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PubMed1983

Vanadate binding to the gastric H,K-ATPase and inhibition of the enzyme's catalytic and transport activities.

Vanadate inhibition of the catalytic and transport activities of the gastric magnesium-dependent, hydrogen ion transporting, and potassium-stimulated adenosinetriphosphatase (EC 3.6.1.3) (H,K-ATPase) has been studied. The principal experiment observations are the following: (1) Inhibition of adenosine 5'-triphosphate (ATP) hydrolysis is biphasic. Vanadate binding with a stoichiometry of 1.5 nmol mg-1 approximately halves K+-stimulated ATPase activity at physiological temperature. The remaining activity is inhibited by binding an additional 1.5 nmol mg-1 vanadate with lower apparent ions bind specifically to gastric vesicles with two affinities. Vanadate binding in the presence of nucleotide is compatible with competition for the kinetically defined high-affinity and low-affinity ATP sites. (3) Vanadate inhibits phosphoenzyme formation and the K+-stimulated p-nitrophenyl phosphatase activity of the enzyme monophasically. A maximum of 1.5 nmol mg-1 acid-stable phosphoenzyme is formed. The half-time for vanadate dissociation from the site that inhibits p-nitrophenyl phosphate hydrolysis is 5 min (4) At most, 3 nmol mg-1 vanadate is required to inhibit proton transport. The simplest interpretation of the data is that vanadate inhibits the H,K-ATPase by binding competitively with ATP at two catalytic sites. Different catalytic mechanisms at the high-affinity and low-affinity sites are suggested by the different stoichiometries found for vanadate binding and phosphoenzyme formation.

Adenosine Triphosphatases↗
PubMed1998

The mitogen-activated protein kinase pathway contributes to vanadate toxicity in vascular smooth muscle cells.

Vanadate has been considered in the treatment of diabetes because of its insulin-like effects. However, it has severe toxic effects in both animal and man. In cultured cells, vanadate can either cause death or be growth stimulatory, depending on the cell type and growth conditions. Here, we report that in baboon aortic smooth muscle cells (SMCs), vanadate induced p42/p44 mitogen-activated protein kinase (MAPK) activity. This effect was abolished in the presence of the specific MAPK kinase (MAPKK) inhibitor PD098059. Although activation of p42/p44MAPK/MAPKK is generally thought to be necessary for proliferation, in SMCs, vanadate did not promote DNA synthesis and inhibited thymidine incorporation stimulated by platelet-derived growth factor (PDGF)-BB in a dose dependent fashion (IC50: 30 microM). Prolonged exposure to vanadate exerted cytotoxic effects. Cells retracted, rounded up and detached from the substratum. These vanadate-induced morphological changes were blocked in the presence of PD098059. The addition of PDGF-BB further activated p42/p44MAPK/MAPKK in the presence of vanadate and substantially increased vanadate toxicity. We conclude from these observations that activation of the p42/p44MAPK/MAPKK signalling module contributes to the cytotoxic effects induced by vanadate.

Animals↗
PubMed1994

Vanadate has no effect on insulin-extraction by the rat liver.

The mechanism(s) responsible for the insulin-like effects of vanadate are still unclear, although several possible explanations have been raised. However, the possibility that vanadate induces inhibition of insulin degradation in the liver was not examined yet. Therefore, in the present study we examined the effect of vanadate on the extraction of insulin by the perfused rat liver using an open, non-recycling system. Baseline insulin extraction (44 +/- 2% and 37 +/- 3%) was not affected by the administration of 1 and 5 microM vanadate (decreased to 42 +/- 2% and 36 +/- 1, respectively, p = NS). Vanadate had no adverse effects on liver viability, and the bile flow remained stable during and after vanadate administration (0.87 +/- 0.08 microliter/min/g liver prior to Vs. 0.084 +/- 0.11 microliter/min/g liver following vanadate administration). This study shows that vanadate does not inhibit insulin extraction by the perfused liver, and that does of vanadate that effectively inhibit hepatic glucose production possess no adverse effects on liver viability.

