Biomedical subjects

T Ramlal

Publications and source records attributed to T Ramlal.

53 records · Page 3Linked to original sources

PubMed1989

Insulin and glucose-dependent regulation of the glucose transport system in the rat L6 skeletal muscle cell line.

Differentiated rat L6 skeletal muscle cell cultures maintained in glucose-deficient medium containing 25 mM xylose displayed a rapid, reversible, time- and concentration-dependent 3-5-fold increase in glucose transport activity. Glucose deprivation in the continuous presence of insulin (24 h) resulted in an overall 9-10-fold stimulation of glucose transport activity. In contrast, acute (30 min) and chronic (24 h) insulin treatment of L6 cells maintained in high glucose (25 mM)-containing medium resulted in a 1.5- and 4-fold induction of glucose transport activity, respectively. Acute glucose deprivation and/or insulin treatment had no significant effect on the total amount of glucose transporter protein, whereas the long-term insulin- and glucose-dependent regulation of glucose transport activity directly correlated with an increase in the cellular expression of the glucose transporter protein. In situ hybridization of the L6 cells demonstrated a 3-, 4-, and 6-fold increase in glucose transporter mRNA induced by glucose deprivation, insulin, and glucose deprivation plus insulin treatments, respectively. Similarly, Northern blot analysis of total RNA isolated from glucose-deprived, insulin, and glucose-deprived plus insulin-treated cells resulted in a 4-, 3-, and 9-fold induction of glucose transporter mRNA, respectively. The continuous presence of insulin in the medium, either in the presence or absence of glucose, resulted in a transient alteration of the glucose transporter mRNA. The relative amount of the glucose transporter mRNA was maximally increased at 6-12 h which subsequently returned to the basal steady-state level within 48 h. These data demonstrate a role for insulin and glucose in the overall regulation of glucose transporter gene expression which may account for the alteration of glucose transporter activity of muscle tissue observed in pathophysiological states such as type II diabetes mellitus.

Animals↗

PubMed1989

Exercise-induced increase in glucose transporters in plasma membranes of rat skeletal muscle.

A previously developed technique for the isolation of plasma and intracellular membrane fractions from rat skeletal muscle was used to investigate transporter migration after insulin treatment or a bout of exercise (45 min of treadmill). Glucose-inhibitable cytochalasin-B binding was used to estimate the number of glucose transporters. Insulin and exercise caused increases in glucose uptake into the hindlimb muscles of 5- and 3-fold, respectively. Each stimulus also caused a 2-fold increase in the number of glucose transporters in plasma membranes prepared from hindlimb muscles. The insulin-induced increase in plasma membrane transporters was accompanied by a concomitant decrease in transporters from the intracellular pool. In contrast to insulin, there was no concomitant decrease in the number of cytochalasin-B-binding sites in the intracellular membrane fraction from exercised muscles. The ability of both insulin and exercise to increase the number of transporters in the plasma membrane is in accordance with recruitment of transporters as one cause of increased transport activity. However, the inability of exercise to decrease the number of transporters in the insulin-sensitive intracellular pool suggests the existence of either a second recruitable transporter pool or masked glucose transporters in the plasma membrane that are unmasked by the muscle contractile activity.

Animals↗

PubMed1989

Decrease in glucose transporter number in skeletal muscle of mildly diabetic (streptozotocin-treated) rats.

Diabetes is associated with a decrease in glucose uptake into muscle, the primary tissue responsible for whole body glucose uptake in the fed state. To study the basis of such a decrease we estimated the number of glucose transporters in skeletal muscle membranes from control and streptozotocin (STZ)-treated rats. Animals were injected with 65 mg STZ/kg and were clearly diabetic (hyperglycemic and glycosuric) at 1 week. After an overnight fast, animals were killed, and skeletal muscle from hind limbs were removed and used to prepare plasma membranes and internal membranes. The number of glucose transporters was determined by D-glucose-protectable equilibrium binding of [3H]cytochalasin-B. STZ-treated rats showed a 37% decrease in the number of glucose transporters per mg protein in crude membranes. The decrease was more pronounced in plasma membranes (average 50% decrease) than in the intracellular membranes (32% decrease). The reduction in the number of glucose transporters was specific, since it was not paralleled by changes in other plasma membrane markers or in total protein, although plasma membrane protein decreased by 15% in STZ-treated rats. When total recoveries of transporters were calculated (i.e. picomoles of transporters recovered per g tissue), the number of transporters in the plasma membrane fraction from STZ-treated rats was decreased by 68% relative to that in control animals. In the intracellular membranes and in total crude membranes from diabetic rats the transporters were decreased by 45%. This suggests that in STZ-treated rats there is an overall decrease in the number of glucose transporters, and that the plasma membrane is further specifically depleted of transporters. The decrease in glucose transporter number in the plasma membrane could at least in part be the cause of the diminished glucose uptake in diabetic muscle and for overall drop in total body glucose utilization of this condition.

