Whole body protein kinetics in women: effect of pregnancy and IDDM during anabolic stimulation

The effects of pregnancy and type 1 diabetes [insulin-dependent diabetes mellitus (IDDM)] on protein metabolism are still uncertain. Therefore, six normal and five IDDM women were studied during and after pregnancy, using [(13)C]leucine and [(2)H(5)]phenylalanine with a hyperinsulinemic-euglycemic clamp and amino acid infusion. Fasting total plasma amino acids were lower in pregnancy in normal but not IDDM women (2,631 +/- 427 vs. 2,057 +/- 471 and 2,523 +/- 430 vs. 2,500 +/- 440 micromol/l, respectively). Whole body protein breakdown (leucine) increased in pregnancy [change in normal (delta N) and IDDM women (delta D) 0.59 +/- 0.40 and 0.48 +/- 0.26 g. kg(-1). day(-1), both P < 0.001], whereas reductions in protein breakdown due to insulin/amino acids (delta N -0.57 +/- 0.19, delta D -0.58 +/- 0.20 g. kg(-1). day(-1), both P < 0.001) were unaffected by pregnancy. Protein breakdown in IDDM women was not higher than normal, and neither pregnancy nor type 1 diabetes altered the insulin sensitivity of amino acid turnover. Nonoxidized leucine disposal (protein synthesis) increased in pregnancy (delta N 0.67 +/- 0.45, delta D 0.64 +/- 0.34 g. kg(-1). day(-1), both P < 0.001). Pregnancy reduced the response of phenylalanine hydroxylation to insulin/amino acids in both groups (delta N -1.14 +/- 0.74, delta D -1. 12 +/- 0.77 g. kg(-1). day(-1), both P < 0.05). These alterations may enable amino acid conservation for protein synthesis and accretion in late pregnancy. Well-controlled type 1 diabetes caused no abnormalities in the regulation of basal or stimulated protein metabolism.     

Stoichiometry,Kinetics, and Regulation of Glucose and Amino Acid Metabolism of a Recombinant BHK Cell Line in Batch and Continuous Cultures

Batch and continuous cultures were carried out to study the stoichiometry, kinetics, and regulation of glucose and amino acid metabolism of a recombinant BHK cell line, with particular attention to the metabolism at low levels of glucose and glutamine. The apparent yields of cells on glucose and glutamine, lactate on glucose, and ammonium on glutamine were all found to change significantly at low residual concentrations of glucose (<5 mmol/L) and glutamine (<1 mmol/L) . The uptake rates of glucose and glutamine were markedly reduced at low concentrations, leading to a more effective utilization of these nutrients for energy metabolism and biosynthesis and reduced formation rates of lactate and ammonium. However, the consumption of other amino acids, especially the essential amino acids leucine, isoleucine, and valine and the nonessential amino acids serine and glutamate, was strongly enhanced at low glutamine concentration. Quantitatively, it was shown that the cellular yields and rates associated with glucose metabolism were primarily determined by the residual glucose concentration, while those associated with glutamine metabolism depended mainly on the residual glutamine. Both experimental results and analysis of the kinetic data with models showed that the glucose metabolism of BHK cells is not affected by glutamine except for a slight influence under glucose limitation and glutaminolysis not by glucose, at least not significantly under the experimental conditions. Compared to hybridoma and other cultured animal cells, the recombinant BHK cell line showed remarkable differences in terms of nutrient sensitivity, stoichiometry, and amino acid metabolism at low levels of nutrients. These cell-line-specific stoichiometry and nutrient needs should be considered when designing an optimal medium and/or feeding strategy for achieving high cell density and high productivity of BHK cells. In this work, a cell density of 1.1 × 10⁷ cells/mL was achieved in a conventional continuous culture by using a proper feed medium.     