Animals↗
PubMed1999

Vanadate induction of NF-kappaB involves IkappaB kinase beta and SAPK/ERK kinase 1 in macrophages.

The present studies investigated the signaling pathways of vanadate, a vanadium ion with +5 oxidation state, to activate NF-kappaB transcription factor, a pivotal regulator of inflammatory responses. Treatment of macrophages with vanadate results in the activation of both NF-kappaB and c-Jun N-terminal kinase (JNK). The activity of a recently identified cellular kinase, IkappaB kinase-beta (IKKbeta), was significantly elevated concomitant with the increased degradation of IkappaBalpha and enhanced NF-kappaB activity in cells exposed to vanadate. To determine whether the IKK pathway and JNK pathway are interconnected or bifurcate upon vanadate stimulation, cells were transfected with either a kinase inactive form of IKKbeta or a kinase inactive form of SAPK/ERK kinase 1 (SEK1). Inactive IKKbeta was able to block vanadate-induced degradation of IkappaBalpha, yet it was unable to influence the activation of JNK by vanadate. Conversely, blockage of JNK activation by transfection of a kinase-inactive form of SEK1 resulted in partially inhibition of vanadate-induced IkappaBalpha degradation. Both vanadate-induced degradation of IkappaBalpha and activation of JNK were potently inhibited by pretreatment of cells with N-acetylcysteine or dimercaprol. These results demonstrate that early activation of stress kinases or change of cellular redox states plays a key role in vanadate-induced activation of NF-kappaB and JNK.

Animals↗
PubMed1999

Effect of vanadate on force and myosin light chain phosphorylation in skinned aortic smooth muscle.

The effects of vanadate were examined on Ca2+-activated force and phosphorylation of 20-kDa myosin light chain in membrane-permeabilized rabbit aortic smooth muscle strips. Addition of vanadate during maximum contraction reduced the force in a dose-dependent manner, and inhibited it almost completely at 1 mM. Two-dimensional polyacrylamide gel electrophoretic analyses revealed that vanadate also reduced the phosphorylation of 20- kDa myosin light chain in a dose-dependent manner from approximately 50% in the absence of vanadate to approximately 20% in the presence of 1 mM vanadate. The effects of 1 mM vanadate on purified myosin light chain kinase and phosphatase were then examined using purified myosin as substrate, and it was found that vanadate neither inhibited myosin light chain kinase nor activated myosin light chain phosphatase. These results indicate that the reduction in the 20-kDa myosin light chain phosphorylation level by vanadate may be effected through its inhibition of the force generation in skinned smooth muscle strip, as evidenced by the finding that vanadate eliminated the enhancement of myosin light chain kinase activity brought about by the interaction between purified myosin and actin.

Actins↗
PubMed1992

Inhibition of phosphoenzyme formation from phosphate and sarcoplasmic reticulum Ca(2+)-ATPase by vanadate binding to high- or low-affinity site on the enzyme.

In the preceding paper, we suggested that 1 mol Ca(2+)-ATPase of sarcoplasmic reticulum (SR) contains 0.5 ml of high-affinity vanadate binding sites as well as 0.5 ml of low-affinity vanadate binding sites [Yamasaki, K. & Yamamoto, T. (1991) J. Biochem. 110, 915-921]. In the present study, we examined the effects of vanadate binding to the high- and low-affinity sites upon phosphorylation of the enzyme by inorganic phosphate (Pi). When vanadate was added to the reaction medium in which the Ca(2+)-ATPase had been phosphorylated by Pi in the absence of Ca2+, the steady-state level of phosphoenzyme (E2P) decreased due to inhibition of its formation. The decrease of E2P after addition of vanadate exhibited biphasic kinetics consisting of an initial fast decay process followed by a slower first-order decay process. The size of the fast E2P decay, which was estimated by extrapolating the slow phase decay to time 0, varied depending on the vanadate concentration with a dissociation constant of 17 microM, and reached maximum at 50 microM vanadate. The maximum value of the fast E2P decay was almost equal to the initial E2P level. The initial fast decay of E2P was competitively prevented by Pi with a dissociation constant of 7.4 mM, which was very close to Km for the E2P formation under similar conditions. These observations suggested that vanadate inhibits E2P formation by competition with Pi at a phosphorylation site on the Ca(2+)-ATPase. The slow first-order decay of E2P corresponded well to the vanadate binding to the high-affinity site of the Ca(2+)-ATPase.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Actions of vanadate on vascular tension and sodium pump activity in cat isolated cerebral and femoral arteries.

