diamide and vitamin-k5

The effects of insulin and several insulin-mimetic agents on rat adipocyte D-glucose metabolism were studied in an effort to determine if any of the insulin-mimetic agents could be used to define the mechanism of insulin action. Antibodies against rat adipocyte plasma membranes have been characterized as having insulin-mimetic effects on glucose transport and these effects may be caused by divalent clustering of cell surface antigens. In contrast to insulin, antimembrane antibodies had little stimulatory effect on D-glucose conversion to lipids in isolated rat adipocytes, under conditions where both reagents stimulated D-glucose oxidation. Among other insulin-mimetic agents tested, the reagents hydrogen peroxide and concanavalin A most closely resembled insulin in their ability to increase both [14C]-CO2 and [14C]-lipid formation from [14C]D-glucose in rat adipocytes. Vitamin K5 and diamide had the unusual effect of inhibiting [1-14C]D-glucose conversion to [14C]-lipids at a concentration that gave maximal stimulation of glucose oxidation by rat adipocytes. Analysis of the lipid components into which glucose derivatives were incorporated revealed that insulin increased D-glucose incorporation into both nonesterified fatty acids and triglycerides and H2O2 and concanavalin A had similar effects. These findings argue against the possibility that insulin and the antimembrane antibodies or the insulin-mimetic agents other than H2O2 and concanavalin A share the same mechanism of action.

Topics: Adipose Tissue; Animals; Antibodies; Cell Membrane; Chromatography, Thin Layer; Concanavalin A; Diamide; Female; Glucose; Hydrogen Peroxide; Insulin; Lipids; Oxidation-Reduction; Rats; Rats, Inbred Strains; Spermine; Vitamin K; Vitamin K 3

These experiments examined the effects of N-ethylmaleimide on insulin- and oxidant-stimulated sugar transport in soleus muscle in terms of the Thiol-Redox model for insulin-stimulated adipocyte sugar transport (Czech, M.P. (1976) J. Cell. Physiol. 89, 661-668). Brief exposure (1 min) to N-ethylmaleimide (0.3-10 mM) inhibited the stimulatory effect of insulin (0.1 U/ml) on D-[U-14C]xylose uptake by rat soleus muscle. N-Ethylmaleimide also inhibited the stimulatory effects of H2O2 (5 mM), diamide (0.2 mM) and vitamin K-5 (0.05 mM). This effect of N-ethylmaleimide on insulin action was paralleled by the inhibition of 125I-labelled insulin binding by the muscle. N-ethylmaleimide lowered muscle ATP; however, its effects on sugar transport and 125I-labelled insulin binding could be dissociated from its effect on ATP. Exposing muscles to insulin prior to N-ethylmaleimide did not abolish the inhibitory effect of sulphydryl blockade on insulin-stimulated sugar transport, but did reduce the effect of the inhibitor by 20-30%. Conversely, when muscles were first allowed to bind 125I-labelled insulin and then exposed to the inhibitor, there was no effect of N-ethylmaleimide on pre-bound insulin. Exposure to diamide or vitamin K-5 before N-ethylmaleimide (1 mM) attenuated the inhibitory effect of sulphydryl blockade but no protective effect was observed with H2O2. None of the oxidants protected against the inhibitory effect of 3 mM N-ethylmaleimide. It is concluded that there are two N-ethylmaleimide-sensitive sites involved in the activation of muscle sugar transport at the post-receptor level. One of these would appear to be similar to the Thiol-Redox site described in the adipocyte; the other site appears to be an essential sulphydryl group whose function does not involve oxidation to a disulphide.

Topics: Animals; Azo Compounds; Biological Transport, Active; Diamide; Ethylmaleimide; Hydrogen Peroxide; Insulin; Muscles; Rats; Rats, Inbred Strains; Receptor, Insulin; Vitamin K; Vitamin K 3; Xylose

The experiments examined the stimulatory effects of H2O2, diamide and vitamin K-5 on the uptake of D-[U-14C]xylose by rat soleus muscle. All three oxidants stimulated sugar transport to the same extent, which was 30-40% of the maximal stimulatory effect of insulin (0.1 U/ml). Maximum stimulation was achieved with vitamin K-5 (0.01-0.1 mM), diamide (0.3 mM) and H2O2 (5-10 mM); at these concentrations the oxidants did not affect muscle ATP levels. Cytochalasin B (5 microM) abolished oxidant-stimulated xylose uptake. Catalase (20 U/ml) abolished the stimulatory effect of H2O2, but did not affect diamide- or vitamin K-5-stimulated transport. The ability of oxidants to stimulate sugar transport in anaerobic muscle could be demonstrated in short term (30 min) experiments, where muscle contained about 50% of its original ATP, but not after 120 min, when ATP levels were depleted. Oxidant-stimulated sugar transport was diminished and even abolished at supra-optimal oxidant concentrations; at these higher concentrations muscle ATP levels were lowered. All three oxidants inhibited the stimulatory effect of 2,4-dinitrophenol on sugar transport; this effect could be demonstrated using those oxidant concentrations which induced maximal stimulation of basal xylose uptake and which did not affect muscle ATP. It is concluded that: (1) H2O2, diamide and vitamin K-5 stimulate stereospecific sugar transport in soleus muscle by some mechanism other than lowering of ATP levels; (2) stimulation of sugar transport by oxidants is an ATP-dependent process; (3) some oxidant-sensitive sulphydryl group is critically involved in the process which activates muscle sugar transport.

Topics: Adenosine Triphosphate; Animals; Azo Compounds; Carrier Proteins; Diamide; Hydrogen Peroxide; Kinetics; Muscles; Rats; Rats, Inbred Strains; Sorbitol; Vitamin K; Vitamin K 3; Xylose