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PubMed2004

Mitochondria-derived oxidative stress induces a heat shock protein response.

In addition to minimizing native and non-native protein aggregations, heat shock proteins (HSPs) regulate programmed cell death pathways. Members of this conserved protein family are in general activated by oxidative stress, and likely help maintain viability under this stress condition. To further our understanding of heat shock protein physiology, we studied whether a specific subtype of oxidative stress, namely that arising from mitochondrial electron transport chain inhibition, induced a heat shock protein response. We exposed human teratocarcinoma (NT2) cells to varying concentrations of the cytochrome oxidase inhibitor sodium azide. Micromolar exposures resulted in a cytoplasm to nucleus translocation of the inducible Hsp70 and of Hsp40, and this was followed by an overall upregulation. The response did not coincide temporally with the onset of azide exposure, but rather was activated when the degree of cytochrome oxidase inhibition (which was progressive over time) surpassed a threshold. Azide did not affect either Hsp70 or Hsp40 dynamics in NT2 rho0 cells, which lack functional electron transport chains. For the azide-exposed native cell line, addition of the antioxidant trolox to the medium abrogated both Hsp70/Hsp40 translocation and upregulation. We conclude that mitochondrial electron transport chain dysfunction activates a heat shock protein response, and that this response is mediated by oxidative stress.

Blotting, Western↗

PubMed1980

Binding of ethidium monoazide to the chromatin in human lymphocytes.

The azide analog of [14C]ethidium bromide was mixed with lymphocytes and photolyzed with visible light. The distribution of azide in the chromatin fraction was found to be 55% in DNA, 28% in protein and 16% in RNA. Label in the DNA portion was found to be almost exclusively in the region digestible with micrococcal nuclease. The parent compound, ethidium bromide, competed with azide for binding sites, illustrating that the azide analog mimics the action of ethidium bromide.

Affinity Labels↗

PubMed1979

Protected natural peptides as intermediates for preparing semisynthetic peptides and protein analogs. Selective acylation of epsilon-amino groups in cyanogen bromide peptides of cytochrome c using azidoformates.

Treatment of CNBr peptides 66--80, 81--104 and 66--104 from cytochrome c with t-butyloxycarbonyl azide leads to selective acylation of the epsilon-amino groups of lysine residues and the phenolic hydroxyl groups of tyrosine residues with less than 25% acylation of the alpha-amino groups. Similar selectivity was obtained for reactions of benzyloxycarbonyl azide, p-nitrobenzyloxycarbonyl azide and p-methoxybenzyloxycarbonyl azide with peptide 81--104. All of these protective groups can be removed under mild conditions, and thus, the partially protected natural peptides are desirable intermediates for the preparation of semisynthetic peptides. Model condensation reactions of N alpha t-butyloxycarbonyl methionine N-hydroxysuccinimide ester with Z-protected peptide 81--104 produced a peptide corresponding to residues 80 to 104 of cytochrome c in 94% yield.

Acylation↗

PubMed1984

Effects of decoquinate and clopidol on electron transport in mitochondria of Eimeria tenella (Apicomplexa: Coccidia).

Resistance of the chicken coccidium Eimeria tenella to the anticoccidial agents decoquinate and clopidol, and the synergistic activity of mixtures of these compounds have been confirmed in vivo. Inhibition of electron transport by decoquinate and clopidol has been studied in mitochondria isolated from unsporulated oocysts of the same lines of E. tenella as those used in the in vivo studies. Electron transport in mitochondria of a drug sensitive line was susceptible to inhibition by both decoquinate and clopidol, and mitochondria isolated from lines made resistant to one or the other of these compounds showed a corresponding resistance at the level of electron transport. Combinations of low concentrations of decoquinate and clopidol caused a greater inhibition of electron transport than expected from summation of their individual actions. Isobolograms showed that decoquinate and clopidol in fact potentiated each other's effect on electron transport. Induced resistance to either decoquinate or clopidol resulted in an increased sensitivity of electron transport to inhibition by the other drug. Cytochrome spectra of E. tenella mitochondria and a biphasic response of NADH-oxidase and terminal oxidase activity to inhibition by cyanide or azide suggest the presence of two functional terminal oxidases. There is a correlation between the resistance of electron transport to inhibition by decoquinate or clopidol and the susceptibility to inhibition by cyanide or azide. Mitochondrial electron transport that is more resistant to inhibition by decoquinate exhibits greater sensitivity to cyanide and azide; electron transport that is more resistant to inhibition by clopidol exhibits a decreased sensitivity to cyanide and azide. Resistance to decoquinate and clopidol is discussed in view of a possibly branched electron transport chain in mitochondria of E. tenella.

