The effects of calcium on protein turnover in skeletal muscles of the rat.

Several experimental procedures were used to increase the intracellular concentration of Ca2+ and determine its effects on protein turnover in isolated extensor digitorum longus and soleus muscle. These methods included the use of ionophore A23187, caffeine, dibucaine, thymol and procaine, all agents known to induce the release of calcium by acting either on the sarcolemma and/or on the sarcoplasmic reticulum. Another approach involved varying the external concentration of Ca2+ in the media in which the muscles were incubated. The changes in muscle Ca2+ concentrations after exposure to the various calcium-releasing agents were in keeping with accepted modes of action of these agents on muscle membranes. The findings suggest that increasing the sarcoplasmic concentration of Ca2+ inhibits protein synthesis and enhances protein breakdown. These catabolic effects of Ca2+ are compared with the changes induced in muscle protein turnover after exposure to insulin or cyclic nucleotides, and in myopathic muscle and situations of work overload. Attention is also drawn to some of the difficulties involved in definitively implicating Ca2+ as a factor involved in the normal regulation of protein turnover.     

Antioxidant alpha-lipoic acid and protein turnover in insulin-resistant rat muscle

We have shown previously that the antioxidant alpha-lipoic acid (ALA) can stimulate glucose transport and can enhance the stimulation of this process by insulin in skeletal muscle from insulin-resistant obese Zucker rats. As insulin can also acutely activate general protein synthesis and inhibit net protein degradation in skeletal muscle, we hypothesized that ALA could directly affect protein turnover and also increase the effect of insulin on protein turnover in isolated skeletal muscle from developing obese Zucker rats. In epitrochlearis muscles isolated from obese Zucker rats, insulin (2 mU/ml) significantly (p < 0.05) increased in vitro protein synthesis (phenylalanine incorporation into protein) and decreased net protein degradation (tyrosine release), whereas a racemic mixture of ALA (2 mM) had no effect on either process. Interestingly, rates of protein synthesis in muscle from obese Zucker rats were substantially lower compared to those values observed in age-matched insulin-sensitive Wistar rats, whereas rates of protein degradation were comparable. Obese Zucker rats were also treated chronically with either vehicle or ALA (50 mg/kg/d for 10 d). Again, insulin significantly increased net protein synthesis and decreased net protein degradation in epitrochlearis muscles isolated from vehicle-treated obese Zucker rats; however, this stimulatory effect of insulin was not improved by prior in vivo ALA treatment. These results indicate that the previously described effect of the antioxidant ALA to increase insulin-stimulated glucose transport in skeletal muscle of obese, insulin-resistant rats does not apply to another important insulin-regulatable process, protein turnover. These findings imply that the cellular mode of action for ALA is restricted to signaling factors unique to the activation of glucose transport, and does not involve the pathway of stimulation of general protein synthesis and net protein degradation.     

Superoxide-forming NADPH oxidase preparation of pig polymorphonuclear leucocyte.

The effect of corticosterone on myofibrillar protein breakdown in diabetic rats was investigated in order to assess the possible counteracting effects of the secondary rise in plasma insulin concentrations which normally accompanies such treatment. N^τ-Methylhistidine excretion, an index of myofibrillar protein breakdown, was compared before and after corticosterone treatment (4.0 mg/100 g body wt. per day) of normal control, adrenalectomized, 10-day-streptozotocin-diabetic and adrenalectomized diabetic rats. Diabetic rats received 1.5 units of insulin/100 g body wt. per day throughout the experiment and showed marked hyperglycaemia and glucosuria during corticosterone treatment, whereas non-diabetic rats had only mild hyperglycaemia but elevated insulin concentrations. Corticosterone treatment increased the average rate of myofibrillar protein breakdown by 68% and 95% respectively in non-diabetic and diabetic rats. Net loss of muscle non-collagen protein for the same 7-day period was greater in diabetic than in non-diabetic animals (4.15 versus 2.84% per day), and the calculated average synthesis rates were lowest in diabetic rats. Adrenalectomy had little effect except to decrease slightly the rate of muscle protein breakdown. These results show that the rise in plasma insulin concentrations that accompanies exogenous corticosterone administration to non-diabetic rats diminishes the catabolic effect of this glucocorticoid on muscle. Insulin appears to antagonize the effects of the glucocorticoid by attenuating the increased rates of myofibrillar protein breakdown and, to a lesser extent, by limiting the decrease in synthesis rates.     

