Chip-scale dispersion engineering using chirped vertical gratings.

A strongly coupled, chirped Bragg grating made by sinusoidally modulating the sidewalls of a silicon waveguide is designed, fabricated, and experimentally characterized. By varying the device parameters, the operating wavelength, device bandwidth, sign (normal or anomalous), and magnitude of group-velocity dispersion may be engineered for specific photonic applications. Asymmetric Blackman apodization is best suited for maximizing the useable bandwidth while providing good ripple suppression. Dispersion values up to 7.0 x 10(5) ps/nm/km are demonstrated at 1.55 microm.

Processing advantages of linear chirped fiber Bragg gratings in the time domain realization of optical frequency-domain reflectometry.

The inclusion of a linear chirped fiber Bragg grating for short pulse dispersion is shown to enhance the time domain realization of optical frequency-domain reflectometry. A low resolution demonstrator is constructed with single surface scans containing 140 resolvable spots. The system dynamic range meets that shown in earlier demonstrations without digital post-processing for signal linearization. Using a conjugate pair of chirped pulses created by the fiber grating, ranging is performed with position and velocity information decoupled. Additionally, by probing the target with short pulses and introducing grating dispersion just before photodetection, velocity immune ranging is demonstrated.

Beta(3)-adrenoceptor agonist-induced increases in lipolysis, metabolic rate, facial flushing, and reflex tachycardia in anesthetized rhesus monkeys.

The effects of two beta(3)-adrenergic receptor agonists, (R)-4-[4-(3-cyclopentylpropyl)-4,5-dihydro-5-oxo-1H-tetrazol-1-yl]-N-[4-[2-[[2-hydroxy-2-(3-pyridinyl)ethyl]amino]ethyl]phenyl]benzenesulfonamide and (R)-N-[4-[2-[[2-hydroxy-2-(3-pyridinyl)- ethyl]amino]ethyl]phenyl]-1-(4-octylthiazol-2-yl)-5-indolinesulfonamide, on indices of metabolic and cardiovascular function were studied in anesthetized rhesus monkeys. Both compounds are potent and specific agonists at human and rhesus beta(3)-adrenergic receptors. Intravenous administration of either compound produced dose-dependent lipolysis, increase in metabolic rate, peripheral vasodilatation, and tachycardia with no effects on mean arterial pressure. The increase in heart rate in response to either compound was biphasic with an initial rapid component coincident with the evoked peripheral vasodilatation and a second more slowly developing phase contemporaneous with the evoked increase in metabolic rate. Because both compounds exhibited weak binding to and activation of rhesus beta(1)-adrenergic receptors in vitro, it was hypothesized that the increase in heart rate may be reflexogenic in origin and proximally mediated via release of endogenous norepinephrine acting at cardiac beta(1)-adrenergic receptors. This hypothesis was confirmed by determining that beta(3)-adrenergic receptor agonist-evoked tachycardia was attenuated in the presence of propranolol and in ganglion-blocked animals, under which conditions there was no reduction in the evoked vasodilatation, lipolysis, or increase in metabolic rate. It is not certain whether the beta(3)-adrenergic receptor-evoked vasodilatation is a direct effect of compounds at beta(3)-adrenergic receptors in the peripheral vasculature or is secondary to the release or generation of an endogenous vasodilator. Peripheral vasodilatation in response to beta(3)-adrenergic receptor agonist administration was not attenuated in animals administered mepyramine, indomethacin, or calcitonin gene-related peptide(8-37). These findings are consistent with a direct vasodilator effect of beta(3)-adrenergic receptor agonists.

Activation of insulin signal transduction pathway and anti-diabetic activity of small molecule insulin receptor activators.

