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1.
The effects of 5-hydroxytryptamine and 5-hydroxytryptophan on 86Rb+ efflux from prelabelled ob/ob-mouse islets were studied to better understand the cellular mechanisms underlying the effects of 5-hydroxytryptamine and 5-hydroxytryptophan on insulin release. 5-Hydroxytryptophan (4 mM) had no effect on 86Rb+ efflux either at a low (3 mM) or at a high (20 mM) d-glucose concentration, whereas 5-hydroxytryptamine (4 mM) stimulated 86Rb+ efflux at both glucose concentrations. These results indicate that 5-hydroxytryptamine may reduce glucose-induced insulin release by inhibiting early steps in the β-cell stimulus-secretion coupling. 相似文献
2.
Anita M. Hennige Nicolas Lembert Martin A. Wahl Hermann P.T. Ammon 《Free radical research》2013,47(5):507-516
Oxidative stress to B-cells is thought to be of relevance in declining B-cell function and in the process of B-cell destruction. In other tissues including heart, brain and liver, oxidative stress has been shown to elevate the intracellular free calcium concentration and to provoke potassium efflux. We studied the effect of oxidative stress on Ca2+ and K+ (Rb+) outflow from pancreatic islets using the thiol oxidants DIP and BuOOH. Both compounds reversibly increased 86Rb+ efflux in the presence of 3 and 16.7 mmol/l glucose. Stimulation of 86Rb+ efflux was also evident in the absence of calcium. DIP evoked release of 45Ca2+ from the pancreatic islets both in the presence or absence of extracellular calcium. Employing inhibitors of the calcium-activated potassium channel (KCa) and the high conductance K+-channel (BKCa), the effect of DIP on 86Rb+ efflux was slightly diminished. Tolbutamide had no effect on 86Rb+ efflux in the presence of DIP. On the other hand thapsigargin, a blocker of the Ca2+-ATPase of the endoplasmic reticulum, completely suppressed the DIP-mediated 86Rb+ outflow. The data suggest that thiol oxidant-induced potassium efflux from pancreatic islets is mainly mediated through liberation of intracellular calcium and subsequent stimulation of calcium-activated potassium efflux. 相似文献
3.
Using detached leaves of potato plants (Solanum tuberosum cv. Bintje) it was shown that abscisic acid (ABA) induced an oriented transport of ions (86Rb,32P and3SS) towards the hormone-treated leaflet. In order to understand the hormone effect on cell permeability, pretreatment of discs
with hormonal solution was conducted. The pretreatment with ABA stimulated the uptake of K+ simultaneously with the decrease of efflux.
The effect of ABA on ion uptake is compared with the action of other hormones, benzylaminopurine (BAP), gibberellic acid (GA3) known to act over a long distance transport, too.
Three kinds of specificity are underlined: - hormonal specificity (stimulation by ABA and inhibiton by BAP of K+ uptake by foliar tissues); - ionic specificity (ABA increase86Rb influx, but inhibits32P and3SS influx); - tissue specificity shown by a comparative study between foliar tissues (source) and tuber tissues (sink). The
autoradiography of foliar discs demonstrated that a treatment of foliar discs can be used, at least for Rb+ (K), to study role of ABA in long-distance transport.
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4.
The effect of glucose on the Ca2+-activated K+ permeability in pancreatic islet cells was investigated by measuring the rate of 86Rb efflux, 45Ca efflux and insulin release from perifused rat pancreatic islets exposed to step-wise increased in glucose concentration. When the glucose concentration was raised from intermediate (8.3 or 11.1 mM) to higher values, a rapid and sustained increase in 86Rb outflow, 45Ca outflow and insulin release was observed. Likewise, in the presence of 8.3 or 16.7 mM glucose, tolbutamide increased 86Rb and 45Ca efflux, as well as insulin release. In the two series of experiments, a tight correlation was found between the magnitude of the changes in 86Rb and 45Ca outflow, respectively. It is concluded that, at variance with current ideas, glucose does not inhibit the response to cytosolic Ca2+ of the Ca2+-sensitive modality of K+ extrusion. On the contrary, as a result of its effect upon Ca2+ handling, glucose stimulates the Ca2+-activated K+ permeability. 相似文献
5.