Amino Acid Transport across the Tonoplast of Vacuoles Isolated from Barley Mesophyll Protoplasts : Uptake of Alanine, Leucine, and Glutamine

Mesophyll protoplasts from leaves of well-fertilized barley (Hordeum vulgare L.) plants contained amino acids at concentrations as high as 120 millimoles per liter. With the exception of glutamic acid, which is predominantly localized in the cytoplasm, a major part of all other amino acids was contained inside the large central vacuole. Alanine, leucine, and glutamine are the dominant vacuolar amino acids in barley. Their transport into isolated vacuoles was studied using ¹⁴C-labeled amino acids. Uptake was slow in the absence of ATP. A three- to sixfold stimulation of uptake was observed after addition of ATP or adenylyl imidodiphosphate an ATP analogue not being hydrolyzed by ATPases. Other nucleotides were ineffective in increasing the rate of uptake. ATP-Stimulated amino acid transport was not dependent on the transtonoplast pH or membrane potential. p-Chloromercuriphenylsulfonic acid and n-ethyl maleimide increased transport independently of ATP. Neutral amino acids such as valine or leucine effectively decreased the rate of alanine transport. Glutamine and glycine were less effective or not effective as competitive inhibitors of alanine transport. The results indicate the existence of a uniport translocator specific for neutral or basic amino acids that is under control of metabolic effectors.     

Defective insulin secretion in NIDDM: integral part of a multiplier hypothesis.

Non-insulin dependent diabetes mellitus (NIDDM) is characterized by a specific defect in glucose recognition by the pancreatic islet beta cell. This is in clear distinction to patients with insulin dependent diabetes mellitus (IDDM) who undergo pancreatic islet beta cell death and no longer have the ability to synthesize, store, and release insulin. Defective glucose-induced first phase insulin responses in patients with NIDDM can be partially restored by exogenous insulin treatment and by other pharmacologic therapy. These observations provide strength for the theory of glucose desensitization of the pancreatic beta cell as an important secondary defect in the pathogenesis of abnormal insulin secretion in NIDDM. However, even though defective insulin secretion is an essential part of the pathogenesis of NIDDM, in itself it is not sufficient. A multiplicative effect is required involving interaction between tissue resistance to insulin action and defective insulin secretion whose product is the syndrome of NIDDM.     

Amino acid regulation of insulin action in isolated adipocytes. Selective ability of amino acids to enhance both insulin sensitivity and maximal insulin responsiveness of the protein synthesis system

Using the number and concentration of amino acids in Dulbecco's modified Eagle's medium as reference (DMEM = 100%), we found that a maximally effective concentration of insulin (10 ng/ml) stimulated protein synthesis by 125% over basal rate in the presence of 50% amino acids (EC50 = 19%), but by only 48% in amino acid-free buffer. Moreover, time course experiments revealed that amino acid regulation of insulin action was very rapid (t1/2 of 9.5 min) and readily reversible (less than 30 min). This effect was specific in that basal rates of protein synthesis were unaltered by amino acids. A second effect of amino acids was to markedly enhance insulin sensitivity of the protein synthesis system in a dose-dependent manner. Thus, the half-maximally effective concentrations of insulin required to stimulate protein synthesis fell from 0.43 to 0.25 to 0.15 ng/ml in the presence of 0, 50, and 150% amino acids. Neither insulin sensitivity nor maximal insulin responsiveness of the glucose transport system was altered by amino acids, nor did amino acids affect the insulin binding capacity of cells. When we divided the 14 amino acids found in DMEM into two groups, we found that one group of 7 amino acids had little or no effect on insulin sensitivity or responsiveness, whereas the other group was fully active (a 157% increase in insulin responsiveness, ED50 of 0.21 ng/ml versus a 68% increase, ED50 of 0.51 ng/ml, with no amino acids). Isoleucine and serine together increased both insulin sensitivity and responsiveness to 60-70% of that seen with the full complement of amino acids. In conclusion: 1) amino acids modulate insulin action by enhancing maximal insulin responsiveness and insulin sensitivity of the protein synthesis system, and the regulatory site of amino acid action appears to be distal to the common signal pathway, within the insulin action-protein synthesis cascade, and 2) the effects of amino acids are specific, in that basal rates of protein synthesis are unaffected, only certain amino acids influence insulin action, and amino acids fail to alter insulin binding or the insulin-responsive glucose transport system. These studies, together with those in the companion paper, demonstrate that the pleiotropic actions of insulin on enhancing glucose uptake and protein synthesis are mediated through divergent pathways that can be independently regulated.(ABSTRACT TRUNCATED AT 400 WORDS)     

