A potassium ion diffusion potential causes adrenaline uptake in chromaffin-granule 'ghosts'.
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ABSTRACT:
SUBMITTER: Njus D
PROVIDER: S-EPMC1161097 | biostudies-other | 1979 Jun
REPOSITORIES: biostudies-other
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Project description:Resealed chromaffin-granule ;ghosts' were used to study the steady-state kinetics of catecholamine transport. The pump has a high affinity for (-)-noradrenaline, (-)-adrenaline, tyramine and 5-hydroxytryptamine (serotonin), but a lower affinity for (+)-noradrenaline. The measured rates of incorporation do not conform to Michaelis-Menten kinetics, but affinity constants for the former substrates are in the range 8-18mum. Reserpine is a potent inhibitor. Incorporation as a function of ATP concentration also fails to show simple kinetics; the affinity constant for ATP is deduced to be about 3mm at 1mm-MgCl(2). Adenylyl (betagamma-methylene)diphosphonate is a competitive inhibitor at low concentrations, but inhibits more strongly at high concentrations. The pump has a transition temperature at 29 degrees C and does not seem to be identical with the Mg(2+)-stimulated adenosine triphosphatase of chromaffin granules.
| S-EPMC1168499 | biostudies-other
Project description:1. Ca(2+) ions decreased the surface charge of isolated adrenal-medullary chromaffin granules whether the granules were untreated or previously incubated with neuraminidase. 2. Ca(2+) binding in both cases followed a Langmuir adsorption isotherm. 3. The chromaffin-granule surface was essentially polyanionic with about 10800 anionic sites per granule, of which 4200 sites are capable of binding Ca(2+) ions with an electrochemical free energy of adsorption of -1.32kJ.mol(-1). 4. The surface region of the chromaffin granule was calculated to bind 1306 Ca(2+) ions at 2.2mm-Ca(2+) (ionic strength 0.16mol.litre(-1)). 5. The importance of Ca(2+) binding to the chromaffin-granule surface is discussed in relation to the hypothesis of secretion by exocytosis.
| S-EPMC1168329 | biostudies-other
Project description:'Ghosts' of bovine chromaffin granules, in which the complex mixture of proteins and solutes normally found in the granule matrix is replaced by buffered sucrose are osmotically sensitive. They shrink when the osmotic pressure of the suspension medium is increased, and swell if solute entry is facilitated by the addition of ionophores. Swelling in the presence of ionophores has been used to investigate the passive ion permeability of these membranes. They have a very low permeability to K+ ions (of the order of 10(-10) cm/s); their permeability to protons, Na+ and choline ions is too low to be detected by these methods. Their passive permeability to anions decreases in the order: CNS- greater than I- greater than CCl3CO2- greater than Br- greater than Cl- greater than SO4(2)- greater than CH3CO2-, HCO3-, F-, PO4(3)- the permeability to hiocyanate being of the order of 10(-7) cm/s. Coupled proton and anion entry is extremely slow, except for weak acids. Fluoride, unexpectedly, also appears to enter rapidly when proton/K+ exchange is facilitated by nigericin. In the presence of K+ salts, nigericin, like valinomycin, induces lysis of intact granules, an effect that is not dependent on the presence of a permeant anion, but is dependent on the pH gradient across the membrane.
| S-EPMC1183763 | biostudies-other
Project description:1. Highly purified resealed chromaffin-granule ;ghosts' were assayed for ATPase and ATP-driven H(+)-translocation and 5-hydroxytryptamine-uptake activities, and for 5-hydroxytryptamine uptake driven by an imposed transmembrane H(+)-gradient. The effects of several inhibitors on these activities were studied. 2. Dicyclohexylcarbodi-imide inhibits all of these activities, but not in parallel; at low concentrations it decreases the permeability of the membrane to protons. 3. 4-Chloro-7-nitrobenzofuran (Nbf-Cl) and silicotungstate inhibit ATP-dependent activities, without effect on 5-hydroxytryptamine uptake driven by an imposed H(+)-gradient. 4. Tributyltin chloride inhibits all of the activities. 5. Treatment of the ;ghosts' with low concentrations of urea inhibits 5-hydroxytryptamine uptake and ATP-dependent generation of a transmembrane H(+)-gradient, without inhibiting ATPase activity. 6. Nbf-Cl and silicotungstate are without effect on the rate of leakage of 5-hydroxytryptamine from preloaded ;ghosts', whereas dicyclohexylcarbodi-imide and tributyltin chloride accelerate the rate of leakage. 7. Treatment of the membranes with (14)C-labelled Nbf-Cl labels several proteins; membranes treated with dicyclohexyl[(14)C]carbodi-imide are labelled predominantly in a protein of low molecular weight, which may be analogous to the mitochondrial H(+)-conducting proteolipid. 8. It is concluded that Nbf-Cl and silicotungstate inhibit the H(+)-translocating ATPase of the granule membrane; that dicyclohexylcarbodi-imide inhibits the ATPase, and inhibits 5-hydroxytryptamine accumulation by accelerating leakage of the amine; and that the effects of tributyltin chloride are due to inhibition of the ATPase, and collapse of the transmembrane H(+)-gradient through OH(-)-anion exchange.