1. The mechanisms involved in the responses induced by sodium vanadate (Va3 VO4) on cat cerebral and femoral arteries were studied. The possibility that these responses were due to Na+, K+-ATPase inhibition was investigated by measuring the effect of vanadate on [3H]-ouabain binding to arterial membrane fractions, K+-induced vasodilatation and ouabain-sensitive 86Rb+ uptake. 2. The vanadium compounds (Na3VO4, VOSO4, VCl3 and O5V3) induced similar, concentration-dependent contractions in each kind of artery, the cerebral vessels being the most sensitive to these compounds. 3. Exposure of the arteries to a low-Na+ (25 mM) solution suppressed the contraction caused by vanadate in femoral but not in cerebral arteries. 4. Vanadate-induced contractions were reduced in Ca2+-free medium but remained unaffected by 3 x 10(-6) M phentolamine, reserpine pretreatment or 3 x 10(-6) M verapamil in both kinds of artery. 5. The addition of 7.5 mM K+ to the arteries immersed in a K+-free solution induced vasodilatation, which was not modified by 10(-3) M vanadate. 6. The consecutive administration of ouabain (10(-4) M) and vanadate (10(-3) M) (or vice versa), or the simultaneous administration of both agents (10(-8) to 10(-3) M) appeared to produce an additive contraction in both types of artery. 7. Vanadate (10(-7) to 10(-3) M) did not displace the [3H]-ouabain binding to arterial membrane fractions of these arteries, whereas 10(-4) M ouabain did. 8. In both kinds of artery, total 86Rb+ uptake was reduced by ouabain (10(-8) to 10(-3) M), in a concentration-dependent manner, whereas it was not modified by vanadate (10(-8)-10(-3) M). 9. These results suggest that vanadate induces contraction in both types of artery by a mechanism unrelated to Na+, K+-ATPase inhibition. Such a mechanism is likely to be related to inhibition of the Ca2-ATPase of the cell membrane and/or the sarcoplasmic reticulum.

Animals↗
PubMed1993

Effects of vanadate on responses of guinea-pig isolated trachea to spasmogens.

The effects of vanadate on the contractility of the guinea-pig isolated trachea was examined. Vanadate (0.1 mM) produced a sustained contraction that was abolished in Ca(2+)-free EGTA (0.1 mM)-containing physiological salt solution but was resistant to verapamil (1 microM). Vanadate (0.1 mM) depressed tracheal responses to CaCl2 (in Ca(2+)-free depolarizing solution), KCl, acetylcholine, histamine and 5-hydroxytryptamine. For vanadate (10 microM), the inhibition of spasmogenic responses only reached statistical significance for histamine and 5-hydroxytryptamine. Caffeine (1 mM)-induced spasm (trachea at 20 degrees C in the presence of indomethacin (2.8 microM)) was not affected by vanadate (10 microM-0.1 mM). Vanadate (0.1 mM) slightly depressed the responses to KCl (50 mM), acetylcholine (1 mM), histamine (1 mM) or 5-hydroxytryptamine (0.1 mM) observed in Ca(2+)-free EGTA (0.1 mM)-containing physiological salt solution. Vanadate (0.5 mM) depressed Ca2+ (20 microM)-induced contraction of trachea which had been chemically skinned of its plasmalemmal membranes. The mechanism of the inhibitory effect of vanadate on tracheal responses to a variety of spasmogens remains obscure, but, under in-vitro conditions, vanadate clearly does not induce hyper-reactivity of airway smooth muscle to spasmogens.

