Diabetologia 9 by Springer-Verlag 1979

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1 Diabetlgia 16, (1979) Diabetlgia 9 by Springer-Verlag 1979 Interrelatinships f Glucse and Insulin Uptake by Muscle f Nrmal and Diabetic Man vidence f a Difference in Metablism f ndgenus and xgenus Insulin N. Kalant, T. Leibvici, I. Rhan, and S. zaki Lady Davis Institute fr Medical Research, Jewish General Hspital, Mntreal, Quebec, Canada Summary. A frearm perfusin technique was used t study glucse and insulin uptake by muscle. In nrmal subjects at glycaemic levels abve 13 mg/ 1 ml, glucse uptake was independent f glucse cncentratin; it was directly related t insulin cncentratin but nt t insulin uptake. In nn-bese maturity-nset diabetic subjects, glucse uptake was dependent n glucse cncentratin and insulin uptake, but nt n insulin cncentratin. In bth grups there was a strng crrelatin between insulin cncentratin and insulin uptake; diabetics had a nrmal insulin uptake in relatin t cncentratin. Fr a given change in insulin cncentratin the increase in glucse uptake was as great in diabetics as in cntrls, but the effect f insulin was mediated thrugh a mechanism invlving its uptake. Thus in the nn-bese maturity-nset diabetic, frearm muscle is nt insulin resistant. The apparent uptake f insulin measured by a radiimmunassay in relatin t its arterial cncentratin was lwer and mre variable fr heterlgus than fr endgenus insulin. With a receptr assay the venus insulin cncentratins were lwer than with the immunassay and differences in uptake between endgenus and exgenus insulin disappeared. It is cncluded that in muscle exgenus insulin was less severely degraded than endgenus insulin. Key wrds: Glucse cncentratin, glucse uptake, insulin cncentratin, insulin uptake, muscle, diabetes, radireceptr assay. A number f studies f the perfused frearm have demnstrated an insulin-stimulated increase in glucse uptake by muscle in nrmal subjects [1-5]. Hwever there is little infrmatin n the tissue uptake f insulin and the pssible relatinship f this t glucse utilizatin. Butterfield, Garratt and Whichelw [6] measured the tissue uptake f glucse and insulin fllwing intra-arterial injectin f 131Iinsulin; they fund that insulin uptake was directly related t plasma insulin cncentratin and that glucse uptake was linearly related t insulin uptake. Since this was an acute study in a nn-steady state, the "uptake" largely reflected mvement int the extravascular space rather than cellular uptake r utilizatin. Rasi et al. [7] subsequently studied the steady state respnse t a prlnged (1 hur) infusin f insulin; while again bserving a linear relatinship between plasma insulin cncentratin and insulin uptake, they did nt find a relatinship between cell glucse uptake and either insulin uptake r insulin cncentratin. These authrs als fund that intra-arterial injectin f glucse led t a fall in arterial insulin cncentratin. Because f these unexpected findings and because f unusually high and variable bld flw in the subjects studied, we have re-examined the steady-state relatinships between glucse and insulin uptake, in nrmal and diabetic subjects. Materials and Methds Studies were made n 9 nn-diabetic and 11 diabetic men. A detailed explanatin f the gals and prcedures f the experiment was given and cnsent btained. The nn-diabetics were healthy vlunteers, aged 4 t 73 (mean + s. e. m. = 54 _+ 5); they had n family histry f diabetes r ther metablic disease and tk n medicatins. Standard glucse tlerance tests perfrmed in 6 subjects were nrmal~ Bdy weight was 12_+3% f ideal (range 94-17). Preparatin f the subjects was as previusly described [8]; a diet cntaining 25-3 g f carbhydrate was taken fr 3 clays preceding the experimental prcedure, and n fd was taken fr h immediately preceding the study. Diabetic subjects X/79/16/365/$1.6