Extremity hyperinsulinemia stimulates muscle protein synthesis in severely injured patients

Insulin has a well-recognized anabolic effect on muscle protein, yet critically ill, severely injured patients are often considered "resistant" to the action of insulin. The purpose of this study was to assess the in vivo effects of hyperinsulinemia on human skeletal muscle in severely injured patients. To accomplish this goal, 14 patients with burns encompassing >40% of their body surface area underwent metabolic evaluation utilizing isotopic dilution of phenylalanine, femoral artery and vein blood sampling, and sequential muscle biopsies of the leg. After baseline metabolic measurements were taken, insulin was infused into the femoral artery at 0.45 mIU.min(-1).100 ml leg volume(-1) to create a local hyperinsulinemia but with minimal systemic perturbations. Insulin administration increased femoral venous concentration of insulin (P < 0.01) but with only a 4% (insignificant) decrease in the arterial glucose concentration and a 7% (insignificant) decrease in the arterial concentration of phenylalanine. Extremity hyperinsulinemia significantly increased leg blood flow (P < 0.05) and the rate of muscle protein synthesis (P < 0.05). Neither the rate of muscle protein breakdown nor the rate of transmembrane transport of phenylalanine was significantly altered with extremity hyperinsulinemia. In conclusion, this study demonstrates that insulin directly stimulates muscle protein synthesis in severely injured patients.     

Effect of insulin on human skeletal muscle protein synthesis is modulated by insulin-induced changes in muscle blood flow and amino acid availability

Insulin promotes muscle anabolism, but it is still unclear whether it stimulates muscle protein synthesis in humans. We hypothesized that insulin can increase muscle protein synthesis only if it increases muscle amino acid availability. We measured muscle protein and amino acid metabolism using stable-isotope methodologies in 19 young healthy subjects at baseline and during insulin infusion in one leg at low (LD, 0.05), intermediate (ID, 0.15), or high (HD, 0.30 mUxmin(-1)x100 ml(-1)) doses. Insulin was infused locally to induce muscle hyperinsulinemia within the physiological range while minimizing the systemic effects. Protein and amino acid kinetics across the leg were assessed using stable isotopes and muscle biopsies. The LD did not affect phenylalanine delivery to the muscle (-9 +/- 18% change over baseline), muscle protein synthesis (16 +/- 26%), breakdown, or net balance. The ID increased (P < 0.05) phenylalanine delivery (+63 +/- 38%), muscle protein synthesis (+157 +/- 54%), and net protein balance, with no change in breakdown. The HD did not change phenylalanine delivery (+12 +/- 11%) or muscle protein synthesis (+9 +/- 19%), and reduced muscle protein breakdown (-17 +/- 15%), thus improving net muscle protein balance but to a lesser degree than the ID. Changes in muscle protein synthesis were strongly associated with changes in muscle blood flow and phenylalanine delivery and availability. In conclusion, physiological hyperinsulinemia promotes muscle protein synthesis as long as it concomitantly increases muscle blood flow, amino acid delivery and availability.     

Mechanism of insulin's anabolic effect on muscle: measurements of muscle protein synthesis and breakdown using aminoacyl-tRNA and other surrogate measures

Despite being an anabolic hormone in skeletal muscle, insulin's anticatabolic mechanism in humans remains controversial, with contradictory reports showing either stimulation of protein synthesis (PS) or inhibition of protein breakdown (PB) by insulin. Earlier measurements of muscle PS and PB in humans have relied on different surrogate measures of aminoacyl-tRNA and intracellular pools. We report that insulin's effect on muscle protein turnover using aminoacyl-tRNA as the precursor of PS and PB is calculated by mass balance of tracee amino acid (AA). We compared the results calculated from various surrogate measures. To determine the physiological role of insulin on muscle protein metabolism, we infused tracers of leucine and phenylalanine into 18 healthy subjects, and after 3 h, 10 subjects received a 4-h femoral arterial infusion of insulin (0.125 mUxkg(-1)xmin(-1)), while eight subjects continued with saline. Tracer-to-tracee ratios of leucine, phenylalanine, and ketoisocaproate were measured in the arterial and venous plasma, muscle tissue fluid, and AA-tRNA to calculate muscle PB and PS. Insulin infusion, unlike saline, significantly reduced the efflux of leucine and phenylalanine from muscle bed, based on various surrogate measures which agreed with those based on leucyl-tRNA (-28%), indicating a reduction in muscle PB (P < 0.02) without any significant effect on muscle PS. In conclusion, using AA-tRNA as the precursor pool, it is demonstrated that, in healthy humans in the postabsorptive state, insulin does not stimulate muscle protein synthesis and confirmed that insulin achieves muscle protein anabolism by inhibition of muscle protein breakdown.     