We recently described the identification of a non-peptidyl fungal metabolite (l-783,281, compound 1), which induced activation of human insulin receptor (IR) tyrosine kinase and mediated insulin-like effects in cells, as well as decreased blood glucose levels in murine models of Type 2 diabetes (Zhang, B., Salituro, G., Szalkowski, D., Li, Z., Zhang, Y., Royo, I., Vilella, D., Diez, M. T. , Pelaez, F., Ruby, C., Kendall, R. L., Mao, X., Griffin, P., Calaycay, J., Zierath, J. R., Heck, J. V., Smith, R. G. & Moller, D. E. (1999) Science 284, 974-977). Here we report the characterization of an active analog (compound 2) with enhanced IR kinase activation potency and selectivity over related receptors (insulin-like growth factor I receptor, epidermal growth factor receptor, and platelet-derived growth factor receptor). The IR activators stimulated tyrosine kinase activity of partially purified native IR and recombinant IR tyrosine kinase domain. Administration of the IR activators to mice was associated with increased IR tyrosine kinase activity in liver. In vivo oral treatment with compound 2 resulted in significant glucose lowering in several rodent models of diabetes. In db/db mice, oral administration of compound 2 elicited significant correction of hyperglycemia. In a streptozotocin-induced diabetic mouse model, compound 2 potentiated the glucose-lowering effect of insulin. In normal rats, compound 2 improved oral glucose tolerance with significant reduction in insulin release following glucose challenge. A structurally related inactive analog (compound 3) was not effective on insulin receptor activation or glucose lowering in db/db mice. Thus, small molecule IR activators exert insulin mimetic and sensitizing effects in cells and in animal models of diabetes. These results have implications for the future development of new therapies for diabetes mellitus.

A selective human beta3 adrenergic receptor agonist increases metabolic rate in rhesus monkeys.

Activation of beta3 adrenergic receptors on the surface of adipocytes leads to increases in intracellular cAMP and stimulation of lipolysis. In brown adipose tissue, this serves to up-regulate and activate the mitochondrial uncoupling protein 1, which mediates a proton conductance pathway that uncouples oxidative phosphorylation, leading to a net increase in energy expenditure. While chronic treatment with beta3 agonists in nonprimate species leads to uncoupling protein 1 up-regulation and weight loss, the relevance of this mechanism to energy metabolism in primates, which have much lower levels of brown adipose tissue, has been questioned. With the discovery of L-755,507, a potent and selective partial agonist for both human and rhesus beta3 receptors, we now demonstrate that acute exposure of rhesus monkeys to a beta3 agonist elicits lipolysis and metabolic rate elevation, and that chronic exposure increases uncoupling protein 1 expression in rhesus brown adipose tissue. These data suggest a role for beta3 agonists in the treatment of human obesity.

Desensitization of beta3-adrenergic receptor-stimulated adenylyl cyclase activity and lipolysis in rats.

The beta3-adrenergic receptor is an integral membrane protein consisting of seven transmembrane domains. Unlike the beta1 and beta2 receptors, this subtype lacks the consensus phosphorylation sites required for desensitization by serine kinases. Using the rodent specific beta3 agonist BRL 35135, our initial data indicated that beta3 receptor-mediated glycerol levels progressively decreased following daily oral doses of 5 mg/kg. Therefore, we initiated studies designed to delineate the possible mechanism(s) for this decreased response. Within 3 hours following a single oral dose of BRL 35135, serum glycerol levels and UCP (uncoupling protein) RNA levels were significantly increased whereas beta3 RNA levels were significantly decreased. Rats were dosed daily for 5 days with either vehicle or BRL 35135 (5 mg/kg, p.o.) and blood samples were collected for glycerol analysis. Adipose tissue was excised for lipolysis and adenyl cyclase measurements. In addition, UCP and beta3 receptor RNA levels were assessed. No effect on adipocyte BRL 37344-stimulated adenylyl cyclase activity was observed 3 hours following the initial dose of BRL 35135. Although a slight decrease (approximately 25%) in adenylyl cyclase activity could be observed 24 hours following the initial dose, it wasn't until day 4 of dosing that a significant decrease (50%) was observed. In contrast, beta3- stimulated lipolysis in adipocytes from BRL 35135-treated rats was decreased 85% within 24 hours and this decrease persisted through four days of treatment. These data indicate that the lipolytic response to beta3 receptor activation is decreased after only a single oral dose of BRL 35135, whereas receptor-mediated adenylyl cyclase activation, although initially unaffected, also desensitizes by day four of treatment.