2 366 N. Kalant et al.: Glucse and Insulin Metablism in Muscle Table 1. Clinical infrmatin n diabetic subjects ~"6 ~ Hypglycaemic agent ther ~ ~ ~ ~ (daily dse) medicatin z U,v 1 g Amytriptyline Recent - Digxin. 9 Chlrprpamide (2 mg) - 2 Chlrprpamide (1 rag) - 1 Tlbutamide (1 g) Chlrprpamide (2 mg) Tlbutamide (.5 g) ~ ~ RRA I J I I I I 1:32 1:16 1:8 1:4 1:2 1 DILUTIN Fig. 1. Dilutin curves f mncmpnent insulin added t serum and assayed by immunassay (RIA) and receptr assay (RRA)., Standard curve. Dilutin curve f serum cntaining 3 ~tu/ml f endgenus insulin and 1 ~tu/ml f mncmpnent insulin were all f the maturity-nset type; they were in gd health apart frm the diabetes and were leading nrmally active lives. Pertinent infrmatin is shwn in Table 1. All were under treatment with diet, alne r with an ral hypgiycaemic agent; the diet was liberal (225-3 g) in carbhydrate and was nt altered prir t the experiment. Nne had previusly received insulin. The last dse f hypglycaemic drug was taken n the mrning prir t the prcedure. Bdy weights f all subjects were within 1% f ideal. The technique and validity f the frearm perfusin prcedure have been described previusly [8]. Briefly, bld is sampled frm a deep vein draining the frearm muscle and frm a vein n the drsum f the cntralateral hand which was warmed t increase bld flw s that the venus bld was "arterialized" [9]; bld flw was measured by strain-gauge plethysmgraphy [1]. Infusins f glucse and insulin were intrduced int an antecubital vein. Subjects were studied in the basal state and during steady-state perids f hyperglycaemia prduced by glucse infusins. Fllwing intrductin f the indwelling needles fr bld sampling, arterialized bld glucse was measured at 5 min intervals fr the remainder f the experiment. After a 3-6 rain rest perid t ensure a steady state, bld sampling and bld flw measurements were made. A new glycaemic steady state was then established by an intravenus pulse (.125 g/kg) and subsequent infusin f 5 g/1 ml glucse slutin. A glucse clamp prcedure was used t maintain cnstancy f the plasma glucse cncentratin [8]. When the plasma glucse had been cnstant fr 1 hur at the new level sampling and flw measurements were made. The rate f infusin was varied frm 1 subject t anther t achieve a range f plasma glucse cncentratin and thus f endgenus plasma insulin cncentratin. In 3 nn-diabetic and 2 diabetic subjects a secnd level f hyperglycaemia was established by a further pulse f glucse and a faster infusin f glucse slutin; stabilizatin at the new level fr 1 h was then fllwed by sampling and flw measurements as befre. ne subject in each grup was studied nly in the basal state while bservatins in the basal state were mitted fr 2 nn-diabetics and 3 diabetics. Tw f the nrmal and six f the diabetic subjects were given intravenus infusins f mncmpnent insulin (.75 mu/kg/min) with the glucse infusin t extend the range f plasma insulin cncentratins during the hyperglycaemic perids. During each perid (basal and hyperglycaemic) sampling f arterialized and deep venus bld and measurements f bld flw were made 3-4 times at 15 rain intervals. Bld flw was first measured times ver a 5-min interval; immediately fllwing this, while the wrist cuff was still inflated, bld was drawn simultaneusly frm the deep vein and the arterialized vein int heparinized tubes, fr subsequent measurement f glucse and insulin. Glucse uptake and insulin uptake were calculated fr each sampling interval; the mean cncentratin and uptake values fr each steady-state perid were used in subsequent statistical analyses. Plasma glucse was measured with an xygen electrde glucse analyzer (Beckman Instruments) at the bedside during the curse f the experiment and the result was subsequently verified by measurement in the labratry. Plasma insulin was measured by radiimmunassay (RIA) using a charcal separatin f unbund insulin [11] and in sme experiments by a radireceptr assay (RRA) as well [12]; the characteristics f the tw assays were (fr the RIA and RRA respectively): sensitivity, fr standard insulin,.35 ~tu/ml and.5 IxU/ml; sensitivity fr plasma insulin,.5 IxU/ ml and.7 ~tu/ml; precisin +.3 IxU and _+.4 ~tu at 5 p.u/m.1, ~tu and + 1.6/xU at 25 ~tu/ml, ~tu and --_ 1.8 ~tu/ml at 4 ~xu/ml; between-assay cefficient f variatin, 6.1% and 5.7%. Dilutin-recvery curves f mncmpnent insulin added t whle serum were indistinguishable frm the standard curve in bth assays (Fig. 1). The haematcrit was measured n all samples t exclude the pssibility f dilutin by the saline slutins used fr washing ut the venus catheters. The hypthesis tested during analysis f the data was that arterial glucse cncentratin determines arterial insulin cncentratin which in turn determines insulin uptake by muscle cells and that bth glucse cncentratin and insulin uptake cntrl glucse uptake. Results were analyzed by the methds f multiple regressin [13] and partial crrelatin [14].

