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1.
When sperm of the sea urchin, Hemicentrotus pulcherrimus, are diluted into K+-free seawater, the pH of the suspension gradually decreases, whereas a rapid decline in pH is observed following dilution into regular seawater. Sperm motility and respiration are also activated after dilution into K+-free seawater, but levels of activity are less than those observed following dilution into regular seawater. Upon addition of 10 mM K+ to K+-free seawater, rapid acid release occurs and motility and respiratory rate in sperm are reactivated. The effect of K+ on respiration was competitive with respect to the external Na+ concentrations. Harmaline, a potent inhibitor of Na+/K+-ATPase, causes a decrease in movement and respiration of the sperm. Harmaline does not inhibit the rapid decline in pH, although it depresses the release of acid from mitochondria. These results suggest that external K+ plays an important role in intracellular alkalinization of sea urchin sperm.  相似文献   

2.
The effects of stimulants of fluid secretion on net transepithelial transport of the MRP2 substrate Texas Red and the p-glycoprotein substrate daunorubicin were examined in Malpighian tubules of Drosophila melanogaster. Fluid secretion rates were determined using the Ramsay assay and secreted fluid concentrations of Texas Red and daunorubicin were determined using a microfluorometric technique. Nanoliter droplets of secreted fluid were collected in optically flat glass capillaries and dye concentration was determined from fluorescence intensity measured by confocal laser scanning microscopy. Net transepithelial flux of each compound was then calculated as the product of its concentration in the secreted fluid and the fluid secretion rate. Net transepithelial flux of Texas Red increased when fluid secretion was stimulated by tyramine, cyclic AMP or hypoosmotic saline. Net flux decreased when fluid secretion rate of cAMP-stimulated tubules was reduced by elevating saline osmolality with sucrose. Net transepithelial flux of daunorubicin increased when fluid secretion was stimulated by cAMP. Significant increases in dye flux were seen only when the dyes were present at concentrations close to or greater than the concentration required for half maximal transport. Regression analyses showed that 57- 88% of the change in dye flux was attributable to the change in fluid secretion rate when tubules were stimulated with cAMP, cGMP, or tyramine. The results do not suggest that the effects of tyramine and cAMP are mediated through changes in transepithelial potential, nor do they indicate the direct effects of the stimulants on MRP2-like or p-glycoprotein-like transporters (e.g., via protein kinases). Instead, the results suggest that increases in fluid secretion rate minimize diffusive backflux of these dyes and, thus, facilitate higher rates of net transepithelial transport indirectly.  相似文献   

3.
Abstract: Calcium is required to sustain fast axonal transport in sensory neurons of frog and cat. We studied the Ca2+ dependence of fast axonal transport in the motoneurons of the lower spinal cord from frog. The accumulation of acetylcholinesterase at a crush on the ventral roots was used to follow axonal transport. Two types of experiments were performed: modification of the medium bathing the ventral roots, alone, and modification of the medium bathing the spinal cord and ventral roots. Incubation (17-18 h) of the ventral roots in Ca2+-free medium markedly inhibited acetylcholinesterase transport, a finding that demonstrates a Ca2+ requirement for fast axonal transport in motoneurons; when 4 m M MgCl2 was added to the Ca2+-free medium, transport was also greatly reduced. During incubation of the ventral roots in normal medium supplemented with 0.18 m M CoCl2 transport proceeded normally; but when the Co2+ concentration was raised to 1.8 m M , transport was diminished as drastically as in the Ca2+-free medium. Incubation of the spinal cord and ventral roots in medium containing 0.18 m M CoCl2 did not reduce the accumulation of acetylcholinesterase at the crush. Similarly, accumulation of acetylcholinesterase at a crush on the dorsal root was not significantly reduced by exposure of the dorsal root ganglion and root to 0.18 m M Co2+. Exposure of sensory cell bodies to 0.18 m M Co2+ thus produces differential effects on transport of acetylcholinesterase and on transport of newly synthesized radiolabeled protein.  相似文献   