Thiazolidinediones produce a conformational change in peroxisomal proliferator-activated receptor-gamma: binding and activation correlate with antidiabetic actions in db/db mice.

The thiazolidinediones are novel insulin sensitizers that serve as orally active antidiabetic agents, in rodents, nonhuman primates, and man. We have examined the effects of 4-week oral administration of three thiazolidinediones (AD-5075, BRL 49653, and CS-045) on plasma glucose and triglyceride concentrations in obese hyperglycemic db/db mice. All three agents lower plasma glucose and triglyceride concentrations. Normal levels of glucose are achieved after treatment with AD-5075 (> 1.7 mg/kg) or BRL 49653 (> or = 30 mg/kg), whereas CS-045 (100 or 300 mg/kg) produces only modest reductions in either parameter. Although the thiazolidinediones have demonstrated insulin-sensitizing activities both in vivo and in vitro, their primary molecular target has been unclear. We have compared the in vivo antidiabetic actions described above with the in vitro activities on peroxisomal proliferator-activated receptor-gamma (PPAR gamma). Hamster PPAR gamma 1 was transiently expressed in COS-1 cells to study the binding of [3H]AD-5075. The concentrations of compounds needed to displace radiolabeled AD-5075 from PPAR gamma correlate with their in vivo potency; the Ki values for displacement by cold AD-5075, BRL 49653, and CS-045 are 22, 68, and 1600 nM, respectively. To examine activation of the receptor, it was transiently cotransfected into COS-1 cells with a reporter plasmid containing two copies of a peroxisome proliferator response element. The EC50 values for activation are 2, 6, and 140 nM for AD-5075, BRL 49653, and CS-045, respectively. We have also analyzed limited proteolytic digests of in vitro translated hamster PPAR gamma. The thiazolidinediones produce a conformational change in PPAR gamma analogous to those produced by agonists of other nuclear hormone receptors. In the presence of saturating concentrations of either AD-5075 or BRL 49653, a receptor fragment of 27 kDa is protected from proteolysis by trypsin. These data support the conclusion that the antidiabetic actions of the thiazolidinediones are directly mediated through binding to PPAR gamma and the resulting active conformation of the receptor. Therefore, binding and transactivation assays using PPAR gamma should serve to identify other novel therapeutic agents with potential antidiabetic activities.

A novel insulin secretagogue is a phosphodiesterase inhibitor.

The arylpiperazine L-686,398 was described as an oral hypoglycemic agent and is shown to be an insulin secretagogue in vitro. The characteristics of its activity were similar to those of the incretin glucagon-like peptide I (GLP-I). We demonstrate that both the peptide and L-686,398 increase the accumulation of cAMP in isolated ob/ob mouse pancreatic islet cells, but by different mechanisms. Although GLP-I activates adenylate cyclase, the arylpiperazine has no effect on this enzyme or on the binding of 125I-labeled GLP-I to its receptor on RINm5F rat insulinoma cell membranes. However, L-686,398 inhibits the total cAMP phosphodiesterase (PDE) activity in homogenates of ob/ob mouse pancreatic islets with an EC50 of approximately 50 mumol/l. To determine the mechanism of PDE inhibition by the arylpiperazine and to examine its specificity, we studied the kinetics of arylpiperazine inhibition of two recombinant PDEs. The arylpiperazine is a competitive inhibitor of both a human heart type III PDE and a rat type IV-D PDE. Inhibition of the type III and IV isozymes are characterized by Ki values of 27 and 5 mumol/l, respectively. Although not extremely potent, the arylpiperazine does exhibit modest selectivity between these PDEs. The observation that L-686,398 acts as a PDE inhibitor suggests that exploration for beta-cell-specific PDE isoforms may reveal novel PDEs as targets for the development of therapeutically useful glucose-dependent insulin secretagogues.