3 N. Kalant et al.: Glucse and Insulin Metablism in Muscle 367 (nss!l ItU Urn.m/Ira) axij ~ms~id c.i c-i ~,. (nss!l I m t/u.au/fl~) ~I~ldn u!lnsui (~/n ~) mlnsu! Ieul.rv w.4 i~.- r162 I~ ~ tt3 r~'3~,4~ ~r-q.=. cl X (nss!l i m UU.UU/'&u) ~Imdn sdir" ~ N t"q "~ 9 r ~ r-- tt3 ",~ 1r I'~ sni~ I~.UalaV ~ (",1r162 t"q I?-- '~ r t',l. (nss.n Im I/m.tu/ItU) mlj ems~id (nsstl lul [/u.r.~/fl~ ) ~leldn u!insui z t-? (~/n~) u!insm. c[ X (nss!l I m t/u.~u/~tu) ~lmdn snl ;> e., e., ~ (w/~u) S311[~ l~.ul.w (nss!l lm l/m.m/im) mil. etus~id (nss!l I m T/u.nu/gl~) e~ "G. u.qnsui (~u/n~) u!lnsu! I 8 r162 8 cl X (nss!~ ItU l/lq.m/~m) ~Ieldn ~snl r el (tp/~u SDD[~ I~.Ul.tV,-"~(",ler163

4 368 N. Kalant et al.: Glucse and Insulin Metablism in Muscle A 1.25 n 125 % '~. 1 2 e- ~.75 - ~ U. (.: < _z 25 #:. z I L I ~ I ~t Z. [ 4 J I I 8 12 GLUCS CNCNTRATIN (mg/dl) INSULIN CNCNTRATIN (pu/ml) Fig. 2. A Relatinship f glucse uptake t arterial glucse cncentratin in nn-diabetic subjects. This demnstrates the crrelatin befre cntrlling fr the influence f insulin cncentratin n uptake. Regressin equatin: Y = X. Crrelatin cefficient, r =.51. B Relatinship f insulin uptake t arterial insulin cncentratin in nn-diabetic subjects. When radireceptr assay values are used fr the 2 data pints btained during insulin infusin, the regressin equatin is: Y = X and crrelatin cefficient, r =.86. When immtmassay results are used fr all data the regressin equatin is: Y = X, with a regressin cefficient, r =.78 Results All individual results are shwn in Table 2. Fr the 2 experiments the glucse cncentratins were cnstant within _+ 2 mg/dl f the mean during each perid; insulin cncentratins were within _+ 1 ~tu/ ml during the basal perids and within + 2 ~tu/ml during hyperglycaemia. Plasma flw was cnstant within _+ 5% f the mean. These results are in keeping with thse reprted previusly [8]. 1. Glucse Utilizatin The rate f glucse utilizatin by the whle bdy during a steady-state perid btained by glucse infusin was taken as the mean infusin rate fr that perid; these utilizatin rates were examined in relatin t glucse and insulin cncentratin. The regressin f the utilizatin rate n the tw variables was cnsiderably greater in the nn-diabetics: fr the nrmal and diabetic grups respectively, the regressin cefficients fr glucse utilizatin n glucse cncentratin were.11 and.1, and fr utilizatin n insulin cncentratin,.468 and.4. The verall difference between the tw grups was significant (P <.5). Since the average glucse cncentratins were similar in the tw grups (21 _+ 14 mg/dl and 29 _+ 2 mg/dl fr nn-diabetic and diabetic grups) an additinal estimate f insulin effectiveness was btained frm the rati, glucse utilizatin (mg/min/kg)/mean insulin cncentratin; this averaged.18_+.2 and.1_+.2 (mean _+ s. e.m.) fr the nn diabetic and diabetic subjects, respectively (P <.25). These analyses demnstrate that the diabetic subjects shwed insulin unrespnsiveness as well as decreased glucse tlerance. 2. Muscle Uptake f Glucse and Insulin In the nn-diabetics, there was a significant relatinship between cncentratin and uptake fr bth glucse and insulin (Fig. 2). The relatinship between insulin uptake and insulin cncentratin in the diabetics is seen in Figure 3 A. When immunassay values were used fr samples btained during insulin infusin, all estimates f uptake fell belw the regressin line f uptake n cncentratin f endgenus insulin and sme f the uptake values were clearly negative. Plasma insulin in the samples frm the insulin infusin experiments was then measured by the radireceptr assay, tgether with 3 randmly selected samples frm diabetic and nn-diabetic subjects wh did nt receive an insulin infusin. In the samples frm nn-insulin infused subjects and in arterialized venus samples frm the insulin infused

