Insulin‐induced translocation of glucose transporters in rat hindlimb muscles

Klip, Amira; Ramlal, Toolsie; Young, Douglas A.; Holloszy, John O.

doi:10.1016/0014-5793(87)80452-0

Cited by 330 publications

(245 citation statements)

References 22 publications

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“…The fold enrichments of both plasma membrane enzymes in the 25% sucrose fraction compare well with those reported originally by Klip and coworkers [14]. The specific activity of both enzymes was lower in membranes recovered on top of the 30 and 35% sucrose fractions and in the membrane pellet (Table 1) relative to those recovered from the 25% sucrose fraction indicating that the latter fraction was enriched with plasma membranes.…”

Section: Membrane Characterizationsupporting

confidence: 88%

“…The procedure for isolating rat muscle membrane fractions enriched with plasma membranes (PM) and intracellular membranes (IM) endowed with the insulin sensitive pool of GLUT4 and ~2-Na,K-ATPase subunit has been reported in detail [9,12,14]. Briefly, 10-15 g of skeletal muscle tissue was isolated from rat hindlimbs, homogenized, and sub-…”

Section: Subcellular Fractionation Of Rat Skeletal Musclementioning

confidence: 99%

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Sedimentation and immunological analyses of GLUT4 and α2‐Na,K‐ATPase subunit‐containing vesicles from rat skeletal muscle: evidence for segregation

Aledo

Hundal

1995

FEBS Letters

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In skeletal muscle insulin induces the translocation of both the GLUT4 glucose transporter and the 62 subunit of the Na,K-ATPase from an intraceHnlar membrane (154) compartment to the plasma membrane (PM). Fractionation studies of rat skeletal muscle using a discontinuous sucrose gradient have indicated that the insulin-induced loss of both proteins occurs from a fraction containing intracellular membranes (IM) of common density. This raises the possibility that both proteins may be colocalized in a single intracellular compartment or are present in separate membrane vesicles that are of similar buoyant density. In this study we report that membrane vesicles from the insulinresponsive IM fraction can in fact be separated on the basis of differences in their sedimentation velocities; immunoblot analyses of fractions collected from a sucrose velocity gradient revealed the presence of two separate peaks for GLUT4 and the a2 subunit of the Na,K-ATPase. One of these peaks representing a fast sedimenting population of vesicles (with a sedimentation coefficient of 2697 + 57 S) reacted against antibodies to the 62 subunit of the Na,K-ATPase, whereas, the second peak contained a population of much slower sedimenting vesicles (with a sedimentation coefficient of 209 -+ 4 S) were practically devoid of the a2-subunit. By contrast, the slow sedimenting vesicles were enriched by ~32-fold in GLUT4 relative to the starting IM fraction when the fractional protein content was taken into account. Immunoprecipitation of GLUT4-containing vesicles from the insulin-sensitive IM fraction revealed that no immunoreactivity towards either the a2 or the ~1 subunits of the Na,K-ATPase could be observed, signifying that the insulin-responsive subunits of the Na,K-ATPase and GLUT4 were present in different membrane vesicles and that it was unlikely, therefore, that the insulin-induced redistribution of these proteins to the PM occurs from a common intracellular pool.

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Section: Membrane Characterizationsupporting

confidence: 88%

Section: Subcellular Fractionation Of Rat Skeletal Musclementioning

confidence: 99%

Sedimentation and immunological analyses of GLUT4 and α2‐Na,K‐ATPase subunit‐containing vesicles from rat skeletal muscle: evidence for segregation

Aledo

Hundal

1995

FEBS Letters

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“…This method is a modification of that previously used to demonstrate insulin-mediated translocation of glucose transporters in rat muscle (Klip et al, 1987). All preparative steps were conducted at 4 1C in the presence of a protease inhibitor cocktail.…”

Section: Subcellular Fractionation By Sucrose Gradient Density Centrimentioning

confidence: 99%

Expression of the skeletal muscle dystrophin–dystroglycan complex and syntrophin-nitric oxide synthase complex is severely affected in the type 2 diabetic Goto–Kakizaki rat