Glucose-dependent alterations of intracellular free calcium by glucagon-like peptide-1(7-36amide) in individual ob/ob mouse beta-cells.

Depolarizing concentrations of glucose produce characteristic alterations of intracellular free Ca2+ ([Ca2+]i) in pancreatic beta-cells. The effects of the proposed incretin, glucagon-like peptide-1(7-36amide) (GLP-1a) on [Ca2+]i were determined from Fura-2 fluorescence ratio imaging of cultured ob/ob mouse pancreatic beta-cells. In control cells, [Ca2+]i is low in 3 mM glucose; increasing [glucose] to 8-12 mM results in an initial dip in [Ca2+]i followed by slow oscillating increases in [Ca2+]i. GLP-1a (0.03-10,000 pM) does not alter [Ca2+]i in 3 mM glucose, but does change the response to elevated glucose (8-12 mM). The time integral of the initial dip is reduced ([GLP-1a] 10-100 pM), and the integral of the [Ca2+]i signal is increased ([GLP-1a] > or = 1 pM). GLP-1a increases the frequency of sustained, stable plateau responses to elevated glucose, and the frequency of large, rapid spikes of increased [Ca2+]i associated with either plateaus, or oscillations. Application of a cAMP analog mimics most of the actions of GLP-1a. Activation of the GLP-1a receptor, or application of cAMP alters pancreatic beta-cell [Ca2+]i only when [glucose] is high.

Synthesis and hypoglycemic activity of substituted 8-(1-piperazinyl)imidazo[1,2-a]pyrazines.

A series of alkyl- and halo-substituted 8-(1-piperazinyl)imidazo[1,2-a]pyrazines were prepared using two approaches, the condensation of alpha-halocarbonyl derivatives with an aminopyrazine or the oxidation-dehydration of a [(beta-hydroxyalkyl)amino]pyrazine. These imidazo[1,2-a]pyrazines were evaluated for their binding affinity to the alpha 1, alpha 2, beta 1, and beta 2 adrenergic receptors as well as their ability to lower blood glucose in insulin resistant hyperglycemic ob/ob mice. Modifications on 8-(1-piperazinyl)imidazo[1,2-a]pyrazine (4) reduced alpha 2 binding, lowered hypoglycemic potency, and showed variations in binding to the alpha 1, beta 1, and beta 2 adrenergic receptors. In addition to 4, the 2-methyl, 3-methyl, and 5-methyl 8-(1-piperazinyl)imidazo[1,2-a]pyrazines (16k, 25m, and 16f, respectively) displayed high affinity for the alpha 2 receptor and were potent hypoglycemic agents when compared to 2-amino-7,8-dihydro-4-(1-piperazinyl)-6H-thiopyrano[3,2- d]pyrimidine (MTP-1403, 2). Receptor binding was modified by use of a 4-methylpiperazine moiety which reduced alpha 1 and beta 1 binding while retaining some hypoglycemic activity. The structure-activity relationship for heterocyclic alkyl and halo substitution on biological activity is discussed.

The function but not the expression of rat liver inhibitory guanine nucleotide binding protein is altered in streptozotocin-induced diabetes.

Adenylate cyclase activity was examined as a measure of inhibitory guanine nucleotide binding protein (Gi) function in liver plasma membranes from rats made chemically diabetic by streptozotocin (STZ) treatment. Clonidine activation of the alpha 2 adrenergic receptor, which activates Gi, inhibited forskolin--stimulated adenylate cyclase activity in control membranes. However, there was no effect on adenylate cyclase activity in membranes from STZ diabetic animals. Also, a polyclonal antipeptide antibody was raised to a highly conserved segment of the Gi alpha 2 subunit. This antibody specifically recognizes a 41 kilodalton protein, is blocked by an excess of peptide, does not recognize the alpha-subunit of transducin, and immunoprecipitates a 41 kilodalton protein which was ADP-ribosylated by pertussis toxin. Immunoblots using this antibody detect no difference between normal and STZ diabetic animals in the level of liver plasma membrane Gi expression. Therefore, STZ-induced diabetes altered the function of Gi but had no effect on Gi expression.