5 N. Kalant et al.: Glucse and Insulin Metablism in Muscle 369 %- 1- A B 9 " 8.~. 6.c 4 [] / / [] '~ 2 1,..,. _z z_ -2C ~ D D [] D I I I I 1 I INSULIN CNCNTRATIN (pu/rnl) Fig. 3A and B. Relatinship f insulin uptake t arterial cncentratin fr exgenus mad endgenus insulins. The slid line is the regressin line f uptake n cncentratin fr endgenus insulin in nn-diabetic subjects, taken frm Fig. 2B. ndgenus insulin in diabetic subjects. ~, xgenus insulin in diabetics,, xgenus insulin in nn-diabetics; tw f these are frm experiments included in Table 2 while the ther tw, frm yunger subjects, are shwn here nly as additinal infrmatin. A All measurements by radiimmunassay. Regressin equatin fr diabetics: Y= X. Crrelatin cefficient, r=.35 (P >.1). B xgenus insulin measured by radireceptr assay; brken line is regressin line fr all diabetic data. Regressin equatin, Y = X. Crrela~ tin cefficient, r =.88 (P <.1) subjects the receptr assay gave slightly higher values than the immunassay (3--4 ~tu/ml); hwever, in deep venus sera frm insulin infused subjects the receptr assay gave distinctly lwer values than the immunassay (mean + s.e.m, f difference = --12_+2 gu/ml). Fr bth diabetics and nn-diabetics estimates f insulin uptake based n the receptr assay results were all higher than thse based n immunassay and much clser t the regressin line fr the nn-diabetic data btained withut insulin infusin (Fig. 3B). 3. Crrelatin and Regressin Analyses Fr each f the tw grups, nn-diabetic and diabetic, an analysis was made f the data pled frm all experiments, using the receptr assay values f insulin cncentratin in thse experiments in which an insulin infusin had been given. Fr the nndiabetic subjects (Table 3), the dependence f glucse uptake n glucse cncentratin was nt statistically significant when the influence f insulin cncentratin was remved. Glucse uptake was strngly dependent n insulin cncentratin, but nt n insulin uptake as seen when the influence f insulin cncentratin was excluded.insulin uptake was highly dependent n glucse uptake and n insulin cncentratin, but the dependence n each f these tw factrs was greatly diminished by cntrlling the ther. Insulin cncentratin was psitively crrelated t glucse cncentratin, particularly when insulin uptake was cntrlled. Partial crrelatin analysis f data frm the diabetics (Table 4) shws tw majr differences frm that f the nn-diabetics: (a) the relatinship between glucse uptake and glucse cncentratin was significant even when insulin cncentratin was cntrlled; (b) glucse uptake was related t insulin uptake, but nt t insulin cncentratin. As with the nrmal subjects there was a strng negative crrelatin between insulin uptake and glucse cncentratin. Multiple regressin analyses f glucse uptake demnstrated that the influence f glucse cncentratin was small in bth grups f subjects (regressin cefficients f.1 and.2 fr nn-diabetics and diabetics respectively). In the nn-diabetics the nly significant determinant f glucse uptake was insulin cncentratin (regressin cefficient.6, P <.1), while in the diabetics nly insulin uptake was effective (regressin cefficient.13, P <.l). Similar analyses f insulin uptake shwed n significant differences between the tw grups: fr nrmal and diabetic subjects, respectively, the regressin cefficients f insulin uptake n insulin cncentratin