Animals↗

PubMed1994

Effects of deuterium labeling on azido amino acid mutagenicity in Salmonella typhimurium.

The mutagenic effects of azide (N3-) anion in bacterial test systems require the formation of the novel mutagenic metabolite, 3-azido-L-alanine (AZAL). Although the mechanism of AZAL-induced mutagenicity is unknown, subsequent bioactivation of this metabolite appears likely. Earlier studies have shown that other azide-containing amino acids are mutagenic as well. In fact, the mutagenic potency of the synthetic AZAL homologue, L-2-amino-4-azidobutanoic acid (HomoAZAL), was several times that of AZAL. To gain insight into the biochemical processing and mutagenicity of azido amino acids in Salmonella typhimurium, several specifically deuterium-labeled azido amino acids have been prepared and tested for mutagenic potency. In addition, the effect of (aminooxy)acetic acid (AOA) (a potent inhibitor of pyridoxal-dependent processes) on AZAL-induced mutagenesis was examined. The results showed that 2-deuterium substitution of AZAL resulted in a slight increase in mutagenic potency, while AOA treatment resulted in no change in AZAL potency. Taken together these findings did not support the involvement of pyridoxal-dependent processes in AZAL bioactivation. In contrast, deuterium substitution adjacent to the azide group in HomoAZAL and 5-azido-L-norvaline (N3-Norval) resulted in a large decrease in mutagenic potency when compared to the non-deuterium labeled compounds. These observations are consistent with a bioactivation mechanism involving rate-limiting C-H bond cleavage in the formation of the ultimate mutagen. Moreover, this effect of deuterium labeling points to processing of the azide-containing side chain as a key feature in the mutagenic activation mechanism.

Alanine↗

PubMed1993

Albumin grafting on to polypropylene by thermal activation.

4-azido-2-nitrophenyl albumin (ANP-albumin) was prepared by reacting 4-fluoro-3-nitrophenyl azide with albumin. The thermal decomposition kinetics of phenyl azide of ANP-albumin was studied at various temperatures by Fourier-Transform Infrared (FTIR) spectroscopy. The decomposition rate of the phenyl azide increased with temperature. The activation energy for the first-order decomposition of the phenyl azide was 128.0 kJ/mol. Albumin was grafted on to polypropylene (PP) films by thermolysis of the azido groups of ANP-albumin with no premodification of the PP surface. The albumin-grafted surface was characterized by electron spectroscopy for chemical analysis (ESCA) and by quantitative determination of platelet adhesion and activation. The bulk concentration of ANP-albumin used for adsorption varied from 0.001 to 30 mg ml-1, and the albumin-adsorbed PP films were incubated at 100 degrees C for up to 7 h. The carbon and nitrogen peaks resulting from the grafted albumin were used to compare the surface albumin concentrations as a function of the concentration of ANP-albumin in the adsorption solution. When the PP film was adsorbed with ANP-albumin at the concentration of 5 mg ml-1 or higher and incubated at 100 degrees C for longer than 5 h, the surface became resistant to platelet adhesion. The ANP-albumin can be grafted on to chemically inert surfaces such as PP surface through simple thermolysis of azido groups to prevent platelet adhesion and activation.

Adsorption↗

PubMed2004

Differential accumulation of Glut1 in the non-DRM domain of the plasma membrane in response to the inhibition of oxidative phosphorylation.

Approximately 50% of Glut1 in the plasma membrane of Clone 9 cells is localized to the detergent-resistant membrane (DRM) fraction. Acute exposure (90 min) to 5mM azide stimulated glucose transport by approximately 4.7-fold and increased the abundance of Glut1 in the non-DRM fraction of the plasma membrane by approximately 2.9-fold while the abundance of Glut1 in the DRMs was not changed. In parallel experiments, approximately 17 h exposure to azide further increased the rate of glucose transport over that observed at 90 min by approximately 33% and increased plasma membrane Glut1 content by approximately 3.5-fold over control. The increase in total plasma membrane Glut1 reflected a approximately 4.7-fold increase of Glut1 content in the non-DRM fraction and a approximately 2.6-fold increase in the DRMs. We conclude that acute exposure to azide increases Glut1 content in the non-DRM fractions, while prolonged exposure to azide increases the Glut1 content in both non-DRM and DRM fractions. These changes may play an important role in the stimulation of glucose transport in response to the inhibition of oxidative phosphorylation.