Vanadate treatment of streptozotocin diabetic rats restores expression of the insulin-responsive glucose transporter in skeletal muscle.

Streptozotocin-treated rats were diabetic, as assessed by blood glucose and plasma insulin values, while vanadate treatment restored blood glucose values to normal. Immunoblot analysis using a monoclonal antibody to the insulin-responsive glucose transporter demonstrated a 70% decline in transporter expression in skeletal muscle of diabetic rats. Subsequent treatment of diabetic animals with vanadate resulted in renewed expression of the transporter to 87% of control levels. Northern blot analysis of total skeletal muscle RNA from diabetic animals revealed a 55% decline in the steady state level of muscle glucose transporter mRNA, while vanadate treatment led to a 187% increase in transporter mRNA over normal levels. These results support the conclusion that vanadate acts to relieve diabetic hyperglycemia by inducing expression of the insulin-responsive glucose transporter at the pretranslational level.

Effects of an alpha 2-adrenoceptor antagonist on glucose tolerance in the genetically obese mouse (C57BL/6J ob/ob).

This study examines the effects of a relatively selective alpha 2-adrenoceptor antagonist, 8-(L-piperazinyl)imado-[1,2-alpha] pyrazine (compound A), and the preferential alpha 2-agonist clonidine on blood glucose, glucose tolerance, and plasma insulin levels in the C57BL/6J ob/ob mouse and its lean littermate. While clonidine raised blood glucose levels and impaired glucose tolerance, oral administration of compound A resulted in decreased blood glucose levels, as well as improved glucose tolerance in ob/ob mice. Insulin levels in ob/ob mice treated with clonidine were significantly reduced, while compound A raised insulin levels threefold and blocked the effects of clonidine when co-administered to the same animals. Clonidine-induced hyperglycemia in lean littermates was not accompanied by a decrease in insulin levels, while a small but significant increase in insulin levels was observed by compound A administration. Glycogen synthesis in diaphragm of ob/ob mice was enhanced after oral administration of compound A and was accompanied by an increase in plasma insulin levels. Concomitant treatment with a potent somatostatin analog to inhibit insulin release blocked the effects of the alpha 2-adrenoceptor antagonist, compound A. These observations suggest that the alpha 2-antagonist studied, increased plasma insulin levels with an accompanying reduction in blood glucose and an improvement in glucose tolerance in a genetic model of insulin resistance. Differential sensitivity to alpha 2-agonist in these genetically obese mice, ob/ob, was demonstrated by decreased insulin levels due to clonidine administration.

The effect of ponalrestat on sorbitol levels in the lens of obese and diabetic mice.

Sorbitol levels were determined in lens of genetically obese (ob/ob) and diabetic (db/db) mice, as well as in lean mice (+/db, +/ob) made diabetic by administration of streptozotocin (STZ). Treatment of lean mice with STZ resulted in hypoinsulinemia, whereas the ob/ob and db/db mice were hyperinsulinemic. Hyperglycemia was present in STZ-treated +/db and +/ob mice and in db/db mice, whereas relative euglycemia was observed in ob/ob mice and untreated +/db and +/ob mice. Sorbitol levels were elevated in lens tissue of db/db mice and STZ-treated +/db. In contrast, no changes in sorbitol content were observed in ob/ob mice and +/ob mice treated with STZ, suggesting that aldose reductase activity in lens of this animal model is considerably less than that present in db/db mice. Oral treatment of db/db mice and STZ-treated +/db mice with Ponalrestat reduced hyperglycemia-induced sorbitol accumulation significantly in lens, indicating that aldose reductase inhibitors may ameliorate long-term complications associated with sorbitol accumulation in diabetes.