Acetylcholine↗
PubMed1986

Effects of sodium vanadate on various types of vascular smooth muscles.

Effects of sodium vanadate on various vascular smooth muscles of guinea pigs, rabbits, and Wistar Kyoto rats (WKY) were studied. Sodium vanadate of concentrations higher than 10(-5) M induced contractions in the aortae of all animals. The contractile effects varied among vascular smooth muscles, and mesenteric arteries showed no or only weak contractile response to the drug, while aortae showed higher contractile responses. In the portal veins, potentiation of spontaneous contractions was observed by the application of sodium vanadate. These responses were not blocked by treatments with adrenergic blocking agents or indomethacin, indicating the direct action of the drug on vascular smooth muscles. Treatment with 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) blocked completely the contractile effects of sodium vanadate, whereas it showed no effect on K-contractures. Partial depolarization of the membrane by elevations of K concentration potentiated sodium vanadate-induced contractions and minimized the variations of responses among preparations. In K-depolarized preparations, sodium vanadate often induced relaxation of preparations. The contractile effects of sodium vanadate were not blocked by treatment with ouabain, though ouabain also showed contractile actions in a number of preparations. It was suggested that vanadate acts directly on vascular smooth muscles and causes contractions without relation to the inhibition of Na,K-ATPase. It may cause contractions inhibiting Ca-ATPase of sarcoplasmic reticulum and/or of cell membrane, and cause relaxation by inhibiting ATPase of contractile proteins. The variations of the responses may be explained by differences of membrane permeability to vanadate.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1997

Vanadate, but not insulin, inhibits insulin receptor gene expression in rat hepatoma cells.

Insulin and vanadate treatments have recently been shown to reverse the overexpression of the hepatic insulin receptor (IR) gene in streptozotocin-induced diabetic rats. To better understand the mechanisms underlying these effects, the abilities of insulin and vanadate to affect IR gene expression have been comparatively examined in Fao hepatoma cells, an insulin-responsive cell line. Exposure of Fao cells to insulin (1 microM) or vanadate (500 microM) for 24 h led to a 2-fold decrease in IR number in total cellular membranes. Insulin treatment did not affect IR messenger RNA (mRNA) level regardless of time of exposure and concentration. In contrast, vanadate treatment caused a time- and dose-dependent decrease in IR mRNA level, which was maximal (4-fold change) after a 24-h exposure to 500 microM vanadate and was fully reversible. Insulin treatment increased from 28 to 39% the relative expression of isotype A IR mRNA, but vanadate treatment did not significantly affect this parameter. Vanadate treatment did not modify mRNA half-life (3.5 h) in 5, 6 dichlorobenzimidazole riboside-treated cells but decreased by 4-fold the transcriptional activity of the IR gene. These data show for the first time that, although both insulin and vanadate decrease total cellular IR number in Fao cells, only vanadate decreases IR mRNA level. It does so by inhibiting transcription of the IR gene, suggesting an action on the gene promoter which could be mediated by changes in the level of expression and/or of phosphorylation of trans-acting factors.

Animals↗
PubMed1998

Vanadate elevates lipogenicity of starved rat adipose tissue: mechanism of action.