4.
The organic anion salicylate is a plant secondary metabolite that can protect plants against herbivores. Transport of salicylate across the basolateral membrane of the Malpighian tubules of Drosophila melanogaster was studied using a radioisotope tracer technique. The uptake of [(14)C]salicylate by the Malpighian tubules was active, saturable and Na(+)-dependent; the maximum uptake rate (J(max)) and the half saturation concentration (K(t)) were 12.6 pmoltubule(-1)min(-1) and 30.7micromoll(-1), respectively. In contrast to organic anion transport by vertebrate renal tissues, salicylate uptake was not trans-stimulated by glutarate (0.01-1.0 mmoll(-1)) or cis-inhibited by high concentrations (5 mmoll(-1)) of various alpha-keto acids (glutaric acid, alpha-ketoglutaric acid, succinic acid, and citric acid). Changes in basolateral membrane potential or physiologically relevant changes in bathing saline pH did not affect the rate of [(14)C]salicylate uptake. Ring-structure monocarboxylic acids (benzoic acid, nicotinic acid, gentisic acid, unlabelled salicylic acid, alpha-cyano-4-hydroxycinnamic acid, probenecid, fluorescein, and P-aminohippuric acid) strongly inhibited [(14)C]salicylate uptake rate. In contrast, short-chain monocarboxylic acids had little (butyric acid) or no effect (lactic acid, pyruvic acid, and propionic acid). Our results suggest that salicylate uptake across the basolateral membrane of D. melanogaster Malpighian tubules is mediated by a non-electrogenic, alpha-cyano-4-hydroxycinnamic acid-sensitive, Na(+):salicylate cotransport system.  相似文献   

5.
The organic anion salicylate is a plant secondary metabolite that protects plants against phytophagous insects. In this study, a combination of salicylate-selective microelectrodes and a radioisotope tracer technique was used to study the transepithelial transport of salicylate by the Malpighian tubules of 10 species of insects from five orders. Our results show that salicylate is transported into the lumen of the Malpighian tubules in all the species evaluated, except Rhodnius prolixus. The transepithelial transport of salicylate by the Malpighian tubules of Drosophila simulans, Drosophila erecta, Drosophila sechellia, and Acheta domesticus was saturable, Na+-dependent and inhibited by α-cyano-4-hydroxycinnamic acid. This transport system resembles that previously found in tubules of Drosophila melanogaster. In contrast, transepithelial transport of salicylate by Malpighian tubules of Tenebrio molitor, Plagiodera versicolora, Aedes aegypti, and Trichoplusia ni was unaffected by Na+-free bathing saline. The presence of both salicylate and salicylate metabolites in the secreted fluid samples from the Malpighian tubules of A. domesticus, R. prolixus, T. molitor, and T. ni indicates that insect Malpighian tubules may both transport and metabolize salicylate. The highest capacities to rid the hemolymph of salicylate were found in T. molitor, P. versicolora and Drosphila spp. Our results suggest that transport of salicylate by the Malpighian tubules might contribute to elimination of this organic anion from the hemolymph, particularly in some species that encounter high levels of organic anion in the diet.  相似文献   

6.
Modulation of renal epithelial ion transport allows organisms to maintain ionic and osmotic homeostasis in the face of varying external conditions. The Drosophila melanogaster Malpighian (renal) tubule offers an unparalleled opportunity to study the molecular mechanisms of epithelial ion transport, due to the powerful genetics of this organism and the accessibility of its renal tubules to physiological study. Here, we describe the use of the Ramsay assay to measure fluid secretion rates from isolated fly renal tubules, with the use of ion-specific electrodes to measure sodium and potassium concentrations in the secreted fluid. This assay allows study of transepithelial fluid and ion fluxes of ~20 tubules at a time, without the need to transfer the secreted fluid to a separate apparatus to measure ion concentrations. Genetically distinct tubules can be analyzed to assess the role of specific genes in transport processes. Additionally, the bathing saline can be modified to examine the effects of its chemical characteristics, or drugs or hormones added. In summary, this technique allows the molecular characterization of basic mechanisms of epithelial ion transport in the Drosophila tubule, as well as regulation of these transport mechanisms.  相似文献   