Animals↗

PubMed1988

Insulin-mediated translocation of glucose transporters from intracellular membranes to plasma membranes: sole mechanism of stimulation of glucose transport in L6 muscle cells.

Plasma membranes and light microsomes were isolated from fused L6 muscle cells. Pre-treatment of cells with insulin did not affect marker enzyme or protein distribution in isolated membranes. The number of glucose transporters in the isolated membranes was calculated from the D-glucose-protectable binding of [3H]cytochalasin B. Glucose transporter number was higher in plasma membranes and lower in intracellular membranes derived from insulin-treated cells than in the corresponding fractions from untreated cells. The net increase in glucose transporters in plasma membranes was identical to the net decrease in glucose transporters in light microsomes (2 pmol/1.23 x 10(8) cells). The fold increase in glucose transporter number/mg protein in plasma membranes (2-fold) was similar to the fold increase in glucose transport caused by insulin. This suggests that recruitment of glucose transporters from intracellular membranes to the plasma membrane is the major mechanism of stimulation of hexose transport in L6 muscle cells. This is the first report of isolation of the two insulin-sensitive membrane elements from a cell line, and the results indicate that, in contrast to rat adipocytes, there is not change in the intrinsic activity of the transporters in response to insulin.

Biological Transport↗

PubMed1988

Inhibition by forskolin of insulin-stimulated glucose transport in L6 muscle cells.

The cardioactive diterpene forskolin is a known activator of adenylate cyclase, but recently a specific interaction of this compound with the glucose transporter has been identified that results in the inhibition of glucose transport in several human and rat cell types. We have compared the sensitivity of basal and insulin-stimulated hexose transport to inhibition by forskolin in skeletal muscle cells of the L6 line. Forskolin completely inhibited both basal and insulin-stimulated hexose transport when present during the transport assay. The inhibition of basal transport was completely reversible upon removal of the diterpene. In contrast, insulin-stimulated hexose transport did not recover, and basal transport levels were attained instead. This effect of inhibiting (or reversing) the insulin-stimulated fraction of transport is a novel effect of the diterpene. Forskolin treatment also inhibited the stimulated fraction of transport when the stimulus was by 4 beta-phorbol 12,13-dibutyrate, reversing back to basal levels. Half-maximal inhibition of the above-basal insulin-stimulated transport was achieved with 35-50 microM-forskolin, and maximal inhibition with 100 microM. Forskolin did not inhibit 125I-insulin binding under conditions where it caused significant inhibition of insulin-stimulated hexose transport. Forskolin significantly elevated the cyclic AMP levels in the cells; however its inhibitory effect on the above basal, insulin-stimulated fraction of hexose transport was not mediated by cyclic AMP since: (i) 8-bromo cyclic AMP and cholera toxin did not mimic this effect of the diterpene, (ii) significant decreases in cyclic AMP levels caused by 2',3'-dideoxyadenosine in the presence of forskolin did not prevent inhibition of insulin-stimulated hexose transport, (iii) isobutylmethylxanthine did not potentiate forskolin effects on glucose transport but did potentiate the elevation in cyclic AMP, and (iv) 1,9-dideoxyforskolin, which does not activate adenylate cyclase, inhibited hexose transport analogously to forskolin. We conclude that forskolin can selectively inhibit the insulin- and phorbol ester-stimulated fraction of hexose transport under conditions where basal transport is unimpaired. The results are compatible with the suggestions that glucose transporters operating in the stimulated state (insulin or phorbol ester-stimulated) differ in their sensitivity to forskolin from transporters operating in the basal state, or, alternatively, that a forskolin-sensitive signal maintains the stimulated transport rate.

1-Methyl-3-isobutylxanthine↗

PubMed1988

Insulin-induced decrease in 5'-nucleotidase activity in skeletal muscle membranes.