We have established an experimental system in rats in which the lipogenic capacity of adipose tissue was decreased in vivo by prolonged fasting, and restored in vitro by glucose together with insulin or vanadate. Incubation of fasted adipose explants for 5 h at 37 C with 2 mM glucose alone did not elevate lipogenic capacity. However, glucose with insulin (17 nM) or vanadate (100 microM), led, respectively, to 2.2- and 8- to 10-fold elevation. Actinomycin D (50 microM) completely blocked this increase, while low concentrations (ED50 = 4.0 +/- 0.4 microM) of vanadate potentiated it. Neither insulin nor vanadate elevated fasted adipose explants' lipogenic capacity in the absence of glucose, or in the presence of the nonmetabolizable glucose analog 3-O-methylglucose. Upon replacing glucose with 2-deoxyglucose (1 mM), a glucose analog that undergoes phosphorylation to 2-deoxyglucose-6-phosphate with no further metabolism, vanadate was nearly as potent as with glucose in elevating lipogenic capacity. Vanadate was superior to insulin in increasing glucose-6-phosphate level in fasted-adipose explants. Adipose glucose-6-phosphatase activity was inhibited by vanadate (IC50 = 7.0 +/- 0.4 microM). We have concluded that glucose-6-phosphate is the key metabolite of glucose involved in the transcriptionally regulated elevation of lipogenic capacity of fasted adipose explants. Vanadate has a more profound effect than insulin, as it elevates glucose-6-phosphate to higher levels and the subsequent increase in lipogenic capacity is four to five times greater than that induced by insulin. The mechanism involved is the combined action of vanadate in enhancing glucose entry and in inhibiting dephosphorylation of endogenously formed glucose-6-phosphate. The latter effect is not exerted by insulin.

3-O-Methylglucose↗
PubMed1999

Independent signal-transduction pathways for vanadate and for insulin in the activation of glycogen synthase and glycogenesis in rat adipocytes.

The activating effect of vanadate on glycogenesis and on glycogen synthase (uridine diphosphate-glucose-glycogen glucosyl transferase) activity was studied in rat adipocytes and compared with that of insulin. Using several approaches and specific blockers, we found that vanadate and insulin resemble each other, in the activation of glycogen synthase, in several aspects: both require nonarrested protein phosphatase 1 activity; they are equally suppressed by conditions that elevate cAMP-levels; and both depend on the activation of phosphatidylinositol-3 kinase. The basic differences between them are as follows: 1) vanadate promotes glycogenesis through the activation of a cytosolic protein tyrosine kinase, in an insulin-receptor-independent manner; 2) vanadate elevates glucose-6-phosphate (G-6-P) to a higher level than insulin; 3) vanadate-activated glycogenesis is accompanied by an increase in the cellular content of immunoreactive glycogen synthase, an effect less noticeable with insulin; 4) adipose glucose-6-phosphatase is inhibited by vanadate (dose for 50% inhibition, IC50 = 7 +/- 0.7 microM) but not by insulin. We have concluded that insulin and vanadate activate glycogenesis through a phosphatidylinositol-3 kinase and dephosphorylation-dependent mechanism. Vanadate, however, uses a receptor-independent pathway and is superior to insulin in elevating the level of G-6-P, a key metabolite for activating glycogen synthase. This is attributed to the combined effect of vanadate in enhancing glucose entry and in inhibiting dephosphorylation of endogenously formed G-6-P. The latter effect is not exerted by insulin.

Adipocytes↗
PubMed1992

Anticoagulant action of vanadate.

Sodium orthovanadate (vanadate) prolonged the clotting time of normal human plasma in a dose-dependent manner. The prolongation of clotting time by vanadate linearly decreased with an increase in the concentration of amiloride. Vanadate also was completely additive to prolongation by heparin. When factor Xa or thrombin was incubated with vanadate, the amidolytic activity of each decreased in a dose-dependent manner with vanadate. Amiloride protected the decrease of amidolytic activity of both factor Xa and thrombin by vanadate. The amidolytic activity of trypsin also was inhibited by vanadate, but that of alpha-chymotrypsin was not inhibited, suggesting that vanadate preferentially inhibits the amidolytic activity of trypsin and trypsin-like enzymes. These results show that vanadate prolongs the clotting time of plasma through mechanisms involving in part the inhibition of the activity of both factor Xa and thrombin.

Amino Acid Sequence↗
PubMed1988

Insulin-mimetic effects of vanadate in primary cultures of rat hepatocytes.

