Biomedical subjects

J Shemer

Publications and source records attributed to J Shemer.

At least 145 records · Page 8Linked to original sources

PubMed1988

Insulin-sensitive tyrosine kinase is increased in livers of adult obese Zucker rats: correction with prolonged fasting.

Adult obese Zucker rats (fa,fa) are hyperinsulinemic and insulin resistant. Specific insulin binding to crude membranes prepared from livers was 2.8% (per mg protein) in fatty animals compared with 7.9% in homozygous lean (Fa,Fa) and 9.0% in heterozygous lean (Fa,fa) animals. Insulin binding increased in liver membranes from fatty animals after a 72-h fast to 6.4%. The reduced insulin binding in livers from fatty rats was associated with elevated insulin-sensitive tyrosine kinase activity, which fell towards control values after the fast. The elevated tyrosine kinase activity was associated with an increased maximum velocity (Vmax) without a change in Michaelis-Menten constant (Km) for its substrates, ATP and poly(Glu,Tyr)4:1. These findings suggest that, in adult fatty rats, insulin-sensitive tyrosine kinase has increased intrinsic activity. Further, the effect of the prolonged fast on both insulin binding and kinase activity, suggest that in this model environmental factors, and not necessarily a genetic abnormality, may regulate liver insulin receptors and their kinase. Whether the inverse relationship of the kinase and insulin receptor number is the result of a compensatory mechanism remains to be elucidated.

Animals↗

PubMed1987

Insulin and insulin-like growth factor-I stimulate a common endogenous phosphoprotein substrate (pp185) in intact neuroblastoma cells.

Mouse neuroblastoma N18 cells contain specific high affinity insulin and insulin-like growth factor-I (IGF-I) receptors. Insulin and IGF-I induce phosphorylation, in intact cells, of their respective receptor beta subunits. The insulin receptor beta subunit is represented by a 95-kDa phosphoprotein that is recognized by a specific antiserum (B10). The IGF-I receptor beta subunit is represented by two phosphoproteins of molecular mass 95 and 105 kDa. The hormone-induced phosphorylation was rapid and dose-dependent occurring on both phosphoserine and phosphotyrosine residues. In addition, both insulin and IGF-I induced phosphorylation of an endogenous protein of molecular mass 185 kDa (pp185). The rapidity and dose dependency of the phosphorylation of pp185 suggested that it may represent a common endogenous substrate for the insulin and IGF-I receptors in these neural-derived cells. Phosphorylation was primarily on phosphoserine and phosphotyrosine residues. pp185 did not absorb to wheat germ agglutinin-agarose and was not stimulated by either epidermal growth factor or platelet-derived growth factor. The finding of pp185 in these neural-related cells as well as in non-neural tissues suggests that it may represent a ubiquitous endogenous substrate for both the insulin and IGF-I receptor kinases.

Animals↗

PubMed1987

Insulin receptors and insulin action in dissociated brain cells.

The present study was conducted to characterize insulin receptors and insulin action in rat brain cells. Binding of [125I]insulin to cells obtained by mechanically dissociating rat brains was 86% specific, time-dependent and reached equilibrium within 90 min. The t1/2 of association was 14 min and t1/2 of dissociation was 8 min. Scatchard analysis demonstrated the typical curvilinear plot providing high affinity (0.03 nM) and low affinity (6.6 nM) binding sites. The total number of binding sites were 0.15 pmol/mg protein. Crosslinking of [125I]insulin to its receptors on dissociated brain cells followed by SDS-PAGE and autoradiography showed that the alpha-subunit of the receptor had a molecular weight of 122,000. This was in contrast with a molecular weight of 134,000 for the liver alpha-subunit. Incubation of dissociated brain cells with insulin resulted in a concentration-dependent inhibition of total [3H]norepinephrine (NE) uptake. This inhibitory effect of insulin on [3H]NE uptake was sodium ion-dependent suggesting that 80-90% of the sodium ion-dependent uptake was insulin-sensitive. Incubation of lectin-purified insulin receptors with insulin resulted in a time- and concentration-dependent stimulation of phosphorylation of the tyrosine residue of an exogenous substrate poly (Glu, Tyr) (4:1). In addition, insulin also stimulated the autophosphorylation of the beta-subunit of the insulin receptors. These observations corroborate our contention that insulin exerts neuromodulatory effects mediated by the specific insulin receptors in the brain.

Animals↗

PubMed1987

Insulin-like growth factor I receptors in neuronal and glial cells. Characterization and biological effects in primary culture.

Primary cultures of neuronal and glial cells from 1-day-old neonatal rats contain high affinity receptors for insulin-like growth factor I (IGF-I). The IC50 for displacement of 125I-IGF-I binding by unlabeled IGF-I was 3 nM for neuronal cells and 4 nM for glial cells. Unlabeled insulin was 20-50 times less potent. Apparent molecular mass of the alpha subunits of the IGF-I receptor was 125 kDa in neuronal and 135 kDa in glial cells. IGF-I induced autophosphorylation of the IGF-I receptor beta subunit in lectin-purified membrane preparations in a dose-dependent manner. The major phosphoamino acid of the beta subunit in both cell types was tyrosine in the IGF-I-stimulated state and serine in the basal state. Apparent molecular mass of the beta subunits of the IGF-I receptors was 91 kDa for neuronal and 95 kDa for glial cells. Tyrosine kinase activity of the IGF-I receptors was demonstrated by IGF-I-induced phosphorylation of the exogenous substrate poly(Glu, Tyr) 4:1 in both cell types. IGF-I had no effect on 2-deoxyglucose uptake in neuronal cells. In contrast, in glial cells, IGF-I stimulated 2-deoxyglucose uptake at very high doses, presumably acting via the insulin receptor. The effect of IGF-I as a neurotrophic growth factor in both neuronal and glial cells was demonstrated by its stimulation of [3H]thymidine incorporation. These findings suggest the IGF-I is an important growth factor in nervous tissue-derived cells.