Insulin releases inositol phosphoglycans from myocytes in culture [(1986) Science 233, 967-972], which display insulinomimetic activity. Because 5'-nucleotidase is anchored to the membrane through inositol-containing phospholipid glycans, we investigated whether insulin could release the enzyme from the membrane. Membranes prepared from hindquarter muscles of rats perfused with insulin showed a 23% decrease in 5'-nucleotidase activity. Isolated membranes from muscle exposed to insulin in vitro also showed a small but reproducible decrease (9%) in 5'-nucleotidase activity relative to unexposed controls. Phospholipase C from Staphylococcus aureus released 60% of the membrane-bound 5'-nucleotidase. We propose that insulin may activate an endogenous phospholipase C that cleaves phospholipid-glycan-anchored proteins.

5'-Nucleotidase↗

PubMed1988

Stimulation of Na+/H+ exchange by insulin and phorbol ester during differentiation of 3T3-L1 cells. Relation to hexose uptake.

Acute exposure of 3T3-L1 undifferentiated fibroblasts to insulin or 4 beta-phorbol-12,13-dibutyrate (PDB) produced a moderate but significant stimulation of hexose transport (100% stimulation). In differentiated 3T3-L1 adipocytes, stimulation by insulin increased significantly (to 340%), while that by PDB remained at 130%. Total protein kinase C activity was 3-fold higher in 3T3-L1 fibroblast than adipocyte homogenates. PDB, but not insulin, induced migration of protein kinase C from the cytosol to the membrane, in both fibroblasts and adipocytes. Moreover, the hormone increased by 15% the protein kinase C activity of the cytosol. In 3T3-L1 fibroblasts, both insulin and PDB elicited a rapid (2 min lag) cytoplasmic alkalinization, measured with the fluorescent pH indicator bis-carboxyethyl carboxyfluorescein trapped in the cytoplasm. In 3T3-L1 adipocytes, PDB but not insulin elicited the cytoplasmic alkalinization. The alkalinization was prevented by amiloride or by replacing Na+ with either N-methylglucamine+ or K+. Stimulation of hexose transport by insulin or PDB was not affected by amiloride or Na+ substitution. It is concluded that: 1) Insulin and PDB have different effects on protein kinase C activity and subcellular distribution; 2) the responses of Na+/H+ exchange and hexose transport to insulin and PDB develop independently during differentiation of 3T3-L1 cells; 3) stimulation of Na+/H+ exchange and of hexose transport occur in parallel rather than in series in 3T3-L1 cells.

Animals↗

PubMed1987

Insulin-induced translocation of glucose transporters in rat hindlimb muscles.

Insulin causes a translocation of glucose transporters from intracellular microsomes to the plasma membrane in adipocytes. To determine whether insulin has a similar effect in rat hindlimb muscles, we used glucose-inhibitable cytochalasin B binding to estimate the number of glucose transporters in membrane fractions from insulinized and control muscles. Insulin treatment caused an approx. 2-fold increase in cytochalasin B-binding sites in a plasma membrane fraction and an approx. 70% decrease in cytochalasin B-binding sites in an intracellular membrane fraction. In order to detect this effect of insulin, it was necessary to develop a procedure for isolating a plasma membrane fraction and an intracellular membrane fraction that were not contaminated with sarcoplasmic reticulum. Our results show that, as in adipocytes, insulin stimulates translocation of glucose transporters from an intracellular membrane pool to the plasma membrane in hindlimb skeletal muscles.

Animals↗

PubMed1987

Cytoplasmic Ca2+ during differentiation of 3T3-L1 adipocytes. Effect of insulin and relation to glucose transport.

