenon occurs in chronic hyperinsulinaemic states, we have carried out a longitudinal study of total, cell surface and intracellular liver insulin receptors in genetically obese Zucker rats, with spontaneously develop hyperinsulinaemia. Liver plasma membranes, Golgi-endosomal fractions, a microsomal fraction and a total particulate fraction were isolated in 2–14-week old obese (fa/fa) rats and examined for specific insulin binding relative to lean (Fa/?) age-matched animals. In 16-day old rats, which were still normoinsulinaemic, insulin binding was unchanged. Later on, as hyperinsulinaemia developed, three sequential changes in insulin binding activity were observed: first, a 25–30% increase in Golgiendosomal fractions (20 days); then, a 50–60% decrease in Golgi-endosomal fractions (4–5 weeks); and finally, a 50% decrease in plasma membranes (11 weeks), microsomal fraction and total particulate fraction (14 weeks), accompanied by restoration in Golgi-endosomal fractions (8–11 weeks). Unlike insulin receptors, insulin extractable from Golgiendosomal fractions at 4–5 weeks was unchanged or increased. We conclude that, although an early increase in the endocytosis of liver insulin receptors may occur in hyperinsulinaemic Zucker rats, this mechanism does not account for the later decrease in cell surface receptors observed in these animals.     

Glucagon stimulation of hepatic gluconeogenesis in rats fed a high-protein, carbohydrate-free diet

We previously observed increased gluconeogenesis in isolated liver of rats fed a high protein, carbohydrate-free (HP) diet and postulated that this was due to enhanced secretion of glucagon. To test this hypothesis, the effects of glucagon, dibutyryl cyclic AMP (DB-CAMP), and insulin on hepatic gluconeogenesis were examined. In perfused liver of control rats, alanine conversion to carbohydrate amounted to 20.39 ± 3.15 μmole/100 g body wt/90 min. Gluconeogenesis was increased to 35.14 ± 2.66 by glucagon; to 31.40 ± 1.0 by DB-CAMP, and suppressed to 14.56 + 1.15 by insulin. When both glucagon and insulin were added to the medium, only the stimulatory action of glucagon was evident. In perfused liver of HP-fed rats, alanine incorporation into carbohydrate totalled 51.79 ± 4.36. Gluconeogenesis was elevated by glucagon to 65.23 + 1.78, by DB-CAMP to 62.27 + 2.41, and was unaffected by insulin, 49.73 + 3.11. In the presence of both glucagon and insulin, only the glucagon effect was apparent. Levels of intermediary metabolites involved in gluconeogenesis were determined in control and HP livers. Concentrations of alanine, lactate, and pyruvate were reduced in HP liver, while phosphoenol-pyruvate (PEP) was increased, findings that suggest enhanced conversion of pyruvate to PEP. Fructose diphosphate concentration was normal; however, fructose-6-phosphate was greatly elevated, suggesting that fructose diphosphatase was stimulated by HP feeding. Similar changes in hepatic metabolites have been found following exposure of normal liver to glucagon. Plasma insulin values were not altered by HP feeding (C = 36.7 ± 3.7 μU/ml; HP = 32.3 ± 2.8). Plasma-glucagon levels were higher in HP rats than in controls (C = 80 ± 7.5 pg/ml; HP = 137.8 ± 16.7; p < 0.01). When the molar ratio of insulin to glucagon in plasma was calculated, the value in HP-fed rats was reduced, thus demonstrating a relative as well as an absolute rise of glucagon concentration (C = 11.1 ± 1.1; HP = 5.76 ± 0.6; p < 0.001). Liver cyclic AMP was greater in HP-fed rats than in controls (C = 0.83 ± 0.04 nmole/g; HP = 1.17 ± 0.07; p < 0.001), a finding that indicates that the liver was exposed to an increased amount of glucagon.