Skeletal muscle cells from insulin-resistant (non-diabetic) individuals are susceptible to insulin desensitization by palmitate.

We recently demonstrated that in vivo insulin resistance is not retained in cultured skeletal muscle cells. In the present study, we tested the hypothesis that treating cultured skeletal muscle cells with fatty acids has an effect on insulin action which differs between insulin-sensitive and insulin-resistant subjects. Insulin effects were examined in myotubes from 8 normoglycemic non-obese insulin-resistant and 8 carefully matched insulin-sensitive subjects after preincubation with or without palmitate, linoleate, and 2-bromo-palmitate. Insulin-stimulated glycogen synthesis decreased by 27 +/- 5 % after palmitate treatment in myotubes from insulin-resistant, but not from insulin-sensitive subjects (1.50 +/- 0.08-fold over basal vs. 1.81 +/- 0.09-fold, p = 0.042). Despite this observation, we did not find any impairment in the PI 3-kinase/PKB/GSK-3 pathway. Furthermore, insulin action was not affected by linoleate and 2-bromo-palmitate. In conclusion, our data provide preliminary evidence that insulin resistance of skeletal muscle does not necessarily involve primary defects in insulin action, but could represent susceptibility to the desensitizing effect of fatty acids and possibly other environmental or adipose tissue-derived factors.     

Altered responses in skeletal muscle protein turnover during aging in anabolic and catabolic periods

One of the most important effects of aging is sarcopenia, which is associated with impaired locomotion and general weakness. In addition, there is increased susceptibility to illness in aging, which often results in muscle wasting episodes. In such instances, the mobilization of muscle proteins provides free amino acids that are used for energetic purpose, the synthesis of acute phase proteins, and the immune response. However, since muscle protein mass is already depleted, the ability of the aged organism to recover from stress is impaired. Therefore, elucidating the mechanisms that result in sarcopenia is of obvious importance. Age-related changes in protein synthesis and proteolysis are rather small and our current methodology does not enable one to establish unequivocally whether sarcopenia results from depressed protein synthesis, increased proteolysis or both. By contrast, in anabolic and catabolic periods, a number of dysregulations in muscle protein turnover became clearly apparent. The aim of this review is to provide an overview of such altered responses to nutrients and catabolic treatments, which may ultimately contribute to explain sarcopenia. This includes impaired recovery in catabolic states, impaired anabolic effects of nutrients, in particular leucine, and a lack of regulation of the ubiquitin-proteasome proteolytic system. These alterations are discussed with respect to modifications in the insulin/IGF-1 axis and glucocorticoid related effects.     

Regulation of proteolysis during reloading of the unweighted soleus muscle

There is little information on the mechanisms responsible for muscle recovery following a catabolic condition. To address this point, we reloaded unweighted animals and investigated protein turnover during recovery from this highly catabolic state and the role of proteolysis in the reorganization of the soleus muscle. During early recovery (18 h of reloading) both muscle protein synthesis and breakdown were elevated (+65%, P<0.001 and +22%, P<0.05, respectively). However, only the activation of non-lysosomal and Ca(2+)-independent proteolysis was responsible for increased protein breakdown. Accordingly, mRNA levels for ubiquitin and 20S proteasome subunits C8 and C9 were markedly elevated (from +89 to +325%, P<0.03) and actively transcribed as shown by the analysis of polyribosomal profiles. In contrast, both cathepsin D and 14-kDa-ubiquitin conjugating enzyme E2 mRNA levels decreased, suggesting that the expression of such genes is an early marker of reversed muscle wasting. Following 7 days of reloading, protein synthesis was still elevated and there was no detectable change in protein breakdown rates. Accordingly, mRNA levels for all the proteolytic components tested were back to control values even though an accumulation of high molecular weight ubiquitin conjugates was still detectable. This suggests that soleus muscle remodeling was still going on. Taken together, our observations suggest that enhanced protein synthesis and breakdown are both necessary to recover from muscle atrophy and result in catch-up growth. The observed non-coordinate regulation of proteolytic systems is presumably required to target specific classes of substrates (atrophy-specific protein isoforms, damaged proteins) for replacement and/or elimination.     