Animals↗

PubMed2005

Severe cytochrome c oxidase inhibition in vivo does not induce a pyrimidine deficiency; neuroprotective action of oral uridine prodrug PN401 requires supraphysiological levels of uridine.

It has been hypothesized that mitochondrial respiratory chain dysfunction leads to a pyrimidine deficiency since the pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase is coupled to the electron transport chain. The uridine prodrug triacetyluridine (PN401) is neuroprotective in several models of neurodegenerative disease involving respiratory chain toxins. Therefore, the therapeutic effects of PN401 might involve the correction of a pyrimidine deficiency secondary to respiratory chain impairment. We infused mice with the cytochrome c oxidase inhibitor azide, which inhibited brain complex IV activity. Chronic infusion of azide for 2 or 14 days induced significant toxicity and mortality but did not cause a pyrimidine deficit in the brain. In contrast, the pyrimidine synthesis inhibitor N-phosphonoacetyl-l-aspartate (PALA) produced a pyrimidine deficit with minimal mortality. Treatment with 6% PN401 decreased mortality and cerebrocortical apoptosis caused by azide. Previously, we found that optimal neuroprotection against mitochondrial complex II inhibition required 4-6% PN401. PN401 at 1, 3, 6 and 10% in chow induced nonlinear increases in plasma uridine with 6% PN401 elevating plasma uridine up to 80 muM, and these higher micromolar uridine levels were also required for neuroprotection in chemical hypoxia models in vitro. Our results indicate that severe complex IV inhibition in vivo does not lead to a pyrimidine deficiency, and therefore the protective effect of PN401 in the azide toxin model is not mediated through the correction of a pyrimidine deficiency. Furthermore, supraphysiological levels of uridine are required to produce optimal protective effects in disorders involving impairment of mitochondrial respiratory complex II or IV.

Acetates↗

PubMed2004

Catalase inhibition in the Arcuate nucleus blocks ethanol effects on the locomotor activity of rats.

Previous studies have demonstrated that there is a bidirectional modulation of ethanol-induced locomotion produced by drugs that regulate brain catalase activity. In the present study we have assessed the effect in rats of intraperitoneal, intraventricular or intracraneal administration of the catalase inhibitor sodium azide in the locomotor changes observed after ethanol (1 g/kg) administration. Our results show that sodium azide prevents the effects of ethanol in rats locomotion not only when sodium azide was systemically administered but also when it was intraventricularly injected, then confirming that the interaction between catalase and ethanol takes place in Central Nervous System (CNS). Even more interestingly, the same results were observed when sodium azide administration was restricted to the hypothalamic Arcuate nucleus (ARC), a brain region which has one of the highest levels of expression of catalase. Therefore, the results of the present study not only confirm a role for brain catalase in the mediation of ethanol-induced locomotor changes in rodents but also point to the ARC as a major neuroanatomical location for this interaction. These results are in agreement with our reports showing that ethanol-induced locomotor changes are clearly dependent of the ARC integrity and, especially of the POMc-synthesising neurons of this nucleus. According to these data we propose a model in which ethanol oxidation via catalase could produce acetaldehyde into the ARC and to promote a release of beta-endorphins that would activate opioid receptors to produce locomotion and other ethanol-induced neurobehavioural changes.

Analysis of Variance↗

PubMed1999

Cytotoxic potential of the phytochemical genistein isoflavone (4',5',7-trihydroxyisoflavone) and certain environmental chemical compounds on testicular cells.