7.
Abstract: The acute effects of serum on sodium-potassium (Na+-K+) pump activity and glucose uptake in cultured rat skeletal muscle were studied. Addition of serum to myo-tubes in phosphate-buffered saline caused Na+-K+ pump activity (as measured by changes in the ouabain-sensitive component of both membrane potential and 86Rb uptake) to increase, with peak effects obtained after 30 min. The effect was blocked completely by treatment with amiloride, but not by tetrodotoxin, which blocks voltage-dependent Na+ channels. On transfer of myotubes to Na+-free, choline buffer, resting Na+-K+ pump activity decreased to about 10% of that in phosphate-buffered saline. Addition of regular serum, but not Na+-free serum, caused Na+-K+ pump activity to increase slightly. Similar results were obtained with serum on glucose uptake, the peak effect being reached within 15 min. Stimulation of glucose uptake by serum was partially reduced by amiloride and was not altered by tetrodotoxin. Removal of external Na+ also eliminated serum effects on glucose uptake. The results demonstrate that there are similar signals involving Na+-H+ exchange for serum-induced increases in Na+-K+ pump activity and glucose transport. The lack of complete blockade of serum-induced elevation of glucose transport suggests an additional, as yet undefined, intracellular signal for stimulation of this transport system.  相似文献   

8.
Abstract: We have previously reported that insulin/insulin-like growth factor (IGF)-I induced the α1 isoform of Na+,K+-ATPase in cultured astrocytes. In this study the effects of insulin/IGF-I on Na+,K+-ATPase activity and cell proliferation were examined in astrocytes cultured under the various conditions, to test the possible involvement of the enzyme activity in the mitogenic action of IGF-I on astrocytes. Insulin increased Na+,K+-ATPase activity and stimulated cell proliferation in subconfluent astrocytes (cultured for 7–14 days in vitro). In contrast, these effects were not observed in confluent cells (cultured for 28 days). Furthermore, insulin stimulated neither the enzyme activity nor [3H]thymidine incorporation in astrocytes preincubated in fetal calf serum-free medium for 2 days (quiescent cells) and treated with dibutyryl cyclic AMP (differentiated cells). The increases in Na+,K+-ATPase activity and expression of the α1 mRNA preceded the mitogenic effect. 125I-IGF-I binding experiment showed that all the cells used here had similar binding characteristics. The insulin-induced increase in enzyme activity was not affected by 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7), and it was observed even in Ca2+-free medium. The stimulation by IGF-I of [3H]thymidine incorporation was attenuated by ouabain and a low external K+ level. These findings suggest that stimulation of Na+,K+-ATPase activity is involved in the mitogenic action of IGF-I on cultured astrocytes.  相似文献   

9.
Abstract: The effects of several neurotoxins and cholinergic antagonists on the nicotine-induced secretion of catecholamines by adrenal medulla cells in culture were investigated. Aconitine, veratridine, and batrachotoxin, in the presence of 1 μ m -tetrodotoxin inhibited the nicotine-stimulated secretion of catecholamines in a dose-dependent manner in Locke's solution. In Na+-free sucrose medium, tetrodotoxin was not required to inhibit the stimulatory effects of aconitine, veratridine, and batrachotoxin, and these agents by themselves inhibited the nicotine-stimulated secretion of catecholamines. Scorpion venom, which also increases the flux of Na+ through tetrodotoxin-sensitive channels, was not an effective inhibitor of nicotine-stimulated secretion. Histrionicotoxin, atropine, hexamethonium, and decamethoniun–as well as the Na+-channel activators–noncompetitively inhibit nicotine-stimulated secretion. The effects of these agents on nicotine-stimulated secretion appear similar to their effects on the inhibition of depolarization at the neuromuscular junction. Reversibility studies suggest that the stimulatory and inhibitory sites of the neurotoxins are different, while studies in Na+-free media suggest that tetrodotoxin-insensitive sodium channels are not involved in the inhibitory effect of the neurotoxins. A possible site of action for the inhibitory effects of the neurotoxins. A possible site of action for the inhibitory effects of the neurotoxins is the nicotinic-receptor-associated ion channel.  相似文献   

10.
Potassium is a major osmolyte used by plant cells. The accumulation rates of K+ in cells may limit the rate of expansion. In the present study, we investigated the involvement of ion channels in K+ uptake using patch clamp technique. Ion currents were quantified in protoplasts of the elongation and emerged blade zone of the developing leaf 3 of barley ( Hordeum vulgare L.). A time-dependent inward-rectifying K+-selective current was observed almost exclusively in elongation zone protoplasts. The current showed characteristics typical of Shaker-type channels. Instantaneous inward current was highest in the epidermis of the emerged blade and selective for Na+ over K+. Selectivity disappeared, and currents decreased or remained the same, depending on tissue, in response to salt treatment. Net accumulation rates of K+ in cells calculated from patch clamp current–voltage curves exceeded rates calculated from membrane potential and K+ concentrations of cells measured in planta by factor 2.5–2.7 at physiological apoplastic K+ concentrations (10–100 m m ). It is concluded that under these conditions, K+ accumulation in growing barley leaf cells is not limited by transport properties of cells. Under saline conditions, down-regulation of voltage-independent channels may reduce the capacity for growth-related K+ accumulation.  相似文献   