To evaluate possible mechanisms by which insulin inhibits hepatic apolipoprotein B (apoB) secretion, we incubated primary cultures of rat hepatocytes with sodium orthovanadate, a phosphotyrosine phosphatase inhibitor and insulin-mimetic agent. Vanadate (10 microM) and insulin (10 nM) inhibited the medium accumulation of apoB (secretion) by 21 and 37%, respectively, without increasing intracellular apoB. The effects of insulin and vanadate together were not additive. Both insulin and vanadate enhanced intracellular glycogen accumulation by 82 and 37%, respectively. Unlike insulin, vanadate, at a concentration that inhibited apoB secretion (10 microM), had no effect on intracellular lipogenesis, inhibited the secretion of newly synthesized hepatic proteins, and had a delayed onset and termination of action on inhibition of apoB secretion. At higher concentrations (40 and 80 microM), vanadate stimulated intracellular lipogenesis. In conclusion, our data indicate that vanadate mimics insulin action in hepatocytes with regard to the inhibition of medium accumulation of apoB. These data are consistent with the hypothesis that inhibition of apoB secretion may be secondary to an increase in phosphotyrosine content at its site of synthesis. The kinases responsible for this effect have not been identified. Several effects of vanadate, however, are different from those of insulin, suggesting a differential sensitivity to vanadate, a divergence of the signal transfer by insulin and vanadate at the insulin-receptor or postreceptor level, or both.

Acetates↗
PubMed1990

Insulin-mimetic effects of vanadate. Possible implications for future treatment of diabetes.

Vanadate ions, low-molecular-weight phosphate analogues, mimic most of the rapid actions of insulin in various cell types. When administered orally to diabetic hyperglycemic rats, vanadate reaches the circulation, mimics insulin stimulation of glucose uptake and metabolism, and leads to normoglycemic and partial anabolic states. In addition, vanadate restores tissue responsiveness to insulin and hepatic glycogen levels and activates new synthesis of key enzymes for carbohydrate metabolism. This suggests that correcting hyperglycemia is sufficient to correct the typical metabolic alterations found in streptozocin-induced diabetic rats. Several weeks of oral administration of vanadate to diabetic rats has not produced detectable liver or kidney toxicity. The mechanism by which vanadate mimics the actions of insulin is still obscure. Unlike insulin, vanadate does not seem to stimulate the autophosphorylation and endogenous tyrosine phosphorylation of insulin-receptor kinase or other intracellular proteins either directly or by virtue of its known inhibitory effect on protein phosphotyrosine phosphatase. Results from many studies support a model in which vanadate activates glucose metabolism by either utilizing an alternative (insulin-independent) cascade or bypassing the early events of the insulin-dependent cascade. Either of these possibilities is of clinical importance, because early insulin events may become defective, as a result of severe hyperinsulinemia, and may contribute to insulin resistance. Alternative pathways by which vanadate may stimulate glucose metabolism, e.g., by increasing intracellular Ca2+ levels and/or regulating intracellular and intravesicular pH, are discussed. From a clinical perspective, studies should be continued in evaluating the level of vanadate toxicity after prolonged treatment and searching for agents that potentiate its insulin mimetic actions in vitro and in vivo.

Animals↗
PubMed1982

Effects of vanadate on organic ion accumulation in rat renal cortical slices.

Sodium vanadate is a potent inhibitor of Na-K-adenosine triphosphatase. p-Aminohippurate (PAH) and tetraethylammonium accumulation in rat renal cortical slices was inhibited by vanadate in a dose-dependent manner at medium vanadate concentrations from 10(-6) to 10(-3) M. Inhibition was reversible at vanadate concentrations less than 1.5 x 10(-5) M. The slice content of vanadium (7.5-325 micrometer V/g wt. of tissue) was linearly related to medium vanadate concentrations ranging from 10(-5) to 10(-3) M. The ability of slices to generate glucose and ammonia was not impaired by medium vanadate concentrations up to 5 x 10(-4) M, a concentration that maximally inhibited organic ion accumulation. Increasing medium K+ concentrations potentiated vanadate inhibition of PAH accumulation which correlated with inhibition of sodium pump activity, as determined by 42K+ uptake. Intraperitoneal administration of vanadate (1 or 5 mg V/kg) to rats produced a profound diuresis and natriuresis during the 1st hr. Inhibition of PAH accumulation of renal slices from these rats was related to tissue vanadium concentrations. These data suggest that vanadate exerts its action on proximal tubule transport of PAH via inhibition of Na-K-adenosine triphosphatase.