The cytoplasmic concentration of ionized Ca2+ [( Ca2+]i) was determined in 3T3-L1 cells during their differentiation from fibroblasts to adipocytes, suspended and loaded with the fluorescent Ca2+ indicators quin2 or indo-1. In undifferentiated fibroblasts, as well as in differentiated adipocytes up to day 9, [Ca2+]i was steady around 170 nM, and it increased significantly only in old adipocytes (day 12). During differentiation, stimulation of glucose uptake by insulin increased from a few percent to severalfold. Stimulation of uptake was already apparent after 10 min of addition of the hormone, and 10 nM insulin produced maximal stimulation in 30 min. Insulin (10(-6) M) added to quin2- or indo-1-loaded, suspended adipocytes had no detectable effect on [Ca2+]i for at least 10 min. In contrast, addition of the general anesthetic halothane increased [Ca2+]i from 172 to 251 nM in 3 min. In EGTA solution, the Ca2+ ionophore ionomycin elicited release of Ca2+ from intracellular stores that resulted in a transient increase in [Ca2+]i. A smaller but measurable Ca2+ release from intracellular stores (increasing [Ca2+]i by 20 nM) resulted upon addition of 20 micrograms/ml phosphatidic acid. In contrast, insulin did not produce any detectable release of Ca2+ from intracellular stores. Incubation of 3T3-L1 adipocytes with insulin in the presence of EGTA (the latter in excess over the Ca2+ concentration of the medium) did not prevent the stimulation of hexose uptake by the hormone, indicating that extracellular Ca2+ does not play a role in the insulin response. Furthermore, incubation of cells with quin2/AM in EGTA medium during exposure to insulin did not prevent stimulation of hexose uptake. Under these conditions it is demonstrated that intracellular quin2 suffices to chelate cytoplasmic Ca2+ even if releasable Ca2+ from intracellular stores were to pour into the cytoplasm. Thus, quin2 effectively lowers [Ca2+]i without impairing insulin action. It is concluded that insulin does not produce changes in [Ca2+]i and that chelating intracellular Ca2+ does not prevent stimulation of hexose uptake by insulin. These results suggest that it is unlikely that changes in [Ca2+]i may play a role in the transduction of information in insulin stimulation of glucose uptake in 3T3-L1 adipocytes.

Adipose Tissue↗

PubMed1987

Protein kinase C is not required for insulin stimulation of hexose uptake in muscle cells in culture.

The L6 skeletal muscle cell line has been identified as a suitable model to study the action of insulin on glucose uptake in muscle [Klip, Li & Logan (1984) Am. J. Physiol. 247, E291-E296]. The signals that transfer information from occupied insulin receptors to glucose transporters remain unknown. Here we report that activation of protein kinase C by exogenous phorbol esters results in stimulation of glucose uptake. Protein C kinase activity was induced to migrate from the cytosolic fraction to the microsomal fraction after 40 min of exposure of intact cells to 4 beta-phorbol 12,13-dibutyrate. In contrast, incubation with insulin did not alter the subcellular distribution of the kinase. Prolonged preincubation of L6 cells with phorbol esters resulted in depletion of kinase C activity, whereas neither the basal rate of glucose uptake nor its stimulation by insulin were affected. This suggests that protein kinase C is expressed in L6 cells, and that insulin stimulation of hexose transport does not involve protein kinase C.

Cell Line↗

PubMed1987

Selective increase in cytoplasmic calcium by anesthetic in lymphocytes from malignant hyperthermia-susceptible pigs.

Anesthetic-induced malignant hyperthermia in pigs and humans is characterized by muscle rigidity and rapid, often fatal, increases in body temperature. A defect in Ca2+ homeostasis has been suspected as underlying the disease, based on the preventive effect of dantrolene sodium, an agent thought to reduce Ca2+ levels in the cytoplasm. We describe here direct measurements of cytoplasmic ionized Ca2+ levels in lymphocytes from seven normal and 12 malignant hyperthermia-susceptible pigs, using the fluorescent indicator quin2. No differences in the concentration of cytoplasmic ionized Ca2+ were found in cells from malignant hyperthermia-susceptible pigs (160 +/- 10 nM) relative to the controls (150 +/- 10 nM). However, addition of halothane in vitro caused a significant increase (to 270 +/- 30 nM) in lymphocytes from malignant hyperthermia-susceptible pigs, but not from normal pigs (180 +/- 10 nM). The halothane-mediated increase in cytoplasmic ionized Ca2+ required extracellular Ca2+. It is suggested that general anesthetics such as halothane increase the permeability of the cell surface to Ca2+, and that this increase may, on its own or indirectly, increase the cytoplasmic level of ionized Ca2+ during a malignant hyperthermia crisis. The detection of a halothane-dependent increase in cytoplasmic ionized Ca2+ selectively in malignant hyperthermia-susceptible pigs could be the basis for a noninvasive test for malignant hyperthermia.

Aminoquinolines↗

PubMed1986

Insulin stimulation of glucose uptake and the transmembrane potential of muscle cells in culture.