Decreased expression of the insulin-responsive glucose transporter in diabetes and fasting.

Cellular resistance to insulin caused by a reduction in insulin-mediated glucose uptake can be produced in rats by chemically inducing diabetes with streptozotocin and by fasting. Two glucose transporter isoforms are expressed in fat cells: (1) the insulin-responsive species which is found only in fat and muscle, and (2) a species corresponding to the erythrocyte/Hep G2/rat brain transporter. We show here that fat cells isolated from streptozotocin diabetic rats and from fasted rats show a significant (60-80%) decrease in the amount of immunologically detectable insulin-sensitive glucose transporter and no change in the level of the Hep G2/rat brain transporter. Administration of insulin and refeeding, respectively, result in a return of the insulin-sensitive glucose transporter to levels that are normal or slightly above normal. Thus, peripheral tissue insulin resistance could be due to the specific reduction in the amount of insulin-sensitive glucose transporter.

Impaired insulin-like growth factor I-mediated stimulation of glucose incorporation into glycogen in vivo in the ob/ob mouse.

The ability of insulin to modulate glucose metabolism is impaired in insulin resistant ob/ob mice. It has been shown that insulin-like growth factor I stimulates the uptake and metabolism of glucose in muscle through the insulin-like growth factor receptor not the insulin receptor. Thus, we have compared the abilities of insulin-like growth factor I and insulin to stimulate the in vivo incorporation of [14C]-glucose into glycogen in the diaphragm of ob/ob mice and their lean littermates. The animals used in these studies were 12-14 weeks old and the serum insulin levels of the ob/ob mice were 16-fold higher than in their lean littermates. There were no differences in the serum levels of glucose or insulin-like growth factor I. Both insulin and insulin-like growth factor I stimulate the incorporation of [14C]-glucose into glycogen in lean mice. Significant stimulation occurs at doses as low as 1 micrograms/kg of either peptide. The effective doses of insulin and insulin-like growth factor I are quite similar, which indicates that the effect of insulin-like growth factor I is mediated by the insulin-like growth factor receptor and not the insulin receptor. In contrast, greater than 100 micrograms/kg of insulin-like growth factor I is required to stimulate [14C]-glucose incorporation into glycogen in the diaphragm of ob/ob mice. Thus, ob/ob mice are resistant to the action of both insulin and insulin-like growth factor I.(ABSTRACT TRUNCATED AT 250 WORDS)

Design of a selective insulin receptor tyrosine kinase inhibitor and its effect on glucose uptake and metabolism in intact cells.

An inhibitor of the insulin receptor tyrosine kinase (IRTK), (hydroxy-2-naphthalenyl-methyl) phosphonic acid, was designed and synthesized and was shown to be an inhibitor of the biological effects of insulin in vitro. With a wheat germ purified human placental insulin receptor preparation, this compound inhibited the insulin-stimulated autophosphorylation of the 95-kDa beta-subunit of the insulin receptor (IC50 = 200 microM). The ability of the kinase to phosphorylate an exogenous peptide substrate, angiotensin II, was also inhibited. Half-maximal inhibition of basal and insulin-stimulated human placental IRTK activity was found at concentrations of 150 and 100 microM, respectively, with 2 mM angiotensin II as the peptide substrate. The inhibitor was found to be specific for tyrosine kinases over serine kinases and noncompetitive with ATP. The inhibitor was converted into various (acyloxy)methyl prodrugs in order to achieve permeability through cell membranes. These prodrugs inhibited insulin-stimulated autophosphorylation of the insulin receptor 95-kDa beta-subunit in intact CHO cells transfected with human insulin receptor. Inhibition of insulin-stimulated glucose oxidation in isolated rat adipocytes and 2-deoxyglucose uptake into CHO cells was observed with these prodrugs. Our data provide additional evidence for the involvement of the insulin receptor tyrosine kinase in the regulation of glucose uptake and metabolism. These results and additional data reported herein suggest that this class of prodrugs and inhibitors will be useful for modulating the activity of a variety of tyrosine kinases.