11.
Abstract: The temporal resolution of carbon-fiber microelectrodes has been exploited to examine the plasticity of quantal secretory events at individual adrenal medullary cells. The size of individual quantal events, monitored by amperometric oxidation of released catecholamines, was found to be dependent on the extracellular ionic composition, the secretagogue, and the order of depolarization delivery. Release was observed with either exposure to 60 m M K+ in the presence of Ca2+ or exposure to 3 m M Ba2+ in solutions of different pH, with and without external Ca2+. Ba2+ was demonstrated to induce Ca2+-independent exocytotic release for an extended period of time (>4 min) relative to release induced by K+ (∼30 s), which is Ca2+ dependent. In all cases, simultaneous changes of intracellular divalent cations, monitored by fura-2 fluorescence, accompanied quantal release and had a similar time course. Exocytosis caused by Ba2+ in Ca2+-free medium had a larger mean spike area at pH 8.2 than at pH 7.4. When Ba2+-induced spikes measured at pH 7.4 were compared, the spikes in Ca2+-free medium were found to be broader and shorter but had the same area. Release induced by K+ after exposure to Ba2+ was comprised of larger quantal events when compared with preceding K+ stimulations. Finally, spikes obtained with Ba2+ exposure at an extracellular pH of 5.5 had a different shape than those obtained in more basic solutions. These changes in spike size and shape are consistent with the interactions between catecholamines and other intravesicular components.  相似文献   

12.
The K+(86Rb) uptake into the roots and the translocation to the shoots of 11-day-old intact wheat seedlings ( Triticum aestivum L. cv. Martonvásári 8) were investigated using plants grown with different K+ supplies. The effects of environmental conditions (darkness, humidity) and of metabolic and transport inhibitors (oligomycin, disalicylidene-propanediamine, 2,4-dinitriphenol, diethylstilbestrol, colchicine) were also studied. Plants with K content of about 0.2 mmol/g dry weight in the root and 0.5 mmol/g dry weight in the shoot (low K status) showed high K+ uptake into the roots and high translocation rates to the shoots. Both transport processes were very low in plants with K content of more than 1.5 and 2.2 mmol/g dry weight in the root and shoot, respectively (high K status).
Darkness and a relative humidity of the air of 100% did not influence K+ uptake by roots, but did inhibit upward translocation and water transport. Inhibition of photosynthesis and treatments with diethylstilbestrol (10−5 mol/dm3), as well as with colchicine resulted in inhibition of translocation in plants of low K status, but these inhibitors had little effect on K+ uptake by the roots. Oligomycin, 2,4-dinitrophenol and diethylstilbestrol (10−4 mol/dm3), however, inhibited K+ uptake by the roots. In general, K+ transport processes were almost unchanged in plants of high K status. It is concluded that only plants of low K status operating with active K+ transport mechanisms are responsive to environmental factors. In high K+ plants the transport processes are passive and are uncoupled from the metabolic energy flow.  相似文献   

13.
Abstract. Cultivars of hexaploid wheat ( Triticum aestivum cvs. Chinese Spring or PI 178704) and derivatives containing chromosomes from both a cultivar and a wild, salt-tolerant species ( Lophopyrum elongatum or L. ponticum ) were compared to determine differences in growth, ion transport and ion accumulation under salt-stress. Two experiments were conducted in which plants were grown under saline and non-saline conditions and harvested at various lime intervals throughout ontogeny. Under salt-stress the growth rate of the cultivars, as compared to the growth rate of the derivatives, decreased more rapidly later in development. Transport rates from root to shoot of Na+ and Cl reached higher levels in the cultivars. The cultivars accumulated more Na+ and Cl and relatively less K+ in the shoot. The K+/Na+ ratio was higher in the derivatives than in the cultivars from which they were derived. The addition of chromosomes from Lophopyrum species into wheat altered ion accumulation, growth rates, and ion transport rates from root to shoot.  相似文献   