Mulvey

Harno

Keenan

et al. 2005

European Journal of Cell Biology

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The inability of insulin to stimulate glucose metabolism in skeletal muscle fibres is a classic characteristic of type 2 diabetes. Using the non-obese Goto-Kakizaki rat as an established animal model of this type of diabetes, sucrose gradient centrifugation studies were performed and confirmed the abnormal subcellular location of the glucose transporter GLUT4. In addition, this analysis revealed an unexpected drastic reduction in the surface membrane marker b-dystroglycan, a dystrophin-associated glycoprotein. Based on this finding, a comprehensive immunoblotting survey was conducted which showed a dramatic decrease in the Dp427 isoform of dystrophin and the a/b-dystroglycan subcomplex, but not in laminin, sarcoglycans, dystrobrevin, and excitation-contraction-relaxation cycle elements. Thus, the backbone of the trans-sarcolemmal linkage between the extracellular matrix and the actin membrane cytoskeleton might be structurally impaired in diabetic fibres. Immunohistochemical studies revealed that the reduction in the dystrophin-dystroglycan complex does not induce obvious signs of muscle pathology, and is neither universal in all fibres, nor fibre-type specific. Most importantly, the expression of a-syntrophin and the syntrophinassociated neuronal isoform of nitric oxide synthase, nNOS, was demonstrated to be severely reduced in diabetic fibres. The loss of the dystrophin-dystroglycan complex and the syntrophin-nNOS complex in selected fibres suggests a weakening of the sarcolemma, abnormal signalling and probably a decreased cytoprotective mechanism in diabetes. Impaired anchoring of the cortical actin cytoskeleton via dystrophin might interfere with the proper recruitment of the glucose transporter to the surface membrane, following stimulation by insulin or muscle contraction. This may, at least partially, be responsible for the insulin resistance in diabetic skeletal muscles.

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“…Previous studies have shown that an appreciable amount of GLUT1 and GLUT4 protein is located in intracellular vesicles. However, only the glucose transporters which are inserted into the plasma membrane are functionally active [9]. Therefore, we determined the presence of the glucose transporter proteins GLUT1 and GLUT4 in the plasma membrane of rat skeletal muscle to assess whether in vivo glucose infusion induced the translocation of the transporters.…”

Section: Insulin Signalling Cascadementioning

confidence: 99%

Prolonged glucose infusion into conscious rats inhibits early steps in insulin signalling and induces translocation of GLUT4 and protein kinase C in skeletal muscle

et al. 2002

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Impairment of the glucose transport in the insulinsensitive tissues contributes to the pathogenesis of Type II (non-insulin-dependent) diabetes mellitus [1,2]. Skeletal muscle represents the most important tissue for the maintenance of a balanced postprandial glucose homeostasis; about 80 % of insulin-stimulated glucose uptake is accounted for by muscle [3±5]. The influx of glucose into muscle is mediated by the glucose transporter proteins GLUT1 and the insulinsensitive GLUT4 [6±11]. Insulin induces the translocation of GLUT4 to the sarcolemma and to special- Abstract Aims/hypothesis. Previous studies on diabetic patients have shown that hyperglycaemia increases glucose uptake in an apparently insulin-independent manner. However, the molecular mechanism has not been clarified. Methods. We studied rats receiving continuous glucose infusion to address this question. In this animal model, rats accommodate systemic glucose oversupply and rapidly develop insulin resistance. Results. Glucose infusion increased both plasma glucose and insulin concentrations to peak after one day. In spite of continuous glucose infusion normoglycaemia was reached after 5 days while insulin concentrations remained higher. Focusing our studies in day 2 (hyperglycaemia/hyperinsulinaemia) and day 5 (normoglycaemia/hyperinsulinaemia) we found, particularly in day 5, that the early steps of the insulin signalling cascade in skeletal muscle of glucose-infused rats were not more activated when compared to control animals as assessed by a comparable phosphorylation of the insulin receptor, IRS-1 and PKB and by an unaltered IRS-1-associated Ptd(Ins) 3' kinase activity. Continuous glucose infusion induced GLUT4 protein expression and translocation to the plasma membrane while neither expression nor translocation of GLUT1 was affected. Translocation of PKC-bI, -bII (> threefold) and -a, -q (to a lesser extent) to the plasma membrane was significantly induced after 2 days but not after 5 days of glucose infusion when normoglycaemia was reached. Conclusions/interpretation. Our data support the hypothesis that continuous glucose infusion induces translocation of GLUT4 while the early steps of the insulin signalling cascade were not increased. These effects could be mediated by activation of PKC. [Diabetologia (2002) 45: 356±368]

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Insulin‐induced translocation of glucose transporters in rat hindlimb muscles

Cited by 330 publications

References 22 publications

Sedimentation and immunological analyses of GLUT4 and α2‐Na,K‐ATPase subunit‐containing vesicles from rat skeletal muscle: evidence for segregation

Sedimentation and immunological analyses of GLUT4 and α2‐Na,K‐ATPase subunit‐containing vesicles from rat skeletal muscle: evidence for segregation

Expression of the skeletal muscle dystrophin–dystroglycan complex and syntrophin-nitric oxide synthase complex is severely affected in the type 2 diabetic Goto–Kakizaki rat

Prolonged glucose infusion into conscious rats inhibits early steps in insulin signalling and induces translocation of GLUT4 and protein kinase C in skeletal muscle

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