Jarmo T. Laitinen 《Journal of neurochemistry》1993,61(4):1461-1469
Abstract: Dopamine (DA) stimulated K+ efflux (assessed as 86Rb+ efflux) in retinal suspensions of posthatched chicken. This effect was dose dependent (EC50= 22 μM), was mimicked by the D1-selective antagonist SKF-38393, and reversed by the D1-selective antagonist SCH-23390, indicating an involvement of D1 receptors. Analogues of cyclic AMP (CAMP) did not mimic the DA action. Moreover, DA failed to affect cAMP levels, suggesting that adenylyl cyclase (AC) was not involved. In contrast, forskolin (FSK) stimulated both K+ efflux and cAMP accumulation in the retina (EC50 of 10 μM for both effects). The FSK-elicited K+ efflux was not mimicked by 1,9-dideoxy-FSK (an analogue of FSK that does not activate AC), suggesting that FSK stimulated K+ efflux through the activation of AC. Both DA and FSK inhibited Na+,K+-ATPase activity in the retina. However, the DA-elicited K* efflux was independent of this inhibition, whereas the FSK effect on K+ efflux was largely due to the inhibitory action of the diterpene of the ion pump. A possible role of protein kinase C (PKC) in the DA action was explored. The PKC activator 4β-phorbol 12-myristate 13-acetate (4β-PMA) potently (EC50= 4 nM) stimulated K+ efflux. This action was not mimicked by the inactive isomer 4α-PMA. When added together, DA and 4β-PMA behaved in an additive manner, suggesting separate mechanisms of action for these two drugs. Moreover, DA failed to stimulate retinal phosphoinositide hydrolysis, a well-known pathway leading to PKC activation. These data suggest that DA acting through D1 receptors and independently of AC can modulate its target cell excitability in the chick retina by stimulating K+ efflux pathways. The mechanism of the DA action remains to be clarified. 相似文献
6.
Abstract: The effects of four K+-channel inhibitors on synaptosomal free Ca2+ concentrations and 86Rb+ fluxes are analysed. 4-Aminopyridine, α-dendrotoxin, charybdotoxin, and tetraethylammonium all increase the free Ca2+ concentration, although their potencies differ widely. In each case, the elevation in free Ca2+ concentration is reversed by the subsequent addition of tetrodotoxin. The transient 86Rb+ efflux from preequilibrated synaptosomes induced with high concentrations of veratridine is partially inhibited by 4-aminopyridine and α-dendrotoxin. In contrast, when 4-aminopyridine or α-dendrotoxin is added to polarized synaptosomes, an enhanced86Rb+ flux is seen, both for uptake and for efflux with no change in the total 86Rb+/K+ content of the synaptosomes and with only a slight time-averaged plasma membrane depolarization (6.4 and 3.3 mV, respectively). The enhancements of flux by 4-aminopyridine or α-dendrotoxin are sensitive to ouabain and/or to tetrodotoxin. Furthermore, these flux changes show the same concentration dependencies as the blocked component of veratridine-stimulated 86Rb+ efflux, the elevation of free Ca2+ concentration, and the facilitation of glutamate exocytosis that are elicited by 4-aminopyridine or α-dendrotoxin. It is concluded that these findings support the proposal of spontaneous, repetitive firing of synaptosomes evoked by K+-channel inhibitors and that the enhanced 86Rb+ flux is a consequence of the activity of 4-aminopyridine- and α-dendrotoxin-insensitive K+ channels during these action potentials. 相似文献
7.
During perifusion with medium deprived of Ca2+, addition of glucose or omission of Na+ resulted in prompt and quantitatively similar inhibitions of 45Ca efflux from β-cell rich pancreatic islets microdissected from ob / ob mice. Glucose had no additional inhibitory effect when Na+ was isoosmotically replaced by sucrose or choline+. When K+ was used as a substitute for Na+, the inhibitory effect of Na+ removal on 45Ca efflux became additive to that of glucose. The observation that glucose can be equally effective in inhibiting 45Ca efflux in the presence or absence of Na+ is difficult to reconcile with the postulate that the Na+-Ca2+ countertransport mechanism is a primary site of action for glucose. 相似文献
8.