Stimulation of phosphatidylcholine breakdown and diacylglycerol production by growth factors in Swiss-3T3 cells.

The effect of a number of growth factors on phosphatidylcholine (PtdCho) turnover in Swiss-3T3 cells was studied. Phorbol 12-myristate 13-acetate (PMA), bombesin, platelet-derived growth factor (PDGF) and vasopressin rapidly stimulated PtdCho hydrolysis, diacylglycerol (DAG) production, and PtdCho synthesis. Insulin and prostaglandin F2 alpha (PGF2 alpha) stimulated PtdCho synthesis, but not its breakdown, whereas epidermal growth factor (EGF) and bradykinin were without effect. Stimulation of PtdCho hydrolysis by the above ligands resulted in increased production of phosphocholine and DAG (due to phospholipase C activity) and significant amounts of choline, suggesting activation of a phospholipase D as well. CDP-choline and glycerophosphocholine levels were unchanged. Down-regulation of protein kinase C with PMA (400 nM, 40 h) abolished the stimulation of PtdCho hydrolysis and PtdCho synthesis by PMA, bombesin, PDGF and vasopressin, but not the stimulation of PtdCho synthesis by insulin and PGF2 alpha. PtdCho hydrolysis therefore occurs predominantly by activation of protein kinase C (either by PMA or PtdIns hydrolysis) leading to elevation of DAG levels derived from non-PtdIns(4,5)P2 sources. PtdCho synthesis occurs by both a protein kinase C-dependent pathway (stimulated by PMA, PDGF, bombesin and vasopressin) and a protein kinase C-independent pathway (stimulated by insulin and PGF2 alpha). DAG production from PtdCho hydrolysis is not the primary signal to activate protein kinase C, but may contribute to long-term activation of this kinase.     

Antisense oligonucleotides against the lipid phosphatase SHIP2 improve muscle insulin sensitivity in a dietary rat model of the metabolic syndrome

The lipid phosphatase SH2 domain-containing lipid phosphatase (SHIP2) has been implicated in the regulation of insulin sensitivity, but its role in the therapy of insulin-resistant states remains to be defined. Here, we examined the effects of an antisense oligonucleotide (AS) therapy directed against SHIP2 on whole body insulin sensitivity and insulin action in liver and muscle tissue in a dietary rodent model of the metabolic syndrome, the high-fat-fed (HF) rat. Whole body insulin sensitivity was examined in vivo by insulin tolerance tests before and after the intraperitoneal application of an AS directed against SHIP2 (HF-SHIP2-AS) or a control AS (HF-Con-AS) in HF rats. Insulin action in liver and muscle was assayed by measuring the activation of protein kinase B (Akt) and insulin receptor substrate (IRS)-1/2 after a portal venous insulin bolus. SHIP2 mRNA and protein content were quantified in these tissues by real-time PCR and immunoblotting, respectively. In HF-SHIP2-AS, whole body glucose disposal after an insulin bolus was markedly elevated compared with HF-Con-AS. In liver, insulin activated Akt similarly in both groups. In muscle, insulin did not clearly activate Akt in HF-Con-AS animals, whereas insulin-induced Akt phosphorylation was sustained in SHIP2-AS-treated rats. IRS-1/2 activation did not differ between the experimental groups. SHIP2 mRNA and protein content were markedly reduced only in muscle. In standard diet-fed controls, SHIP2-AS reduced SHIP2 protein levels in liver and muscle, but it had no significant effect on insulin sensitivity. We conclude that treatment with SHIP2-AS can rapidly improve muscle insulin sensitivity in dietary insulin resistance. The long-term feasibility of such a strategy should be examined further.     

Early development of protein metabolic perturbations in the liver and skeletal muscle of tumour-bearing rats. A model system for cancer cachexia.

In rats into which a fast-growing ascites hepatoma (Yoshida AH-130) had been transplanted, tumour growth elicited a marked loss of body weight until the animal's death in about 2 weeks. Overall tissue protein metabolism was simultaneously studied in vivo in the gastrocnemius muscle and liver after labelling with [14C]bicarbonate. Early and progressive atrophy developed in the gastrocnemius muscle, the underlying metabolic imbalance being expressed by an elevation in the apparent protein-degradation rate, with no changes in the apparent synthesis rate. A transient hyperplastic response preceded waste in the liver, both states being associated with alterations in protein-degradation rate: an initial decrease during liver growth, then an acceleration as liver regressed. Protein-synthesis rates, virtually unchanged during liver growth, were elevated in the subsequent phase, although not sufficient to balance the enhanced breakdown. Thus, in the tumour host tissues examined, altered states of protein turnover appeared to result mostly from changes in rates of protein breakdown. In sharp contrast with the negative protein balance in the host, the ascites hepatoma cells had the ability to grow or at least, in advanced stages, to maintain a stationary state.     