The effects of genistein (Gn), sodium azide (naz), and dexamethasone (dxm) on testicular cells TM3, TM4 and GC-1 spg were studied in vitro. First, a series of experiments were performed to assess the response of the cells to the exposure of Gn, naz, dxm, a combination of Gn with naz and Gn with dxm. Trypan blue exclusion assay was used to determine the percentage of viability, and LDH-cytotoxicity test was used to assess the degree of treatment-induced cytotoxicity on each cell type. A second series of experiments were performed to study cytomorphology and determine the type and percentage of treatment-induced cell death (apoptosis and necrosis) on each cell line, using fluorescent dye technique to detect apoptotic and necrotic cells, and tunnel assay to confirm apoptosis. The results from the data obtained demonstrated: i) that incubation of testis cells with each of the agents (Gn, dxm, naz) alone and in two combinations (Gn-dxm, and Gn-naz) induced significant testicular cell death; ii) that both genistein and dexamethasone mostly and significantly induced apoptotic cell death while sodium azide induced necrotic cell death; iii) that addition of dexamethasone to genistein demonstrated synergism in apoptosis on testis cells; and iv) that combination of naz with Gn demonstrated synergism in necrosis on testis cells even though Gn alone did not induce significant necrosis. It is concluded that the synergistic actions of genistein and dxm, and of genistein + sodium azide in induction of apoptosis and/or necrosis may be of clinical and pathophysiological research interest considering the chemopreventive and chemotherapeutic potential of genistein; and the clinico-pharmacological application of dexamethasone and sodium azide.

Animals↗

PubMed1999

Energy metabolism inhibition impairs amyloid precursor protein secretion from Alzheimer's fibroblasts.

The present study investigates the influence of aglycemia and sodium azide (a Cytochrome c Oxidase inhibitor) on sAPP secretion from skin fibroblasts derived from sporadic AD patients and control subjects. Aglycemia reduced sAPP release in the medium of both AD and control fibroblasts to a similar extent after 2 h incubation. Treatment for 2 h with increasing azide concentrations (1 microM-100 mM) under glucose deprivation did not significantly affect sAPP secretion from control fibroblasts, but was able to significantly inhibit sAPP secretion from AD fibroblasts (maximal inhibition 51%). The failure of antioxidants like glutathione (GSH) or N-acetylcysteine (NAC) to antagonize the azide effect on AD fibroblasts and lipoperoxidation data seemed to rule out the possibility that oxidative stress could mediate the sodium azide effect on sAPP release from AD fibroblasts.

Acetylcysteine↗

PubMed2006

"Click chemistry" in the preparation of porous polymer-based particulate stationary phases for mu-HPLC separation of peptides and proteins.

With the use of the copper(I)-catalyzed (3 + 2) azide-alkyne cycloaddition, an element of "click chemistry," stationary phases carrying long alkyl chains or soybean trypsin inhibitor have been prepared for use in HPLC separations in the reversed-phase and affinity modes, respectively. The ligands were attached via a triazole ring to size monodisperse porous beads containing either alkyne or azide pendant functionalities. Alkyne-containing beads prepared by direct copolymerization of propargyl acrylate with ethylene dimethacrylate were allowed to react with azidooctadecane to give a reversed-phase sorbent. Azide-functionalized beads were prepared by chemical modification of glycidyl methacrylate particles. Subsequent reaction with a terminal aliphatic alkyne produced a reversed-phase sorbent similar to that obtained from the alkyne beads. Soybean trypsin inhibitor was functionalized with N-(4-pentynoyloxy)succinimide to carry alkyne groups and then allowed to react with the azide-containing beads to produce an affinity sorbent for trypsin. The performance of these stationary phases was demonstrated with the HPLC separations of a variety of peptides and proteins.

Alkynes↗

PubMedDate not supplied

A laser flash photolysis study of di-, tri- and tetrafluorinated phenylnitrenes; implications for photoaffinity labeling.

Several fluorinated aryl azides (2,6 and 2,4-difluorophenyl azide, 2,4,6-triflurorophenyl azide, and methyl 4-azido-2,3,5,6-tetrafluorobenzoate) were studied by laser flash photolysis techniques. In each case, the azides from singlet nitrenes which can be intercepted with pyridine to form ylides. This allowed determination of the rate constant for ring expansion of the singlet nitrene to the dehydroazepine and measurement of the absolute rate constants for reaction of the singlet nitrenes with typical quenchers.

Affinity Labels↗

PubMed1993

Alternative model for mechanism-based inhibition of Escherichia coli ribonucleotide reductase by 2'-azido-2'-deoxyuridine 5'-diphosphate.