Sylvia Lindberg 《Physiologia plantarum》1990,79(2):275-282
Three cultivars of sugar beet (Beta vulgaris L.), which are sensitive to aluminium (Al) in the order Primahill > Monohill > Regina, were grown in water culture for 2 weeks. Nutrients were supplied at 15% increase of amounts daily, corresponding to the nutrient demand for maximal growth. The 2.4-dinitrophenol (DNP)-sensitive (metabolic) and DNP-insensitive (non-metabolic) uptake of aluminium, phosphate. 45Ca2+ and K+(86Rb+) in roots were measured as well as transport to shoots of intact plants. All 3 cultivars absorbed more aluminium if DNP was present during the aluminium treatment than in its absence. It is suggested that sugar beets are able to extrude aluminium activity or that they possess an active mechanism to keep Al outside the cell. The presence of Al in the medium during the 1-h experiment affected the metabolic and non-metabolic fluxes of 45Ca2+ and K+(86Rb+) in different ways. In the presence of DNP, the influx of both 45Ca2+ and K+(86Rb+) and the efflux of 45Ca2+ were inhibited by Al in a competitive way. At inhibition of 45Ca2+ influx, 2 Al ions are probably bound per Ca2+ uptake site in cv. Regina (Al-tolerant), but in cvs Primahill and Monohill only one Al ion is bound (more Al sensitive). Aluminium competitively inhibited the active efflux of 45Ca2+ (absence of DNP) in almost the same way in the 3 cultivars. In contrast, aluminium stimulated the influx of K+(86Rb+) in cvs Primahill, Monohill and Regina in the absence of DNP. Thus, the Al effects on active and passive K+(86Rb+) influx are different. The total influx of K+(86Rb+) increased in the presence of Al and might be connected to an active exclusion of Al. Regina is the least Al-sensitive cultivar, probably because Al interferes less with the Ca2+ fluxes and because this cultivar actively excludes phosphate in the presence of Al. Thus Al-phosphate precipitation within the plant could be avoided. 相似文献
9.
S. N. Orlov E. J. Cragoe Jr. O. Hänninen 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1994,164(2):135-140
β-Adrenergic- and volume-dependent regulation of 22Na influx and 86Rb influx and efflux in erythrocytes of brown trout (Salmo trutta m. lacustris) were studied. Norepinephrine (10-6 mol·1-1) increased the rate of 22Na influx 10-to 20-fold via the activation of a Na/H exchanger (ethyl isopropyl amiloride inhibited component of 22Na influx). Unlike carp erythrocytes the activity of the Na, K-pump (ouabain-inhibited 86Rb influx) was only slightly (25–35%) increased by norepinephrine. The norepinephrine-induced increment of Na, K-pump activity
was completely abolished by ethyl isopropyl amiloride thus indicating that this effect was mediated by Na/H exchanger-induced
increase of intracellular Na+ concentration. Cell shrinkage in hyperosmotic media resulted in a several-fold activation of the Na/H exchanger. Cell swelling
in hypotonic media increased both the rate of K, Cl-cotransport [((dihydroindenyl)oxy)alcanaic acidsensitive components of
86Rb influxe and efflux] and passive permeability (leakage) of erythrocyte membranes for Na+ and K+. No volume-dependent regulation of Na, K, 2Cl-cotransport (bumetanide-sensitive components of 86Rb fluxes) was found. It may be concluded that the regulation of monovalent cation transport in erythrocytes of fast-moving
(carnivorous) brown trout differs essentially from that in slowly moving (herbivorous) carp. 相似文献
10.
The effects of an auxin herbicide, 2,4-D, at a concentration of 0.01 mM, on the K+ uptake and efflux of excised roots of wheat (Triticum aestivum L. cv. Rannaya) were investigated at different pH values. The K+ movement was monitored with a K+ (86Rb) tracer. In parallel experiments the ATPase activities of microsomal fractions were determined by the inorganic phosphate liberation method. 2,4-D inhibited the K+ uptake especially at low pH, irrespective of whether Ca2+ was present or not. No marked changes were observed in the K+ efflux properties at pH values above 4. The inhibitory effect on K+ uptake exhibited a correlation with the hydrocarbon solubility of the herbicide, but not with the 2,4-D-induced decrease of the ATPase activity. It is suggested that 2,4-D exerts a non-specific effect on the lipid-protein interactions, giving rise to a generalized alteration of the transport barrier properties of the plasma membrane even at as low a concentration as 0.01 mM. 相似文献
11.