In vivo andin vitro studies of vanadate in human and rodent diabetes mellitus

In vivo vanadate and vanadyl have been shown to mimic the action of insulin and to be effective treatment for animal models of both Type I and Type II diabetes. The molecular mechanism of action of the vanadium salts on insulin sensitivity remains uncertain, and several potential sites proposed for the insulin-like effects are reviewed. In human trials, insulin sensitivity improved in patients with NIDDM, as well as in some patients with IDDM after two weeks of treatment with sodium metavanadate. This increase in insulin sensitivity was primarily due to an increase in non-oxidative glucose disposal, whereas oxidative glucose disposal and both basal and insulin stimulated suppression of hepatic glucose output (HGP) were unchanged. Clinically, oral vanadate was associated with a small decrease in insulin requirements in IDDM subjects. Of additional benefit, there was a decrease in total cholesterol levels in both IDDM and NIDDM subjects. Furthermore, there was an increase in the basal activities of MAP and S6 kinases to levels similar to the insulin-stimulated levels in controls, but there was little or no further stimulation with insulin was seen. Further understanding of the mechanism of vanadium action may ultimately be useful in the design of drugs that improve glucose tolerance.     

Improvement of glucose tolerance with immunomodulators in type 2 diabetic animals

Cytokine-inducers prevent insulin-dependent diabetes mellitus (IDDM) in animal models. We extended this therapy to non-insulin-dependent diabetes mellitus (NIDDM), because it was reported that diabetes of KK-Ay mice, a model for NIDDM, was recovered by allogenic bone-marrow transplantation that also prevented IDDM in animal models. An i.p. or i.v. injection of streptococcal preparation (OK 432) lowered fasting blood glucose (FBG) levels and markedly improved glucose tolerance test (GTT) in KK-Ay mice for more than 32 h regardless of the glucose loading routes (oral, i.v. or i.p.), while an i.v. injection of BCG improved FBG and GTT for more than 4 wks without body weight loss. The improvement of FBG and GTT with OK-432 was brought about in other NIDDM animals, GK rats and Wistar fatty rats. Among various cytokines possibly induced by OK-432 and BCG, IL-1α, TNFα and lymphotoxin significantly improved FBG and GTT in KK-Ay mice, whereas IL-2 and IFN_γ did not. There were no differences between the OK-432-treated KK-Ay mice and control in histology of the pancreas, degree of insulin-induced decrease in blood glucose levels, and muscle glycogen synthase activities. As to insulin secretion, there is a tendency that the OK-432-treatment less than 1 week did not affect insulin levels during GTT, whereas the treatment more than 2 weeks increased the insulin levels. Thus, cytokine-inducers improved FBG and glucose tolerance of NIDDM animals probably via cytokines. The results imply a role of the cytokines in glucose tolerance of NIDDM, although precise immune and metabolic mechanisms remain to be elucidated.     

Insulin action on protein metabolism in acromegalic patients

Insulin resistance in acromegaly causes glucose intolerance and diabetes, but it is unknown whether it involves protein metabolism, since both insulin and growth hormone promote protein accretion. The effects of acromegaly and of its surgical cure on the insulin sensitivity of glucose and amino acid/protein metabolism were evaluated by infusing [6,6-(2)H(2)]glucose, [1-(13)C]leucine, and [2-(15)N]glutamine during a euglycemic insulin (1 mU x kg(-1) x min(-1)) clamp in 12 acromegalic patients, six studied again 6 mo after successful adenomectomy, and eight healthy controls. Acromegalic patients, compared with postsurgical and control subjects, had higher postabsorptive glucose concentration (5.5 +/- 0.3 vs. 4.9 +/- 0.2 micromol/l, P < 0.05, and 5.1 +/- 0.1 micromol/l) and flux (2.7 +/- 0.1 vs. 2.0 +/- 0.2 micromol x kg(-1) x min(-1), P < 0.01, and 2.2 +/- 0.1 micromol x kg(-1) x min(-1), P < 0.05) and reduced insulin-stimulated glucose disposal (+15 +/- 9 vs. +151 +/- 18%, P < 0.01, and 219 +/- 58%, P < 0.001 from basal). Postabsorptive leucine metabolism was similar among groups. In acromegalic and postsurgical subjects, insulin suppressed less than in controls the endogenous leucine flux (-9 +/- 1 and -12 +/- 2 vs. -18 +/- 2%, P < 0.001 and P < 0.05), the nonoxidative leucine disposal (-4 +/- 3 and -1 +/- 3 vs. -18 +/- 2%, P < 0.01 and P < 0.05), respectively, indexes of proteolysis and protein synthesis, and leucine oxidation (-17 +/- 6% in postsurgical patients vs. -26 +/- 6% in controls, P < 0.05). Within 6 mo, surgery reverses insulin resistance for glucose but not for protein metabolism. After adenomectomy, more leucine is oxidized during hyperinsulinemia.     