Ribonucleotide reductase (RDPR) from Escherichia coli is composed of two subunits, R1 and R2, and catalyzes the conversion of nucleotides to deoxynucleotides. The mechanism of inactivation of RDPR by 2'-azido-2'-deoxynucleoside 5'-diphosphate (N3UDP) has been examined using a variety of isotopically labeled derivatives: (1'-, 2'-, 3'-, or 4'-[2H])-N3UDPs and 2'-[15N3, 13C]-N3UDP. Electron paramagnetic resonance (EPR) and electron spin echo envelope modulation (ESEEM) spectroscopy studies using these compounds indicate that the 2' carbon-nitrogen bond to the azide moiety is cleaved prior to or upon formation of the nitrogen-centered radical derived from the azide moiety of N3UDP. EPR studies reveal no hyperfine interactions of the nitrogen-centered radical with the 1', 2', 3', or 4' hydrogens of N3UDP. ESEEM studies however, reveal that the 1' and 4' deuterons are 3.3 +/- 0.2 and 2.6 +/- 0.3 A, respectively, from the nitrogen-centered radical. Further support for carbon-nitrogen bond cleavage is derived from studies of the interaction of oxidized R1, C225SR1, and C462SR1 with R2 and N3UDP. In all three cases, in contrast to the results with the wild type R1, azide is detected. Nitrogen-centered radical is not observed with either oxidized R1 or C225SR1 but is observed with C462SR1. These results suggest that C225 is required for the conversion of azide into N2 and a nitrogen-centered radical. The dynamics of the inactivation of RDPR by N3UDP have also been examined. Use of [3'-2H]N3UDP reveals an isotope effect of approximately 2 on the loss of the tyrosyl radical and the rate of inactivation of RDPR. In both cases the kinetics are complex, suggesting multiple modes of inactivation. In addition, several modes of inactivation are required to explain the observation that loss of the tyrosyl radical is slower than the rate of inactivation. Studies using [5'-3H]N3UDP reveal that the rapid inactivation is the result of the formation of a tight noncovalent complex between modified nucleotide, nitrogen-centered radical and RDPR. Destruction of the nitrogen-centered radical is a slow process which appears to be accompanied by decomposition of the modified nucleotide into PPi, uracil, and 2-methylene-3(2H)-furanone. The latter covalently modifies R1 and ultimately leads to loss of approximately 50% of the activity of R1.

Azides↗

PubMed1975

Kinetic and equilibrium studies of the ligand binding reactions of eight electrophoretic components of sperm whale ferrimyoglobin.

The reactions of eight electrophoretic components of sperm whale ferrimyoglobin with fluoride, azide, and cyanide have been studied. There do not appear to be significant differences in rate constants or equilibrium constants among the various components. We conclude that at pH 7.0 in 0.05 M potassium phosphate these ligand binding kinetics and equilibria are insensitive to the net charge on the protein. The variation of the azide equilibrium constant with ionic strength from mu 0.01 M to 0.11 M is not in accord with the predictions of the Debye-Hückel theory. On the other hand, azide association kinetic and equilibrium constants are respectively six- and threefold greater for beef ferrimyoglobin than for the isoelectric whale ferrimyoglobin (band V). An examination of the data for whale, horse, and beef myoglobins reveals that large differences in azide (but not CO) association rate constants are associated with amino acid substitutions at residues 45 and 99 in the heme cavity.

Animals↗

PubMed2006

Effects of substrate analogues and pH on manganese superoxide dismutases.

The effect of the substrate analogues azide and fluoride on the manganese(II) zero-field interactions of different manganese-containing superoxide dismutases (SOD) was measured using high-field electron paramagnetic resonance spectroscopy. Two cambialistic types, proteins that are active with manganese or iron, were studied along with two that were only active with iron and another that was only active with manganese. It was found that azide was able to coordinate directly to the pentacoordinated Mn(II) site of only the MnSOD from Escherichia coli and the cambialistic SOD from Rhodobacter capsulatus. The formation of a hexacoordinate azide-bound center was characterized by a large reduction in the Mn(II) zero-field interaction. In contrast, all five SODs were affected by fluoride, but no evidence for hexacoordinate Mn(II) formation was detected. For both azide and fluoride, the extent of binding was no more than 50%, implying either that a second binding site was present or that binding was self-limiting. Only the Mn(II) zero-field interactions of the two SODs that had little or no activity with manganese were found to be significantly affected by pH, the manganese-substituted iron superoxide dismutase from E. coli and the Gly155Thr mutant of the cambialistic SOD from Porphyromonas gingivalis. A model for anion binding and the observed pK involving tyrosine-34 is presented.