Arnost Kleinzeller Jonathan Goldstein 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1984,154(6):561-571
Summary Osmotic responses of slices of dogfish rectal gland to hypotonic (urea-free) and hypertonic media were studied. Transfer of tissue from isotonic (890 mosM) to hypotonic (550 mosM) saline produced an osmotic swelling associated with a slow net uptake of cell K+ (and Cl–) and a slow, two-component efflux of urea. Media made hypertonic (1180 mosM) by addition of urea or mannitol produced osmotic shrinkage with a net loss of KCl. The cell osmotic responses in hypotonic media were lower than predicted for an ideal osmometer. No volume regulatory responses were seen subsequent to the initial osmotic effects. The cation influx in hypotonic media lacked specificity: in the presence of 0.5 mM ouabain or in K+-free media a net influx of Na+ was found. At steady state, the cell membrane potential evaluated from the Nernst potentials of K+ and triphenylmethyl phosphonium+, was independent of medium tonicity, suggesting the membrane potential as a determinant in the cellular osmotic response. Zero-time86Rb+ fluxes were measured:86Rb+ influx was not affected by hypotonicity, implying an unchanged operation of the Na+–K+-ATPase. On the other hand,86Rb+ efflux was significantly reduced at hypotonicity; this effect was transient, the efflux returning to the control value once the new steady state of cell volume had been reached. A controlled efflux system is therefore involved in the cell osmotic response. The absence of the volume regulatory phenomenon suggests that the cells are not equipped with a volume-sensing mechanism.Abbreviations and symbols
DW
dry weight
-
E
extracellular (polyethylene glycol) space
-
E
Nernst potential
- H2Oe
H2Oi tissue water, extra- and intracellular
-
TPMP
+
triphenyl methyl phosphonium salt
-
WW
wet weight 相似文献
12.
A C Boschero S Kawazu A Sener A Herchuelz W J Malaisse 《Archives of biochemistry and biophysics》1979,196(1):54-63
In isolated rat pancreatic islets, valinomycin (0.01 to 1.0 μm) caused a dose-related facilitation of 86Rb+ outflow and a dose-related inhibition of the glucose-induced changes in both outflow and net uptake of 86Rb+. At high concentrations (0.1–1.0 μm), the ionophore also inhibited the oxidation of glucose and endogenous nutrients, decreased the adenylate charge, and lowered the concentration of reduced pyridine nucleotides in the islet cells. However, as little at 1.0 to 10.0 nm valinomycin caused anomalies in the handling of 45Ca2+ (suppression of the early inhibitory effect of glucose upon 45Ca2+ efflux, and reduction in the amount of 45Ca2+ recovered in the islets after an extensive washing procedure) and inhibition of insulin release. Moreover, when the effect of glucose upon K+ conductance was abolished by high concentrations of valinomycin (0.1–1.0 μm), the glucose-induced secondary rise in 45Ca2+ efflux was still observed. These findings suggest that the effects of glucose upon 86Rb+ and 45Ca2+ handling, respectively, although normally concomitant with one another, can be dissociated, in part at least, from one another. It is concluded that the glucose-induced reduction in K+ outflow may be unnecessary for the sugar to cause a partial remodeling of Ca2+ fluxes in the islet cells. 相似文献
13.
Spring wheat (Triticum aestivum L. cv. Svenno), oat (Avena sativa L. cv. Brighton) and glasshouse cucumber (Cucumis sativus L. cv. Bestseller F1) were cultured for a week after germination on complete nutrient solutions of three different dilutions (1, 25 and 50% of the full strength medium). K+(86Rb) and 45Ca were present during the whole culture period. Relative humidity (RH) was 50% except during the last day, when half the material was transferred to 90% RH. Efflux of labelled ions was then followed during eight hours on unlabelled solutions of the same composition as before, and at both 50% and 90% RH in the atmosphere. – Uptake of K+(86Rb) during growth tended to be saturated in the 25% medium. Contrariwise, the level of Ca2+ in the roots increased continuously with strength of the medium. At low concentrations cucumber roots were higher in Ca2+ than roots of oat or wheat, whereas all three species showed similar levels of Ca2+ in 50% medium. – At the lowest ionic strength, smooth efflux curves were obtained that could be resolved according to the three-compartment theory. At higher ionic strength, irregularities were observed, and more for Ca2+ than for K+; but for practical purposes compartment analysis with the same time constants could be applied as for the lowest concentration. – Discrimination between K+ and Rb+ differed between the roots, but not much between the shoots of different species. The roots of oat and wheat took up Rb+ preferentially over K+ in the 25% and 50% media; whereas K+ was preferred over Rb+ or little discrimination made in 1% medium and for cucumber. The shoots generally showed less discrimination than the roots. The main variability in discrimination between K+ and Rb+ thus appears to be localized in the tonoplasts of the roots cells. – Low RH around the shoots increased efflux of K+(86Rb) from the cytoplasm and vacuoles of the root cells as compared to the efflux at high RH. DNP (2,4-dinitrophenol) in the medium had the same effect as high RH around the shoots. The signal system that must exist between shoots and roots is discussed as a response to “drought” conditions. In relation to investigations of others, it is assumed that the effect of DNP may indicate that part of the chain between roots and shoots consists of metabolically influenced sites, whose output is influenced by the rate of water transport. 相似文献
14.