6 37 N. Kalant et al.: Glucse and Insulin Metablism in Muscle Table 3. Partial crrelatin cefficients fr nn-diabetic subjects Partial crrelatin excluding effect f Simple Glucse Glucse Insulin Insulin crrelatin cncentratin uptake cncentratin uptake Glucse uptake: glucse cncentratin Insulin cncentratin: glucse cncentratin d Insulin uptake: glucse cncentratin Glucse uptake: insulin cncentratin Glucse uptake: insulin uptake Insulin uptake: insulin cncentratin.51 a b.51 a b a --.5 a ~.91 c -- _.73 c.87 c.9 ~ r.89 ~ a P<.25; b P<.1; c P<.1 Based n 18 values btained in 9 experiments, using radireceptr assay values fr insulin measurement in thse experiments which incrprated an infusin f insulin d The administratin f exgenus insulin distrts any physilgical relatinship f these parameters; crrelatin cefficients are given fr cmpleteness nly. This artefact des nt affect the validity f ther relatinships. Table 4. Partial crrelatin cefficients fr diabetic subjects Partial crrelatin excluding effect f Parameters Simple Glucse Glucse Insulin Insulin crrelatin cncentratin uptake cncentratin uptake Glucse uptake: glucse cncentratin Insulin cncentratin: glucse cncentratin d Insulin uptake: glucse cncentratin Glucse uptake: insulin cncentratin Glucse uptake: insulin uptake Insulin uptake: insulin cncentratin a.65 b a b r.75 c c.87 c b c.87 c.77 ~ - _ a p <.25; u p <.1; c p <.1 Based n 2 values btained in 11 experiments, using radireceptr assay values fr insulin measurement in thse experiments which incrprated an infusin f insulin d See ftnte t Table 3 alne were.312 and.416, while fr insulin uptake n glucse uptake alne they were and Hwever, while n interactin between the insulin cncentratin and glucse uptake was evident in the diabetics, there was strng interactin in the nrmals (F fr interactin significant at.1). Frm the regressins f insulin uptake and glucse uptake n insulin cncentratin it can be calculated that unit change in insulin cncentratin prduced changes in glucse uptake f.6 mg/min/ 1 ml in the nrmal subjects and f.54 mg/min/ 1 ml in diabetics. Discussin Several authrs have previusly examined the interrelatinships f glucse and insulin in the perfused frearm, particularly with regard t the rle f glucse cncentratin and f insulin cncentratin as determinants f glucse utilizatin [ 1-7, 15]. Thugh based n the Fick principle fr calculatin f glucse uptake, a number f the earlier studies [2, 6, 14] did nt meet the criteria fr a steady state, which is essential fr valid applicatin f the Fick principle. In additin insulin was smetimes given as a single pulse r as an infusin in an amunt sufficient t raise the plasma cncentratin t unphysilgically high levels [1, 2]. Mre recently, Christensen and rskv [16] n the basis f experiments in which the glucse cncentratin was fairly cnstant, cncluded that there was n significant crrelatin between arterial glucse cncentratin and glucse uptake in the nndiabetic. ur results, based n the partial crrelatin after remving the effect f insulin cncentratin, are in agreement with this. Hwever, in diabetics we fund a small but significant dependence f glucse uptake n glucse cncentratin. It is pssible that in the nrmal subjects the effect f glucse cncentratin n uptake was already maximal at levels f glycaemia abve apprximately 13 mg/1 ml (i. e. the lwest levels btained with

7 N. Kalant et al.: Glucse and Insulin Metablism in Muscle 371 the glucse infusin) while in the diabetics it was nt yet maximal even at much higher glucse cncentratins. This is cmpatible with a defect in glucse transprt independent f insulin stimulatin. The rle f insulin in cntrlling glucse uptake is mre cntrversial. Zierler and Rabinwitz [17] and Rasi et al. [7] fund n effect n glucse uptake f an infusin which increased the arterial insulin cncentratin up t abut 65 ~U/ml, thugh a definite effect was nted with an infusin rate 1 times greater [17]. n the ther hand a linear relatinship between glucse uptake and insulin cncentratin in the physilgical range has been reprted by Fineberg and