Animals↗

PubMed1987

Characterization of an endogenous substrate related to insulin and insulin-like growth factor-I receptors in lizard brain.

Lizard insulin receptors are evolutionarily highly conserved. Wheat germ agglutinin-purified brain membranes demonstrate the presence of an endogenous substrate (pp 105) for both the insulin and insulin-like growth factor-I receptors. Both insulin and I-insulin-like growth factor-I stimulate the phosphorylation of this endogenous substrate in a dose-dependent manner. Following insulin-stimulated autophosphorylation of the beta subunit, there is a lag period of about 5 min prior to observable phosphorylation of the endogenous substrate. Phosphoamino acid analysis of both the beta subunit as well as pp 105 reveal primarily phosphotyrosine in both the basal as well as the stimulated state.

Amino Acids↗

PubMed1987

Frog brain and liver show evolutionary conservation of tissue-specific differences among insulin receptors.

The insulin receptors of frog brain and liver show features typical of other insulin receptors with regard to affinity and specificity of binding to insulins and proinsulin, solubility in Triton X-100, binding to and elution from wheat germ agglutinin, and insulin-sensitive tyrosine kinase activity. Likewise, the brain and liver receptors differ from one another in electrophoretic mobility and susceptibility to treatment with neuraminidase, analogous to brain and liver receptors of reptiles, birds, and mammals; while the functional implications of these differences are unknown, their evolutionary conservation for 400-500 million years suggests the possibility that they might have importance.

Animals↗

PubMed1987

The interaction of brain insulin receptors with wheat germ agglutinin.

Brain insulin receptors adsorb to and are recoverable from wheat germ agglutinin-agarose (WGA) columns. Similar results are obtained using dissuccinimidyl suberate (DSS)-crosslinked receptors or photo-affinity labeled receptors. WGA can be used for partial purification of brain insulin receptors provided the appropriate WGA preparation is chosen and the optimal ratio of receptor protein to lectin is achieved.

Animals↗

PubMed1987

Structural and functional studies on insulin receptors from alligator brain and liver.

Insulin receptors are present in membranes prepared from Alligator mississippiensis brain and liver. The apparent molecular weight (MW) of the alpha subunits are 132 kDa and 118 kDa in liver and brain respectively. Apparent MW of the beta subunit is 92 kDa in both brain and liver receptors. Despite the structural differences between brain and liver alpha subunits, brain insulin receptors demonstrate the normal coupling between alpha and beta subunits, i.e. following binding of insulin to the alpha subunit the beta subunit undergoes autophophorylation and stimulates tyrosine specific phosphorylation of exogenously added substrates. These findings suggest that functional insulin receptors are evolutionarily well conserved.

Alligators and Crocodiles↗

PubMed1986

Insulin receptors in lizard brain and liver: structural and functional studies of alpha and beta subunits demonstrate evolutionary conservation.

Specific insulin receptors are present in the liver and brain of the lizard Anolis carolinesis. In this study, the specific binding of 125I-insulin to the receptors showed time, temperature and pH dependency. Specific binding to crude membranes prepared from brain was 1-2% of the total radioactivity added compared to 4-5% in the crude membranes prepared from liver. Solubilization and wheat germ agglutinin purification of the membranes resulted in an increase in the specific binding (per mg of protein) between 6 and 32 times for liver membranes and 13-186 for brain membranes. Binding inhibition of tracer insulin by unlabeled porcine insulin was characteristic for insulin receptors with 50% inhibition for liver crude membranes at 60 ng/ml of porcine insulin and 0.7 ng/ml for purified brain insulin receptors. Chicken insulin was 2- to 3-fold more potent and proinsulin about 100 times less potent than porcine insulin. The alpha-subunits of liver and brain had apparent molecular weights on sodium dodecyl sulfate polyacrylamide gel electrophoresis of 135 kDa and 120 kDa respectively. Apparent molecular weights of beta subunits were 92 kDa for both tissues. Insulin stimulated phosphorylation of the beta subunit of both brain and liver receptors. Both tissues demonstrated tyrosine-specific phosphorylation, which was stimulated by insulin, of exogenously added artificial substrates. In addition, purified brain insulin receptor preparations contained an endogenous protein with apparent molecular weight of 105 kDa, whose phosphorylation was stimulated by insulin (10(-7) mol/l). This phosphoprotein was not immunoprecipitated by anti-insulin receptor antibodies. These studies suggest that the structural differences between brain and liver receptors previously demonstrated in the rat are also present in the lizard, which is about 300,000,000 years older than the mammalian species. Thus, there is strong evolutionary conservation of the brain insulin receptor.

Animals↗

PubMed1986