The fluxes of 22Na+ and 86Rb+ in Arbacia sperm and oocytes were studied in order to determine how these cells carry out cation exchange with the sea environment. The uptake of these ions by serum followed a pattern of early rapid influx (initial 0.5 min) and subsequent efflux (1–3 min) followed by a gradual uptake (after 3 min). Neither the uptake nor the efflux of these cations by Arbacia sperm were affected by ouabain, suggesting that influx and efflux of 22Na+ and 86Rb+ in Arbacia sperm occur predominantly by passive transport. The 22Na+ uptake by Arbacia oocytes showed a steady increase after an initial rapid uptake. A slight but significant inhibition of 22Na+ uptake was observed with ouabain. However, 86Rb+ uptake by the oocytes reached an early equilibrium and was not affected by ouabain. The uptake of Rb+ by Arbacia oocyte is by passive transport while that of Na+ is both by passive and active transport. 相似文献
15.
Jaipaul Singh 《生物化学与生物物理学报:生物膜》1984,775(1):77-85
The effect of acetylcholine and the cholecystokinin-like peptide, caerulein on the fractional efflux of 86Rb+ from preloaded isolated segments of mouse pancreas were studied. Both secretagogues evoked a marked transient increase in 86Rb+ efflux. The removal of Ca2+ from the superfusing medium and addition of 10?4 M EGTA, markedly reduced, but did not abolish the responses to either acetylcholine or caerulein. Furosemide () or piretanide (10?4 M) reduced the basal efflux and inhibited the secretagogue-elicited responses. Stimulus-induced 86Rb+ outflow was abolished when the Cl? component of the superfusing solution was replaced by either NO3?, SO42? or I? but not in case of replacement by Br?, When Na+ was replaced with either Li+ or choline+ both acetylcholine and caerulein failed to elicit any detectable increase in 86Rb+ outflow. However, when Tris+ was substituted for Na+, acetylcholine caused a moderate increase in 86Rb+ efflux which was abolished by either furosemide (10?4 M) or chloride depletion (nitrate substitution). The removal of extracellular K+ or pretreatment with 10?3 M ouabain had little effect on secretagogue-evoked 86Rb+ efflux. These results indicate the presence of a diuretic-sensitive Na+-K+-Cl? cotransport system in the mouse pancreatic acinar cell membrane. 相似文献
16.
The mechanisms by which cationic amino acids influence pancreatic B-cell function have been studied by monitoring simultaneously 86Rb+ efflux and insulin release from perifused rat islets. The effects of two reference amino acids arginine and lysine were compared with those of closely related substances to define the structural requirements for recognition of these molecules as secretagogues. Arginine accelerated 86Rb+ efflux and increased insulin release in the absence or in the presence of 7mm-glucose. Its effects on efflux did not require the presence of extracellular Ca2+ or Na+, but its insulinotropic effects were suppressed in a Ca2+-free medium and inhibited in an Na+-free medium. Among arginine derivatives, only 2-amino-3-guanidinopropionic acid mimicked its effects on 86Rb+ efflux and insulin release; citrulline, guanidinoacetic acid, 3-guanidinopropionic acid and guanidine were inactive. Norvaline and valine also increased 86Rb+ efflux, but their effect required the presence of extracellular Na+; they did not stimulate insulin release. Lysine as well as the shorter-chain cationic amino acids ornithine and 2,4-diaminobutyric acid accelerated 86Rb+ efflux in a Ca2+- and Na+-independent manner. Their stimulation of insulin release was suppressed by Ca2+ omission, but only partially inhibited in an Na+-free medium. The uncharged glutamine and norleucine increased the rate of 86Rb+ efflux in the presence of glucose, only if extracellular Na+ was present. Norleucine slightly increased release in a Ca2+- and Na+-dependent manner. The effects of lysine on efflux and release were not mimicked by other related substances such as 1,5-diaminopentane and 6-aminohexanoic acid. The results suggest that the depolarizing effect of cationic amino acids is due to accumulation of these positively charged molecules in B-cells. This causes acceleration of the efflux of K+ (86Rb+) and activation of the influx of Ca2+ (which triggers insulin release). The prerequisite for the stimulation of B-cells by this mechanism appears to be the presence of a positive charge on the side chain of the amino acid, rather than a specific group. 相似文献
17.