Merimee [5] and by Christensen and r [16]. In the present wrk, in the nn-diabetics there was a strng relatinship between glucse uptake and insulin cncentratin, independent f insulin uptake, but any crrelatin between glucse uptake and insulin uptake was greatly diminished by cntrlling insulin cncentratin. This suggests that glucse uptake and insulin uptake are independently related t insulin cncentratin and that the actin f insulin n glucse transprt is nt dependent n insulin metablism. The cnverse situatin btained in the diabetics: glucse uptake was nt related t insulin cncentratin, but was very strngly dependent n insulin uptake. It is assumed that uptake f insulin invlves binding t receptr, fllwed by degradatin; these tw prcesses are separate [18]. It is prbable that binding equilibrium is reached quickly under steady state cnditins in viv; cntinued remval f insulin (uptake) is therefre due t degradatin. ur results suggest that nrmally the actin f insulin n glucse transprt is related directly t the amunt f insulin bund at equilibrium and thus t the extracellular insulin cncentratin, while in the diabetics a new mechanism links degradatin t the glucse transprt prcess: there is n indicatin frm the present wrk f the nature f this link. Current cncepts f respnsiveness t insulin are based n quantitative relatinships between insulin cncentratin and glucse cncentratin r utilizatin. It is evident that in the diabetic subjects the increase in glucse uptake prduced by a unit increase in insulin cncentratin is as great as in the nn-diabetics, even thugh the effect is mediated by uptake f insulin. Thus the respnsiveness f muscle t insulin is nrmal despite a decrease in whle bdy respnsiveness. This suggests that ther tissues, particularly liver and adipse tissue, may be respnsible fr the insulin unrespnsiveness. It has recently been reprted that sulphnylurea treatment leads t increases in insulin respnsiveness and in the insulin binding capacity f islated liver plasma membrane [18]; it is therefre pssible that the nrmal insulin uptake and the nrmal stimulatin f glucse uptake in the muscle f the diabetics were the result f prir sulphnylurea therapy. This seems unlikely fr serveral reasns: (a) the subjects cntinued t shw decreased whle bdy respnsiveness t insulin, (b) the uptake f insulin in relatin t insulin cncentratin was the same fr the subjects treated with sulphnylureas as fr thse treated by diet alne (data pints fr the tw grups f subjects are randmly distributed in Fig. 3 B) and (c) an increase in insulin binding capacity t nrmal wuld nt explain the altered relatinship between insulin uptake and glucse uptake. The significance f the finding in bth nrmal and diabetic subjects f a high, negative crrelatin between glucse cncentratin and insulin uptake is nt clear. There may be an additinal factr assciated with the high plasma glucse cncentratin which inhibits insulin uptake; perhaps catechlamines may play such a rle. An unexpected finding in the subjects receiving insulin infusins was the lw and variable insulin uptake in cmparisn t that seen at cmparable plasma cncentratins f endgenus insulin. It appeared therefre that the metablism f exgenus insulin was different frm that f endgenus insulin. The results btained with the radireceptr assay fr insulin cnfirm this. Mild alteratin f the insulin mlecule leads t lss f ability t bind t receptr withut altering immunreactivity; further degradatin leads t lss f immunreactivity as well [19]. vidently, in viv all endgenus insulin is degraded t the pint where it is n lnger immunreactive, while part f the exgenus insulin is less severely degraded and retains immunreactivity. This wuld lead t an errneusly lw estimate f AV