Skeletal-muscle growth and protein turnover.

Because of turnover, protein synthesis and breakdown can each be involved in the regulation of the growth of tissue protein. To investigate the regulation of skeletal-muscle-protein growth we measured rates of protein synthesis and breakdown in growing rats during development on a good diet, during development on a marginally low-protein diet and during rehabilitation on a good diet after a period of severe protein deficiency. Rates of protein synthesis were measured in vivo with a constant intravenous infusion of [14C]tyrosine. The growth rate of muscle protein was measured and the rate of breakdown calculated as breakdown rate=synthesis rate-growth rate. These measurements showed that during development on a good diet there was a fall with age in the rate of protein synthesis resulting from a fall in capacity (RNA concentration) and activity (synthesis rate per unit of RNA). There was a fall with age in the breakdown rate so that the rate was highest in the weaning rats, with a half-life of 3 days. There was a direct correlation between the fractional growth and breakdown rates. During rehabilitation on the good diet, rapid growth was also accompanied by high rates of protein breakdown. During growth on the inadequate diet protein synthesis rates were lesss than in controls, but growth occurred because of decreased rates of protein breakdown. This compression was not complete, however, since ultimate muscle size was only one-half that of controls. It is suggested that increased rates of protein breakdown are a necessary accompaniment to muscle growth and may result from the way in which myofibrils proliferate.     

Acute attenuation of translation initiation and protein synthesis by glucocorticoids in skeletal muscle

Glucocorticoids are diabetogenic factors that not only antagonize the action of insulin in target tissues but also render these tissues catabolic. Therefore, in rats, we endeavored to characterize the effects in skeletal muscle of glucocorticoids on translation initiation, a regulated process that, in part, governs overall protein synthesis through the modulated activities of eukaryotic initiation factors (eIFs). Four hours after intraperitoneal administration of dexamethasone (100 microg/100 g body wt), protein synthesis in skeletal muscle was reduced to 59% of the value recorded in untreated control animals. Furthermore, translation initiation factor eIF4E preferred association with its endogenous inhibitor 4E-BP1 rather than eIF4G. Dexamethasone treatment resulted in dephosphorylation of both 4E-BP1 and the 40S ribosomal protein S6 kinase concomitant with enhanced phosphorylation of eIF4E. Moreover, the guanine nucleotide exchange activity of eIF2B was unaffected as was phosphorylation of the alpha-subunit of eIF2. Hence glucocorticoids negatively modulate the activation of a subset of the protein synthetic machinery, thereby contributing to the catabolic properties of this class of hormones in vivo.     

Control of peptide-chain initiation in rat skeletal muscle. Development of methods for preparation of native ribosomal subunits and analysis of the effect of insulin on formation of 40 S initiation complexes

A method was developed for isolation of native ribosomal subunits from rat gastrocnemius muscle. Native 40 S subunits which were isolated by this method retained their associated nonribosomal proteins and consisted primarily of particles with equilibrium densities of 1.41 and 1.48 g/cm3. Based on the binding of radiolabeled Met-tRNAmeti, the 1.41 g/cm3 particle was identified as the 40 S initiation complex. Insulin deficiency in vivo resulting from either diabetes or fasting led to a 2-fold increase in 75 S monomers but had no effect on the numbers of native 40 and 60 S subunits or the relative distribution of the 1.41 and 1.48 g/cm3 particles. The rate of protein synthesis in perfused muscle preparations derived from insulin-deficient rats was reduced to about half the control value. Addition of insulin to the perfusate restored protein synthesis and 75 S monomers to control levels. The effect of insulin on protein synthesis was associated with a 1.5-fold increase in the amount of Met-tRNAmeti bound to the 1.41 g/cm3 particle. These findings identify formation of 40 S initiation complexes as a site of action of insulin on protein synthesis in skeletal muscle.     