We have previously reported on the biochemical properties of a Na+,K+,2Cl?-cotransport in HeLa cells and here we deal with aspects of its physiological regulation. Na+,K+,2Cl?-cotransport in HeLa cells was studied by 86Rb+ influx and 86Rb+/22Na+ efflux measurements. The effects of rat atrial natriuretic peptide (ANP), isoproterenol, and amino acids on 86Rb+ flux, mediated by the bumet-anide-sensitive Na+, K+, 2Cl?-cotransport system and the ouabain-sensitive Na+/K+-pump, were investigated. ANP reduced bumetanide-sensitive 86Rb+ influx under isotonic as well as under hypertonic conditions. Similar decrease of bumetanide-sensitive 86Rb+ influx was observed in the presence of 8-bromo-cGMP, while neither isoproterenol as a β-receptor agonist nor 8-bromo-cAMP-could alter bumetanide-sensitive 86Rb+ influx. Furthermore, efflux of 86Rb+ and 22Na+ was greatly reduced in the presence of bumetanide and ANP. Together with our recent findings, showing functionally active, high affinity receptors for ANP on HeLa cells (Kort and Koch, Biochim. Biophys. Res. Commun. 168:148–154, 1990), this study indicates that ANP participates in the regulation of the Na+, K+, 2Cl?-cotransport system in HeLa cells. Further measurements revealed that amino acids as present in the growth medium (Joklik's minimal essential medium) and the amino acid derivative α-methyl-aminoisobutyric acid (metAlB, 1 and 5 mM, respectively) also reduced Na+, K+, 2Cl?-cotransport-mediated 86Rb+ uptake and diminished the stimulatory effect of hypertonicity on the cotransporter. In addition, the Na+/K+-pump was markedly stimulated in the presence of amino acids, while neither ANP and 8-Br-cGMP nor isoproterenol and 8-Br-cAMP had a significant effect on the activity of the Na+/K+-pump. 相似文献
18.
Alice Y. Romans Theresa M. Allen William Meckes Robert Chiovetti Lulu Sheng Henri Kercret Jere P. Segrest 《生物化学与生物物理学报:生物膜》1981,642(1):135-148
Human erythrocyte glycophorin is one of the best characterized integral membrane proteins. Reconstitution of the membrane-spanning hydrophobic segment of glycophorin (the tryptic insoluble peptide released when glycophorin is treated with trypsin) with liposomes results in the production of freeze-fracture intrabilayer particles of 80 Å diameter (Segrest, J.P., Gulik-Krzywicki, T. and Sardet, C. (1974) Proc. Natl. Acad. Sci. U.S.A. 71, 3294–3298), with particles appearing at or above a tryptic insoluble peptide concentration of 4 mmol per mol phosphatidylcholine. In the present study, increasing concentrations of tryptic insoluble peptide were added to sonicated small unilamellar egg phosphatidylcholine vesicles and the rate of efflux of 22Na+ was examined by rapid (30 s) gel filtration on Sephadex G-50. Below a concentation of 3–5 mmol tryptic insoluble peptide/mol phosphatidylcholine, 22Na+ efflux occurs at a constant slow rate at given tryptic insoluble peptide concentrations. Above a concentration of 3–5 mM, the rate of efflux is biphasic at given tryptic insoluble peptide concentrations, exhibiting both an initial fast and a subsequent slow component. On the basis of graphic and computer curve-fitting analysis, with increasing tryptic insoluble peptide concentration, the rate of the slow component reaches a plateau at a tryptic insoluble peptide concentration of 3–5 mM and remains essentially constant until much higher concentrations are reached; the fast component increases linearly with increasing tryptic insoluble peptide concentration well beyond 5 mM. The most consistent interpretation of this data is as follows. The slow 22Na+ efflux component is due to perturbations of small unilamellar vesicle integrity by tryptic insoluble peptide monomers. At a tryptic insoluble peptide concentration of 3–5 mmol/mol, a critical concentration is reached