difference in the cncentratin f intact insulin and thus f insulin utilizatin. These results raise the warning that fr sme studies n insulin metablism, the fate f exgenus insulin may nt reliably mirrr the fate f endgenus insulin. Acknwledgements. This wrk was supprted by a grant frm the Medical Research Cuncil f Canada. We are indebted t Dr. W. atn fr advice and assistance with the statistical analyses. References 1. Andres, R., Baltzan, M. A., Cader, G., Zierler, K. L.: ffect f insulin n carbhydrate metablism and n ptassium in the frearm f man. J. Clin. Invest. 41, (1962) 2. Zampa, G.A., Altilia, F., Bracchetti, D., Geminiani, G.D., Brgatti,., difreddi, M. T.: Studies n peripheral glucse metablism using the experimental human frearm preparatin. Diabetlgia 3, (1967)

8 372 N. Kalant et al.: Glucse and Insulin Metablism in Muscle 3. Pzefsky, T., Felig, P., Tbin, J.D., Seldner, J.S., Cahill, G. F. Jr.: Amin acid balance acrss tissues f the frearm in pst-absrptive man. ffects f insulin at tw dse levels. J. Clin. Invest. 48, (1969) 4. Pzefsky, T., Santis, M.R., Seldner, J.S., Tancredi, R.C.: Insulin sensitivity f frearm tissues in prediabetic man. J. Clin. Invest. 52, (1973) 5. Fineberg, S.., Merimee, T. J.: ffects f cmparative perfusins f equimlar, single cmpnent insulin and prinsulin in the human frearm. Diabetes 22, (1973) 6. Butterfield, W.J.H., Garratt, C.J., Whichelw, M.J.: Peripheral hrmne actin: Studies n the clearance and effect f [131I]Idinsulin in the peripheral tissues f nrmal, acrmegalic and diabetic subjects. Clin. Sci. Ml. Med. 24, (1963) 7. Rasi,., Whichelw, M.J., Butterfield, W.J.H., Hicks, B.H.: Insulin fixatin and glucse uptake by frearm tissues in respnse t infusins f physilgical amunts f insulin in nn-diabetic subjects. Diabetlgia 8, (1972) 8. Kalant, N., Leibvici, T., Rhan, I., McNeill, K.: ffect f exercise n glucse and insulin utilizatin in the frearm. Metablism 27, (1978) 9. Jacksn, R. A., Peters, N., Advani, U., Perry, G., Rgers, J., Brugh, W. H., Pilkingtn, T. R..: Frearm glucse uptake during the ral glucse tlerance test in nrmal subjects. Diabetes 22, (1973) 1. Whitney, R. J.: The measurement f vlume changes in human limbs. J. Physil. (Lnd.) 121, 1-27 (1953) 11. Alban, J. D. M., kins, R.P., Maritz, G., Turner, R.C.: A sensitive precise radiimmunassay f serum insulin relying n charcal separatin f bund and free hrmne mieties. Acta ndcrinl. (Kbh.) 7, (1972) 12. zaki, S., Kalant, N.: A radireceptr assay fr serum insulin. J. Lab. Clin. Med. 9, (1977) 13. Chen, J.: Multiple regressin as a general data-analytic system. Psychl. Bull. 7, (1968) 14. Snedecr, G. W., Cchrane, W. G.: Statistical methds, ed. 6, pp Ames: Iwa State University Press Jacksn, R. A., Perry, G., Rgers, J., Advani, U., Pilkingtn, T. R..: Relatinship between the basal glucse cncentratin, glucse tlerance and frearm glucse uptake in maturity-nset diabetes. Diabetes 22, (1973) 16. Christensen, N.J., rskv, H.: The relatinship between endgenus serum insulin cncentratin and glucse uptake in the frearm muscles f nndiabetics. J. Clin. Invest. 47, (1968) 17. Zierler, K. L., Rabinwitz, D.: ffect f very small cncentratins f insulin n frearm metablism. Persistence f its actin n ptassium and free fatty acids withut its effect n glucse. J. Clin. Invest. 43, (1964) 18. Freychet, P., Kahn, R., Rth, J., Neville, D.M. Jr.: Insulin interactins with liver plasma membranes. J. Bil. Chem. 247, (1972) 19. Feingls, M.N., Lebvitz, H..: Sulfnylureas increase the number f insulin receptrs. Nature 276, (1978) Received: September 4, 1978, and in revised frm: February 12, 1979 N. Kalant, M. D. Lady Davis Institute Jewish General Hspital 3755 Cte Saint Catherine Rad Mntreal, Quebec H3T 12 Canada