The insulin-mimetic effect of vanadate is not correlated with insulin receptor tyrosine kinase activity nor phosphorylation in mouse diaphragm in vivo.

The in vivo administration of sodium orthovanadate stimulated the incorporation of [14C]glucose into [14C] glycogen, in a dose- and time-dependent manner, in mouse diaphragm. Activation of diaphragm insulin receptor was measured by exogenous tyrosine kinase activity and an antibody that recognizes a conformational change in the receptor beta-subunit upon autophosphorylation. Neither method detected insulin receptor activation by in vivo vanadate administration, suggesting that vanadate's insulin-mimetic effect on mouse diaphragm glycogenesis occurs at a site distal to the insulin receptor.

Serum half-life and biological activity of mutants of human insulin-like growth factor I which do not bind to serum binding proteins.

We have characterized the biological properties of two mutants of human insulin-like growth factor I (IGF-I) which, as we have shown previously, have normal affinity for the type I IGF receptor, but drastically reduced affinity for the acid-stable components of human serum binding proteins. [Phe-1,Val1,Asn2,Gln3,His4,Ser8,His9,Glu12 ,Tyr15,Leu16]IGF I (B-chain mutant) and [Gln3,Ala4,Tyr15,Leu16]IGF I have 1000 and 500 times lower affinity than IGF-I for the native 150K binding protein in adult rat serum. Like IGF-I, these two peptides migrate as monomers during size exclusion chromatography on TSK 125. [125I]IGF-I, [125I]B-chain mutant, and [125I] [Gln3,Ala4,Tyr15,Leu16]IGF-I have in vivo serum half-lives of 100, 27.5, and 26.9 min, respectively, after iv injection. These data suggest that serum binding protein-bound peptide is cleared from the serum more slowly than free peptide. The tissue distributions of [125I]IGF-I and [125I]B-chain mutant are similar 10 min after dosing, with more than 80% of the tissue-sequestered intact radioactive peptides in the kidney. Despite decreased serum half-lives, the B-chain mutant and [Gln3,Ala4,Tyr15,Leu16]IGF-I are, respectively, 4 times and twice as active as IGF-I in stimulating the incorporation of [14C]glucose into glycogen in rat diaphragm in vivo. This effect of IGF-I is thought to be mediated by the type 1 IGF receptor in muscle, since the same doses of peptide that stimulated glycogen synthesis more than 30-fold did not stimulate the incorporation of [14C]glucose into total lipid in adipose tissue, an effect known to be mediated by the insulin receptor. These data support the hypothesis that serum- or tissue-derived binding proteins impair the ability of IGF-I to exert its effects through the type 1 IGF receptor in vivo.

A protein phosphotyrosine phosphatase distinct from alkaline phosphatase with activity against the insulin receptor.

Rat liver plasma membranes were found to have a relatively high ratio of acid to alkaline phosphatase activity when compared to rabbit liver and human placental membranes, respectively. The rat liver plasma membranes contained PPTl phosphatase activity against the soluble autophosphorylated insulin receptor beta-subunit. The PPT phosphatase activity of the membranes, using 32P-histone 2b as a substrate, was inhibited by 100 microM Zn+2, insensitive to 10 mM EDTA, and displayed maximal activity at neutral pH. Dephosphorylation of the insulin receptor beta-subunit by rat liver membranes was inhibited by Zn+2, and stimulated by EDTA. These results prove that the plasma membrane of a physiologically relevant insulin target tissue contains a PPT phosphatase, distinct from alkaline phosphatase, which catalyzes the dephosphorylation of the insulin receptor beta-subunit.