following which there is intrabilayer tryptic insoluble peptide self-association. The fast 22Na+ efflux component is due to the increasing presence of tryptic insoluble peptide self-associated multimers the 80-Å particles seen by freeze-fracture electron microscopy) which results in a significantly larger bilayer defect than do tryptic insoluble peptide monomers. The failure of complete saturation of efflux by the fast component is ascribed to the presence of two populations of small unilamellar vesicles, some of which contain tryptic insoluble peptide multimers and some of which do not.Addition of cholesterol to the tryptic insoluble peptide/phosphatidylcholine vesicles decreases the rate of 22Na+ efflux by inhibiting primarily the fast component. Freeze-fracture electron microscopy indicates that the presence of cholesterol has no effect on the size, number or distribution of 80-Å intra-bilayer particles in the tryptic insoluble peptide/phosphatidylcholine vesicles. These results are consistent with a mechanism to explain the fast Na+ efflux component involving protein-lipid boundary perturbations.Efflux of 45Ca2+ from phosphatidylcholine vesicles is also enhanced by incorporation of tryptic insoluble peptide, but only if divalent cations (Ca2+ or Mg2+) are present in the external bathing media as well as inside the sonicated vesicles. If monovalent Na+ only is present in the bathing media no 45Ca2+ efflux is seen. Under conditions where 45Ca2+ efflux is seen, both a fast and a slow component are present, although both appear lower than corresponding rate constants for 22Na+ efflux. These results suggest a coordinated mechanism for ion efflux induced by tryptic insoluble peptide and, together with the 22Na+ efflux studies, may have mechanistic implications for the transbilayer phospholipid exchange (flip-flop) suggesed to be induced at glycophorin/phospholipid interfaces (de Kruiff, B., van Zoelen, E.J.J. and van Deenen, L.L.M. (1978) Biochim. Biophys. Acta 509, 537–542). 相似文献
19.
The mechanism of the protective effect of Ca2+ on cellular K+ content was studied by examination of the effect of Ca2+ on efflux of the K+ analog, 86Rb+, from preloaded cells with the use of compounds which interfere with monovalent cation movements. Ca2+ decreased 86Rb+ efflux to the same extent in the presence and absence of ouabain, suggesting that Ca2+ did not alter the activity of the (Na+ + K+)-adenosine triphosphatase pump. Ca2+ exerted a similar protective effect in the presence of furosemide, an inhibitor of K+-K+ exchange, indicative that Ca2+ was not inhibiting this pathway. Since Ca2+ did not influence these pathways, it is concluded that Ca2+ exerts its primary effect by slowing passive diffusion. In support of this, Ca2+ also slowed 22Na+ efflux. In addition, ethanol-induced leakage of 86Rb+ was reversed by extracellular Ca2+, suggestive of a Ca2+-membrane phospholipid interaction. 相似文献
20.
Distinct effects of acetylcholine and glucose on 45calcium and 86rubidium efflux from mouse pancreatic islets 总被引:1,自引:0,他引:1
The similarities between the effects of acetylcholine and glucose on phospholipid metabolism in pancreatic islet cells prompted the comparison of their effects on ionic fluxes. Acetylcholine (1 μM) consistently increased 45Ca2+ efflux from mouse islets, whereas glucose increased it in the presence, but decreased it in the absence of extracellular Ca2+. Acetylcholine consistently accelerated 86Rb+ efflux, and this effect was augmented by Ca2+ omission. On the other hand, glucose markedly inhibited 86Rb+ efflux, except when its concentration was raised from 10 to 15 mM in the presence of Ca2+. Unlike their effects on phospholipid metabolism, the ionic effects of the two insulin-secretagogues are thus very different. 相似文献