Azides↗

PubMed2000

Total syntheses of (+)-australine and (-)-7-epialexine.

Three approaches were examined for the synthesis of 3-(hydroxymethyl)pyrrolizidines, a class of compounds that includes the polyhydroxylated pyrrolizidine alkaloids alexine (1), australine (2), and various stereoisomers of thereof. In the first approach, the intramolecular cycloaddition of an azide onto an electron-rich 1, 3-diene bearing a terminal alkoxymethyl substituent (i.e., 21) afforded the dehydropyrrolizidines 22a and 22b, with 22a predominating. A rationale for this stereoselectivity was proposed. Transformation of the major diastereomer 22a into a natural 3-(hydroxymethyl)pyrrolizidine was not possible due to difficulties encountered in transforming the phenyl vinyl sulfide functionality into other useful functional groups. A second approach was examined, wherein the intramolecular cycloaddition of an azide with an optically pure S-t-Bu-substituted diene (i.e., 30) was found to produce the pyrrolizidine 31. In this case, the alkoxymethyl substituent was incorporated into the tether between the azide and the diene, rather than on the diene itself. A key transformation in the synthesis of the diene 30 was the use of the allylic borane R(2)BCH(2)CH=C(TMS)(StBu) for the stereoselective conversion of the D-arabinose-derived azido aldehyde 28 to the E-isomer of 30. The cyclization of 30 to 31 also produced the bicyclic triazene 32, the result of 1,3-dipolar cycloaddition of the azide onto the distal double bond of the diene. Again, difficulties in transformation of the vinyl sulfide functionality of 31 into useful oxygen functionality limited this approach to naturally occurring 3-(hydroxymethyl)pyrrolizidines. A third approach to these compounds was successful. The transformation of L-xylose into the azido epoxy tosylate 46 was accomplished using two Wittig reactions and an epoxidation, in addition to other standard functional group manipulations. Reductive double-cyclization of 46 afforded the pyrrolizidines 47a and 47b, which were debenzylated to afford (+)-australine 2 and (-)-7-epialexine 4, respectively. In the preliminary report of this work, erroneous spectroscopic data in the original literature on the structural assignment of australine led to the conclusion that the synthetic material obtained herein was actually (+)-7-epiaustraline. Recently corrected spectroscopic data have appeared which verify that (+)-australine 2 was indeed synthesized for the first time.

Azides↗

PubMed1999

Mechanism of stimulation of glucose transport in response to inhibition of oxidative phosphorylation: analysis with myc-tagged Glut1.

To determine the role of 'translocation' vs. 'activation' of Glut1 in the stimulation of glucose transport in response to inhibition of oxidative phosphorylation, we measured the abundance of myc-tagged Glut1 in plasma membrane of stably transfected Clone 9 cells, a rat liver cell line expressing only the Glut1 isoform. The myc epitope-tag is located between Ile56 and Pro57 in the putative first extracellular loop of Glut1. Under basal conditions, transfected cells expressed approximately 3 fold higher levels of Glut1 and exhibited a approximately 3 fold higher rate of glucose transport than non-transfected cells. To delineate the mechanism mediating the stimulation of glucose transport by a azide we employed two strategies: (1) mild cell surface biotinylation followed by isolation of plasma membranes and quantitation of Glut1 sites in Western blots employing anti-Glut1 and anti-myc antibodies, and (2) quantitative immunofluorescence of myc epitopes in plasma membrane sheets. The rate of glucose transport increased 2.9 +/- 0.5 fold in transfected cells exposed to 5 mM azide for 1 h. Exposure to azide, however, resulted in no significant increase in Glut1 content of plasma membranes using anti-Glut1 or anti-myc antibodies in Western blots (1.0 +/- 0.1 and 0.9 +/- 0.2 fold, respectively; azide/control), and was associated with no detectable increase in immunofluorescence using either anti-Glut1 or anti-myc antibodies (p > 0.1 for both measurements). Treatment of cells with cobalt chloride (employed as a positive control) resulted in marked increases in glucose transport, cell and plasma membrane Glut1 content, and immunofluorescence of plasma membrane sheets (8-10 fold increase in each parameter). We conclude that the stimulation of glucose transport by azide results mainly from activation of Glut1 transporters pre-existing in the plasma membrane.

Animals↗