Ammonia↗
PubMed1993

Role of cytosolic tyrosine kinase in mediating insulin-like actions of vanadate in rat adipocytes.

In previous studies we have identified a cytosolic protein tyrosine kinase (CytPTK) in rat adipocytes that is largely activated in vanadate-pretreated cells (Shisheva, A., and Shechter, Y. (1992) FEBS Lett. 300, 93-96). We report here that staurosporine and its analog K-252a are highly potent (ID50 = 3 and 100 nM, respectively) in inhibiting CytPTK activity of crude cell extract or partially purified enzyme preparations. Staurosporine and K-252a were less effective by more than 2 and 1 orders of magnitude, respectively, in inhibiting insulin receptor-catalyzed PolyGlu4Tyr phosphorylation in cell-free experiments. Preincubation of rat adipocytes with either staurosporine or K-252a selectively blocked the action of vanadate in activating glucose incorporation into lipids and its oxidation. Thus, staurosporine inhibited vanadate-stimulated lipogenesis and glucose oxidation (via glycolysis and the pentose phosphate pathway) in a concentration-dependent manner with ID50 of 75 and 300 nM, respectively. Insulin-stimulated bioeffects were not inhibited at this low range of staurosporine concentration. Staurosporine had no effect on vanadate-stimulated hexose uptake or on vanadate's antilipolytic action. Using staurosporine, we probed those insulinomimetic agents which facilitate their biological activity via the insulin receptor kinase (insulin, wheat germ agglutinin, concanavalin A, and pervanadate) or via CytPTK (vanadate and to a certain degree Mn2+ and Zn2+). These results suggest that (a) vanadate facilitates its insulin-like actions on glucose utilization via the cytosolic tyrosine kinase and (b) this enzyme does not participate in vanadate effects in stimulating hexose uptake and in inhibiting lipolysis. These findings explain further vanadate's post-insulin receptor actions and raise possible application in the management of glucose metabolism in insulin-independent fashion in pathological conditions.

Adipose Tissue↗
PubMed1983

Ciliary but not saltatory movements are inhibited by vanadate microinjected into living cultured cells.

To test the idea that saltatory organelle movements of nonmuscle cells might be driven by microtubule-dynein interactions, we microinjected vanadate into several different types of cultured cell. Solutions of sodium metavanadate made up in a simple buffered salt solution were pressure microinjected into fully spread cells in an open-topped culture chamber placed on the stage of an inverted microscope. The cells were observed by oil-immersion phase-contrast optics and results were recorded on movie film. Vanadate, at 10(-5)-10(-2) M, microinjected into cultured chick embryo fibroblasts, failed to inhibit organelle movements. To test the effectiveness of vanadate's inhibitory action under living cell conditions, ciliated epithelial cells were microinjected. In these cells even the smallest microinjection of 5 X 10(-5) M vanadate caused an immediate cessation of ciliary beating. Moreover, in cells that were well spread it was found that whereas vanadate, at 5 X 10(-5)-5 X 10(-3) M, inhibited ciliary motion, it failed to inhibit organelle saltations in the same cell. To determine whether vanadate would inhibit a living actin-myosin system, myocardial cells were also microinjected. Following microinjection of 5 X 10(-5) and 5 X 10(-4) M vanadate a temporary tonic contraction (which also occurred following microinjection of buffer alone) was followed by regular beating. Taken together these results demonstrate that in living cell systems microtubule-dynein interactions are as sensitive to vanadate inhibition as they are in demembranated model systems, and that a working actin-myosin system in a living muscle cell does not share this great sensitivity. In light of the pronounced differential inhibitory effects of vanadate on the movements of cilia and organelles, our results suggest that saltatory organelle movements in chick embryo fibroblasts and rabbit oviduct epithelial cells are unlikely to be brought about by microtubule-dynein interactions.

Adenosine Triphosphatases↗