The membrane potential of L6 muscle cells was measured with the fluorescent dye bis-oxonol. Hyperpolarizations of up to 15 mV were caused by gramicidin (in N-methyl-D-glucamine+ medium), or by monensin or ionomycin. Depolarization was achieved with gramicidin (in Na+ medium), or with K+. Insulin did not change the resting membrane potential of -70 mV, yet it effectively stimulated 2-deoxy-D-glucose uptake. Conditions that hyperpolarize the cells did not alter the basal rate of hexose uptake. Moreover, insulin was still capable of stimulating hexose uptake in depolarized cells. It is concluded that modulation of the membrane potential is probably not a signalling event in insulin stimulation of hexose uptake.

Biological Transport↗

PubMed1986

Voltage-dependent increase in ionized cytoplasmic calcium in the L6H9 muscle cell line detected with quin2.

The concentration of free Ca2+ ions in the cytoplasm ([Ca2+]i) is a key parameter in the function of muscle cells. This study describes the effect of membrane depolarization on [Ca2+]i in differentiating cells of the L6H9 line of rat skeletal muscle. [Ca2+]i was assessed using the fluorescent indicator quin2. In the presence of 1 mM extracellular Ca2+, [Ca2+]i averaged 250 nM. Replacement of extracellular Na+ with K+ resulted in cellular depolarization from -64 to -20 mV, measured with a fluorescent oxonol indicator. Depolarized cells showed a significant increase in [Ca2+]i, from 250 to 390 nM. The increase was prevented by nifedipine (5 microM) and was in great part dependent on the presence of extracellular Ca2+. A residual significant increase in [Ca2+]i was observed upon depolarization in Ca2+-free medium; this rise may be attributed to Ca2+ release from intracellular organelles. In the presence of extracellular Ca2+, replacement of extracellular Na+ by N-methylglucamine+ did not depolarize the cells, yet resulted in a significant increase in [Ca2+]i. This rise may be ascribed to inhibition or reversal of Na+/Ca2+ exchange activity due to the absence of extracellular Na+. The data are consistent with the presence of voltage-sensitive Ca2+ channels and Na+/Ca2+ antiporters at the cell surface, and of mechanisms of voltage-sensitive Ca2+ release from intracellular organelles.

Animals↗

PubMed1986

Changes in cytoplasmic free calcium caused by halothane. Role of the plasma membrane and intracellular Ca2+ stores.

Malignant hyperthermia is a muscle disease characterized by an abnormal response to anaesthetics, stress, and exercise. It is typified by muscle contracture and a dramatic elevation in body temperature. A defect in the regulation of the concentration of cytoplasmic free calcium, [Ca2]i, is thought to underlie this disease, but the actual [Ca2+]i was not measurable until recently. We have shown that the anaesthetic halothane increases [Ca2+]i in isolated lymphocytes from malignant hyperthermia-susceptible humans and pigs but not in the normal counterparts. In this report we extend these observations to a larger number of cases and analyze the molecular mechanisms responsible for the increase. The halothane-mediated rise in [Ca2+]i required external Ca2+ and was prevented by nifedipine, an inhibitor of the voltage-sensitive Ca2+ channels of the cell membrane. In addition, the effect of halothane on the releasable Ca2+ from intracellular stores was determined by measuring the size of the releasable pool before and after addition of the anaesthetic. After addition of halothane, about 73% of this Ca2+ pool was still available for release by the Ca2+ ionophore ionomycin in cells from normal humans and pigs. In contrast, only about 45% of the free Ca2+ in intracellular stores was left after treatment with halothane in cells from malignant hyperthermia-susceptible humans and swine. These results indicate that halothane acts both at the cell membrane and at intracellular organelles, and that this action results in a net increase in [Ca2+]i in malignant hyperthermia, but not in normal cells. The action at the cell membrane appears to be on the voltage-sensitive Ca2+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)

Aminoquinolines↗

PubMed1986

Insulin-induced cytoplasmic alkalinization and glucose transport in muscle cells.