THE EFFECT OF DIABETES, INSULIN AND WALLERIAN DEGENERATION ON LEUCINE METABOLISM OF ISOLATED RAT SCIATIC NERVES

Abstract– ¹⁴CO₂ production and ¹⁴C incorporation into proteins was studied in isolated rat sciatic nerves during incubation with 0.1 mM-[1-¹⁴C]leucine. Rats were made diabetic with streptozotocin. Nerves from diabetic rats incubated with glucose oxidized more [¹⁴C]leucine than controls. This difference was abolished in the presence of insulin (1 mU/ml). The effects of diabetes and insulin on leucine oxidation could not be demonstrated in the absence of glucose. Insulin stimulated the incorporation of [¹⁴C] from leucine into proteins by nerves from controls and diabetic rats.
Nerves undergoing Wallerian degeneration showed a marked increase in DNA content and stimulated incorporation of [¹⁴C]leucine into proteins. ¹⁴CO₂ production from leucine proceeded at 75% of the rate observed in intact nerves. Neither insulin nor diabetes affected leucine metabolism in degenerating nerves.
Neither the extracellular space nor the concentration of free amino acids were significantly different in nerves obtained from control and diabetic rats, except for lower glutamine content in the latter.
In vitro leucine metabolism of nerves is affected by diabetes, insulin and the integrity of the axon. The Schwann cell is suggested as a possible site of the observed changes in leucine metabolism.     

Metabolite profiles and the risk of developing diabetes

Emerging technologies allow the high-throughput profiling of metabolic status from a blood specimen (metabolomics). We investigated whether metabolite profiles could predict the development of diabetes. Among 2,422 normoglycemic individuals followed for 12 years, 201 developed diabetes. Amino acids, amines and other polar metabolites were profiled in baseline specimens by liquid chromatography-tandem mass spectrometry (LC-MS). Cases and controls were matched for age, body mass index and fasting glucose. Five branched-chain and aromatic amino acids had highly significant associations with future diabetes: isoleucine, leucine, valine, tyrosine and phenylalanine. A combination of three amino acids predicted future diabetes (with a more than fivefold higher risk for individuals in top quartile). The results were replicated in an independent, prospective cohort. These findings underscore the potential key role of amino acid metabolism early in the pathogenesis of diabetes and suggest that amino acid profiles could aid in diabetes risk assessment.     

Hepatic glycogen synthase phosphatase and phosphorylase phosphatase activities are increased in obese (fa/fa) hyperinsulinemic Zucker rats: effects of glyburide administration

The chronically hyperinsulinemic Zucker fatty rat, with peripheral insulin resistance and glucose intolerance, represents a model of noninsulin dependent diabetes mellitus (NIDDM). These animals have elevated hepatic glycogen levels. Hepatic levels of synthase phosphatase and phosphorylase phosphatase, which are diminished in the IDDM rat, were markedly increased in the obese rats. Glyburide, a sulfonylurea used in treatment of NIDDM, resulted in reduced levels of glycemia and increased insulin levels in Zucker rats. Hepatic glycogen levels were increased, as was the activation of glycogen synthase, although there were no effects of drug administration on synthase phosphatase or phosphorylase phosphatase activities. G6P levels were increased by glyburide in lean rats but not in obese animals. These effects of glyburide on liver glycogen metabolism are accounted for via potentiation of the glycogenic effects of insulin.     

Effects of insulin secretagogues on the secretion of insulin during thiamine deficiency

The mechanism by which glucose and other nutrient secretagogues induce the insulin secretion, is still controversial. Thiamine deficient rats, having a block in the glucose and branched chain amino acid metabolism at pyruvate and branched chain keto acids dehydrogenases respectively, were used to study the effects of insulin secretagogues. The levels of fasting blood glucose and serum insulin were estimated. Also, the serum insulin was assayed after intravenous administration of leucine, arginine and tolbutamide. The fasting blood glucose was increased and the serum insulin was decreased in thiamine deficiency. Leucine and arginine did not enhance insulin secretion in thiamine deficient animals. Tolbutamide induces the insulin secretion minimally in thiamine deficient rats. These results suggest that the nutrient secretagogues require an unimpaired glucose metabolism to induce insulin secretion.     