Insulin and insulin antagonists evoke phosphorylation of P20 at serine 157 and serine 16 respectively in rat skeletal muscle

We show here that insulin and insulin antagonists differentially modify phosphorylation of three phospho-isoforms of P20 (termed S1, S2 and S3) in rat skeletal muscle. Precise phosphorylation sites of S1 and S2 were mapped to serine 157 and serine 16 respectively. Insulin evoked phosphorylation of P20 at serine 157 through the phosphatidylinositol (PI) 3-kinase pathway. Epinephrine and calcitonin gene-related peptide decreased phosphorylation at serine 157 and increased phosphorylation at serine 16 and other unidentified sites. These results demonstrate that the PI-3-kinase pathway of anabolic insulin and the cAMP pathway of catabolic hormones converge on P20 and suggest a potential role of this protein in regulating energy metabolism of skeletal muscle.     

Protein synthesis and amino acid accumulation during development in the rat: dissociation of diaphragm and heart muscle sensitivity to insulin.

Insulin is released into the circulation early during fetal life, but the physiological significance of the hormone in the fetus is still unclear. The present study was undertaken to evaluate the possible effects of insulin on the protein metabolism of skeletal and heart muscle during the perinatal period in the rat. When incubating hemidiaphragm in a bicarbonate buffer, 0.1 U/ml ox insulin failed to enhance the incorporation of leucine-4,5-3H into muscle protein before birth and during the immediate neonatal period. From 3 days of age onwards a stimulatory effect of insulin on protein synthesis was constantly observed. While insulin thus was ineffective in promoting protein synthesis in hemidiaphragms before the third postnatal day, it significantly enhanced the synthesis in heart muscle at an earlier stage of development. Insulin stimulated the uptake of alpha-aminoisobutyric acid-1-14C into the diaphragm muscle during late fetal life as well as through the early neonatal period. The role of insulin in protein metabolism during fetal life may thus be limited to the augmentation of amino acid transport in skeletal and heart muscle and to a stimulatory effect on protein synthesis only in certain tissue(s), e.g. heart muscle.     

Differential effects of insulin on peripheral and visceral tissue protein synthesis in neonatal pigs

We recently demonstrated in neonatal pigs that, with amino acids and glucose maintained at fasting levels, the stimulation of protein synthesis in longissimus dorsi muscle with feeding can be reproduced by a physiological rise in insulin alone. In the current report, we determine whether the response of protein synthesis to insulin in the neonatal pig is 1) present in muscles of different fiber types, 2) proportional in myofibrillar and sarcoplasmic proteins, 3) associated with increased translational efficiency and ribosome number, and 4) present in other peripheral tissues and in viscera. Hyperinsulinemic-euglycemic-amino acid clamps were performed in 7- and 26-day-old pigs infused with 0, 30, 100, or 1,000 ng. kg(-0.66). min(-1) of insulin to reproduce insulin levels present in fasted, fed, refed, and supraphysiological conditions, respectively. Tissue protein synthesis was measured using a flooding dose of L-[4-(3)H]phenylalanine. Insulin increased protein synthesis in gastrocnemius muscle and, to a lesser degree, masseter muscle. The degree of stimulation of protein synthesis by insulin was similar in myofibrillar and sarcoplasmic proteins. Insulin increased translational efficiency but had no effect on ribosome number in muscle. All of these insulin-induced changes in muscle protein synthesis decreased with age. Insulin also stimulated protein synthesis in cardiac muscle and skin but not in liver, intestine, spleen, pancreas, or kidney. The results support the hypothesis that insulin mediates the feeding-induced stimulation of myofibrillar and sarcoplasmic protein synthesis in muscles of different fiber types in the neonate by increasing the efficiency of translation. However, insulin does not appear to be involved in the feeding-induced stimulation of protein synthesis in visceral tissues. Thus different mechanisms regulate the growth of peripheral and visceral tissues in the neonate.     

Inhibitors of phospholipase A2 block the stimulation of protein synthesis by insulin in L6 myoblasts.

Insulin at a concentration close to the physiological range (100 mu-units/ml) stimulated protein synthesis in L6 myoblasts by 17%. Pre-treatment with the phospholipase A2 inhibitors mepacrine or dexamethasone prevented this stimulation and decreased the release of prostaglandin F2 alpha, implicating the action of phospholipase A2 and the subsequent metabolism of arachidonic acid to prostaglandins in the stimulation of protein synthesis by physiological doses of insulin. Higher concentrations of insulin (500-1000 mu-units/ml) stimulated protein synthesis in the presence of mepacrine or dexamethasone, suggesting that an alternative pathway may become important in insulin action when phospholipase A2 is inhibited.