Insulin stimulates glucose uptake into muscle within minutes, preceding stimulation of glycolysis. Signals involved in stimulation of glycolysis include cytoplasmic alkalinization and specific intracellular proteolytic products. In contrast, the signals that mediate stimulation of glucose transport remain unknown. Here we explore whether the insulin-induced cytoplasmic alkalinization is an early event that precedes activation of sugar uptake, whether such alkalinization is causally related to stimulation of sugar uptake, and whether proteolytic activity mediates stimulation of hexose transport. Cytoplasmic pH (pHi) was measured in suspended skeletal muscle cells of the L6H9 line with the intracellularly trapped fluorescent pH indicator bis(carboxyethyl)carboxy fluorescein. At 37 degrees C, insulin (1 X 10(-7) M) produced an increase in pHi of 0.11 units in 10 min. This increase became apparent 2 min after addition of the hormone, and maximal elevation of pHi was observed after 10 min, remaining elevated for up to 60 min. Removal of the hormone with anti-insulin antiserum did not reverse pHi back to the resting level. The alkalinization was prevented by amiloride, by 5-(N,N'-disubstituted)amiloride analogues, and by isosmotic replacement of Na+ with N-methylglucamine+ or choline+. This suggests that insulin activates Na+-H+ exchange. In contrast, stimulation of 2-deoxy-D-glucose transport by insulin was not affected by replacement of external Na+ or by addition of amiloride. Monensin, an exogenous Na+-H+ exchanger, did not stimulate sugar transport even though it increased pHi. Proteinase inhibitors that block hormonal stimulation of glycolysis were ineffective in preventing stimulation of 2-deoxy-D-glucose transport by insulin.(ABSTRACT TRUNCATED AT 250 WORDS)

Amiloride↗

PubMed1985

Subcellular distribution of dihydropyridine isothiocyanate binding in guinea-pig ileal smooth muscle.

The subcellular distribution of the 3H--2,6-dimethyl-3,5-dicarbomethoxy-4(2-isothiocyano) phenyl-1,4-dihydropyridine (DPSCN) binding to guinea-pig ileal smooth muscle was studied by subcellular fractionation. Initial experiments on subcellular fractionation of 3H-DPSCN-labelled tissues by differential centrifugation showed that there was an excellent correlation between the levels of the label present in a fraction and the plasma membrane marker phosphodiesterase I (r = 0.98) but not between the label and the putative endoplasmic reticulum marker NADPH: cytochrome-c-reductase (r = 0.56) or the inner mitochondrial marker cytochrome-c-oxidase (r = 0.36). Centrifugation of the microsomes on a continuous sucrose density gradient showed an excellent correlation of the migration of the label with phosphodiesterase I activity (r = 0.93) but not with the activities of NADPH: cytochrome-c-reductase (r = 0.66) or cytochrome-c-oxidase (r = 0.44). Treatment of microsomes with digitonin (1 mg/ml) followed by centrifugation on continuous sucrose density gradients increased the weighted mean densities of the phosphodiesterase activity (plasma membrane marker) and the labelling by similar magnitudes (0.04 to 0.06 g/ml). The weighted mean densities of NADPH: cytochrome-c-reductase and the cytochrome-c-oxidase were not altered significantly. It is concluded that in the guinea-pig ileal smooth muscle, DPSCN labels the plasma membrane specifically.

Animals↗

PubMedDate not supplied

Stimulation of glucose and amino acid transport and activation of the insulin signaling pathways by insulin lispro in L6 skeletal muscle cells.

The monomeric insulin analogue insulin lispro (Lys B28, Pro B29) is a rapid-acting insulin with a shorter duration of activity than human regular insulin. This compound has the advantage of reducing early postprandial hyperglycemia and the accompanying late hypoglycemia, thereby improving overall blood glucose control. To date, all published studies of the functional properties of insulin lispro have been conducted in whole animals. This study aimed to characterize the cellular actions of insulin lispro and the signals it elicits in an insulin-sensitive muscle cell line, L6 cells. Comparing the cellular actions of insulin lispro with those of human regular insulin, a number of observations were made. (1) Insulin lispro stimulated glucose and amino acid transport into L6 myotubes with a dose dependency and time course virtually identical to those of human regular insulin. (2) Insulin lispro was as effective as human regular insulin in stimulating time-dependent phosphorylation of insulin receptor substrate 1 (IRS-1), p70 ribosomal S6 kinase, and two isoforms of mitogen-activated protein kinase (ERK1 and ERK2). (3) Insulin lispro's ability to induce the association of IRS-1 with the p85 subunit of phosphatidylinositol 3-kinase was similar to that of human regular insulin. (4) As with human regular insulin, 100 nmol of the fungal metabolite wortmannin completely inhibited insulin lispro stimulation of glucose uptake. We concluded that the cellular actions of insulin lispro are similar to those of human regular insulin with respect to glucose and amino acid uptake and that the biochemical signals elicited are also comparable.

Amino Acids↗