Misunderstanding in the classification of diabetes mellitus. What's in a name?

To assess whether physicians, residents, medical students, hospital diagnosis coders, and patients properly use the designations insulin-dependent diabetes mellitus (IDDM) and non-insulin-dependent diabetes mellitus (NIDDM) that were established by criteria of the National Diabetes Data Group, we reviewed clinic and hospital records and administered questionnaires. Although essentially all cases of true IDDM were identified as such and most cases of NIDDM not requiring insulin therapy were correctly identified by all groups, patients with NIDDM on insulin therapy were misidentified as having IDDM by 38% of residents in internal medicine clinics and 68% of primary care and surgical subspecialty residents. On a survey, of 22 patients with NIDDM on insulin therapy, 17 (77%) considered themselves to have IDDM. Thus, patients who have NIDDM by the established criteria who are on insulin therapy are commonly mislabeled as having IDDM. We present an approach for dealing with this problem by adapting nomenclature focusing on insulin deficiency and resistance. It would probably also be helpful to separately identify the subset of patients with "insulin-deficient diabetes" who are ketosis-prone. It is important to use immunologic profiling (islet cell antibody testing) and insulin sensitivity or deficiency testing (C-peptide levels).     

Amino acid degradation by the mesophilic sulfate-reducing bacterium Desulfobacterium vacuolatum

Desulfobacterium vacuolatum strain IbRM was able to grow using casamino acids as a source of carbon, energy and nitrogen. Growth was accompanied by utilization of several amino acids and sulfide production. Proline and glutamate were used preferentially and to the greatest extent. Glycine, serine and alanine were used more slowly and only after proline and glutamate were used. Isoleucine, valine, leucine and aspartate decrease was slowest and occurred in a linear fashion throughout the growth phase. Amino acids used from casamino acids, excluding aspartate, were also used as single carbon, energy and nitrogen sources. As a single amino acid, aspartate could only be used as a nitrogen source. Aspartate was not used as an electron acceptor. No growth occurred on any amino acid in the absence of sulfate. As single substrates, isoleucine, proline and glutamate were oxidized without formation of acetate and with molar yields of 13.1, 9.4 and 7.7 g mol^–1, respectively. Received: 24 June 1997 / Accepted: 10 September 1997     

Amino acid synthesis from glucose-U-14C in Glossina morsitans

Amino acid synthesis from glucose-U-¹⁴C was investigated in 2 day post-emergent and pregnant females of Glossina morsitans. This insect can synthesize alanine, aspartic acid, cystine, glutamic acid, glycine, proline, and serine from glucose. Arginine, histidine, hydroxyproline, isoleucine, leucine, lysine, methionine, phenylalanine, taurine, threonine, and valine showed no radioactivity and hence may be classified as nutritionally indispensable amino acids. Although tyrosine and hydroxyproline were not synthesized from glucose, they are at least partially dispensable nutrients for this insect because their synthesis from phenylalanine has been demonstrated. After the labelled glucose injection the highest radioactivity was recovered in the proline fraction. This is probably related to its rôle as an important energy reserve for flight. The radioactive amino acids recovered from females and from their offspring following glucose-U-¹⁴C injection were similar to those recovered from younger females. Radioactivity was also detected in the expired CO₂ and the excreta. The amino acids alanine, arginine, cystine, glycine, histidine, leucine/isoleucine, lysine, methionine, proline, and valine were identified in the excreta, of which arginine and histidine were in the largest amounts. Only excreted alanine, glycine, and proline showed radioactivity.     

Glucose oxidation and contents of free amino acids in pancreatic β-cells stimulated by a non-metabolizable leucine analogue

Oxidation of glucose and the contents of free amino acids were measured in β-cell-rich pancreatic islets exposed to stereoisomers of the non-metabolizable leucine analogue 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid. The insulin-releasing isomer b(−) appeared to stimulate glucose oxidation, whereas the inactive b(+) form was without effect. 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid reduced the islet contents of aspartic acid, γ-aminobutyric acid and glutamic acid and increased that of phenylalanine. Since the two isomers of 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid did not differ in their ability to alter the intracellular contents of free amino acids, these alterations are probably not significant for the regulation of insulin release. It could not be excluded that increased glucose metabolism may contribute to the stimulation of insulin release by the b(−) isomer of 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid.     