| S-EPMC1162091 | biostudies-other
Project description:The Na(+)/K(+)-ATPase restores sodium (Na(+)) and potassium (K(+)) electrochemical gradients dissipated by action potentials and ion-coupled transport processes. As ions are transported, they become transiently trapped between intracellular and extracellular gates. Once the external gate opens, three Na(+) ions are released, followed by the binding and occlusion of two K(+) ions. While the mechanisms of Na(+) release have been well characterized by the study of transient Na(+) currents, smaller and faster transient currents mediated by external K(+) have been more difficult to study. Here we show that external K(+) ions travelling to their binding sites sense only a small fraction of the electric field as they rapidly and simultaneously become occluded. Consistent with these results, molecular dynamics simulations of a pump model show a wide water-filled access channel connecting the binding site to the external solution. These results suggest a mechanism of K(+) gating different from that of Na(+) occlusion.
| S-EPMC4515779 | biostudies-literature
Project description:A method is described for the preparation of resealed chromaffin-granule ;ghosts'. The lysis and rapid purification procedures provide ;ghosts' in approximately 70% yield from crude granules; the preparation contains 0.1mumol of catecholamine/mg of protein (as compared with 2.8mumol/mg in unlysed granules), of which about one third is inside the ;ghosts'. The ;ghosts' retain their ability to accumulate catecholamines, a process dependent on Mg-ATP and inhibited by reserpine, and a simple assay for this transport is described.
| S-EPMC1168498 | biostudies-other
Project description:Granule-plasma membrane docking and fusion can only occur when proteins that enable these reactions are present at the granule-plasma membrane contact. Thus, the mobility of granule membrane proteins may influence docking and membrane fusion. We measured the mobility of vesicle associated membrane protein 2 (VAMP2), synaptotagmin 1 (Syt1), and synaptotagmin 7 (Syt7) in chromaffin granule membranes in living chromaffin cells. We used a method that is not limited by standard optical resolution. A bright flash of strongly decaying evanescent field produced by total internal reflection was used to photobleach GFP-labeled proteins in the granule membrane. Fluorescence recovery occurs as unbleached protein in the granule membrane distal from the glass interface diffuses into the more bleached proximal regions, enabling the measurement of diffusion coefficients. We found that VAMP2-EGFP and Syt7-EGFP are mobile with a diffusion coefficient of ∼3 × 10-10 cm2/s. Syt1-EGFP mobility was below the detection limit. Utilizing these diffusion parameters, we estimated the time required for these proteins to arrive at docking and nascent fusion sites to be many tens of milliseconds. Our analyses raise the possibility that the diffusion characteristics of VAMP2 and Syt proteins could be a factor that influences the rate of exocytosis.
| S-EPMC9265163 | biostudies-literature
Project description:BackgroundCarbon nanoparticles (CNPs) have been reported to boost plant growth, while the mechanism that CNPs enhanced potassium uptake for plant growth has not been reported so far.ResultsIn this study, the function that CNPs promoted potassium uptake in BY-2 cells was established and the potassium accumulated in cells had a significant correlation with the fresh biomass of BY-2 cells. The K+ accumulation in cells increased with the increasing concentration of CNPs. The K+ influx reached high level after treatment with CNPs and was significantly higher than that of the control group and the negative group treated with K+ channels blocker, tetraethylammonium chloride (TEA+). The K+ accumulation was not reduced in the presence of CNPs inhibitors. In the presence of potassium channel blocker TEA+ or CNPs inhibitors, the NKT1 gene expression was changed compared with the control group. The CNPs were found to preferentially transport K+ than other cations determined by rectification of ion current assay (RIC) in a conical nanocapillary.ConclusionsThese results indicated that CNPs upregulated potassium gene expression to enhance K+ accumulation in BY-2 cells. Moreover, it was speculated that the CNPs simulated protein of ion channels via bulk of carboxyl for K+ permeating. These findings will provide support for improving plant growth by carbon nanoparticles.
| S-EPMC6986061 | biostudies-literature
Project description:1. When yeast oxidizes ethanol at different pH values the uptake of K(+) corresponds closely to the amount of acetate accumulated at each pH value. 2. The addition of semicarbazide to the suspension buffered at pH4.75 inhibited both the K(+) uptake and the acetate accumulation by about 50%. 3. The addition of either acetate or propionate to the suspensions markedly increased the K(+) uptake. 4. The addition of acetate to the suspensions lowered the intracellular pH of the yeast from a resting value of pH5.80 to 5.56. 5. The ratio of the initial rate of K(+) uptake to O(2) consumption was 0.77. This ratio was increased to 1.77 in the presence of 10mmol of propionate/l.
| S-EPMC1178272 | biostudies-other
Project description:Nonaqueous potassium-ion hybrid capacitors (KIHCs) are faced with limited redox reaction kinetics of electrodes for accommodation of large-sized K+. Here, dipotassium terephthalate (K2TP) is applied as an organic negative electrode to provide comparable reaction kinetics with a non-faradaic activated carbon (AC) positive electrode to boost the electrochemical performance of KIHCs. It is revealed that the large exchange current density and fast two-dimensional (2D) diffusion pathways of K+ in K2TP determined by density functional theory (DFT) calculations ensure its fast redox reaction and transport kinetics. The as-constructed KIHC presents both high energy and power densities of 101 W h kg-1 and 2160 W kg-1 based on the mass of the two electrodes (41.5 W h kg-1 and 885.2 W kg-1 based on the mass of the two electrodes and electrolyte), respectively, and a superior capacity retention of 97.7% after 500 cycles. The excellent electrochemical performance is attributed to the fast kinetics, good structural flexibility, and small volume change (9.4%) of K2TP upon K+ insertion/extraction, and its good compatibility with the AC positive electrode in 1,2-dimethoxyethane (DME)-based electrolyte. This will promote application of organic materials in hybrid capacitors and the development of KIHCs.
| S-EPMC6375359 | biostudies-literature