Amino acid metabolism in the Zucker diabetic fatty rat: effects of insulin resistance and of type 2 diabetes

We investigated amino acid metabolism in the Zucker diabetic fatty (ZDF Gmi fa/fa) rat during the prediabetic insulin-resistant stage and the frank type 2 diabetic stage. Amino acids were measured in plasma, liver, and skeletal muscle, and the ratios of plasma/liver and plasma/skeletal muscle were calculated. At the insulin-resistant stage, the plasma concentrations of the gluconeogenic amino acids aspartate, serine, glutamine, glycine, and histidine were decreased in the ZDF Gmi fa/fa rats, whereas taurine, alpha-aminoadipic acid, methionine, phenylalanine, tryptophan, and the 3 branched-chain amino acids were significantly increased. At the diabetic stage, a larger number of gluconeogenic amino acids had decreased plasma concentrations. The 3 branched-chain amino acids had elevated plasma concentrations. In the liver and the skeletal muscles, concentrations of many of the gluconeogenic amino acids were lower at both stages, whereas the levels of 1 or all of the branched-chain amino acids were elevated. These changes in amino acid concentrations are similar to changes seen in type 1 diabetes. It is evident that insulin resistance alone is capable of bringing about many of the changes in amino acid metabolism observed in type 2 diabetes.     

Preoperative oral supplementation with carbohydrate and branched-chain amino acid-enriched nutrient improves insulin resistance in patients undergoing a hepatectomy: a randomized clinical trial using an artificial pancreas

Glucose metabolism is adversely affected in patients following major surgery. Patients may develop hyperglycemia due to a combination of surgical stress and postoperative insulin resistance. A randomized trial was conducted to elucidate the effect of preoperative supplementation with carbohydrates and branched-chain amino acids on postoperative insulin resistance in patients undergoing hepatic resection. A total of 26 patients undergoing a hepatectomy for the treatment of a hepatic neoplasm were randomly assigned to receive a preoperative supplement of carbohydrate and branched-chain amino acid-enriched nutrient mixture or not. The postoperative blood glucose level and the total insulin requirement for normoglycemic control during the 16 h following hepatic resection were determined using the artificial pancreas STG-22. Postoperative insulin requirements for normoglycemic control in the group with preoperative nutritional support was significantly lower than that in the control group (P = 0.039). There was no incidence of hypoglycemia (<40 mg/dL) observed in patients, including those with diabetes mellitus, when the STG-22 was used to control blood glucose levels. STG-22 is a safe and reliable tool to control postoperative glucose metabolism and evaluate insulin resistance. The preoperative oral administration of carbohydrate and branched-chain amino acid-enriched nutrient is of clinical benefit and reduces postoperative insulin resistance in patients undergoing hepatic resection.     

Differential Contribution of Insulin and Amino Acids to the mTORC1-Autophagy Pathway in the Liver and Muscle

Autophagy is a highly inducible intracellular degradation process. It is generally induced by nutrient starvation and suppressed by food intake. Mammalian (or mechanistic) target of rapamycin complex 1 (mTORC1) is considered to be the major regulator of autophagy, but the precise mechanism of in vivo regulation remains to be fully characterized. Here, we examined the autophagy-suppressive effect of glucose, insulin, and amino acids in the liver and muscle in mice starved for 1 day. Refeeding after starvation with a standard mouse chow rapidly suppressed autophagy in both tissues, and this suppression was inhibited by rapamycin administration almost completely in the liver and partially in muscle, confirming that mTORC1 is indeed a crucial regulator in vivo. As glucose administration showed no major suppressive effect on autophagy, we examined the role of insulin and amino acids using hyperinsulinemic-euglycemic clamp and intravenous amino acid infusion techniques. Insulin administration showed a clear effect on the mTORC1-autophagy pathway in muscle, but had only a very weak effect in the liver. By contrast, amino acids were able to regulate the mTORC1-autophagy pathway in the liver, but less effectively in muscle. These results suggest that autophagy is differentially regulated by insulin and amino acids in a tissue-dependent manner.