14.
Abstract— Mouse brain slices were depleted of K+ by three 10-min incubations-in oxygenated HEPES-buffered medium lacking glucose and K+. Addition of K+ or Rb+ (or Cs+, to a smaller degree) with glucose, or with succinate, malate, and pyruvate (SMP) before incubation at 37°C with 14C-amino acids restored active low-affinity transport of d -Glu, α-aminoisobutyrate (AIB), GABA, Gly, His, Val, Leu, Lys, and Orn. Ouabain at 1–2μ m with Rb+ was more inhibitory with SMP than with glucose, suggesting that the glycoside may affect specific energy coupling to transport. Valinomycin, in contrast, showed no specificity of inhibition of amino acid uptake with glucose or SMP and K+ or Rb+. Cs+ partially restored amino acid uptake, but Li+ was less effective than Cs +. NaF at 10 m m with SMP + Rb+, or SMP + K+ did not inhibit amino acid uptake. Therefore, it was possible to dissociate glycolysis and Na+, K + -ATPase activity from amino acid transport. The ion replacements for K + that supported active amino acid transport indicate that the specificity of ions in possible ionic gradients for transport energetics should be reexamined.  相似文献   

15.
The effect of phytochrome on K+ transport in guard cells of Commelina communis L. was studied following stomatal movement and using the K+−channel blockers tetraethylammonium (TEA), Cs+ and quinidine. TEA and quinidine prevented stomatal opening and closure in red light, but not when it was supplemented with far-red. This indicates that channels that can be blocked by TEA and quinidine are regulated by phytochrome. Evidence for a phytochrome effect on K+ leakage through other membranal compartments was also found. These phytochrome effects are modified by temperature. Elevated temperature decreases the involvement of channels and increases K+ transport through other membrane compartments, while low temperature causes channel opening and diminishes K+ leakage. The interaction between phytochrome effects and those of temperature is discussed.  相似文献   

16.
Egg jelly induces the degradation of histones as well as the acrosome reaction in the spermatozoa of Asterina pectinifera . Much similar degradation of histones without any apparent morphological changes such as the acrosome reaction was induced in the spermatozoa by merely dispersing them into Na+-free seawater. It required external Ca2+ much less than the jelly-induced one in normal seawater, and was not susceptible to Ca2+-channel antagonists, verapamil and diltiazem. Once spermatozoa were incubated with egg jelly in Ca2+-free seawater, they did not undergo the histone degradation even after subsequent addition of Ca2+, but Na+-free seawater rescued such blockage. Spontaneous acrosome reaction occurred in seawater containing 10–30 mM Na+ in a Ca2+-dependent manner. This reaction was accompanied by a rapid increase in intracellular pH (pHi) followed by a large pHi decrease. Diltiazem blocked a large decrease in pHi but scarcely inhibited the acrosome reaction induced by low-Na+ seawater. Increasing K+ inhibited both pHi changes and the acrosome reaction induced by low-Na+ seawater. Decreasing pH of seawater also inhibited the pHi changes but did not affect the acrosome reaction. Strontium was also effective to induce a rapid increase, followed by a gradual decrease, in pHi and the acrosome reaction.  相似文献   

17.
Shoot activity has been reported to affect rates of ion uptake by plant roots in other ways than merely through supply of assimilates. To elucidate the mechanisms by which a signal from the upper part of the plant controls the rate of K+ and NO3 uptake by roots, both uptake of K+ and NO3 and secretion into the xylem of young sunflower plants ( Helianthus annuus L.) were measured after changes in light intensity.
No close correlation was observed between the uptake of NO3 and that of K+; an increase in light intensity produced a much greater stimulation of NO3 uptake than of K+ uptake. On the other hand, secretion of NO3 into the xylem was tightly coupled to that of K+, and this coupling was strongly disturbed by excision of the root. The results suggest the involvement of the K2-malate shuttle on the regulation by the shoot of K+ and NO3 secretion in the xylem, which is linked to NO3 uptake, while K+ uptake is independent of this regulation mechanism.  相似文献   

18.
It is well known that the motility of spermatozoa in rainbow trout is suppressed by K+. We showed here that although trout sperm are completely immotile in medium containing 5 mM K+, motility was initiated by the subsequent addition of several mM Ca2+, suggesting that both K+and Ca2+are related to the process of the initiation of sperm motility. It was further found that K+channel blockers tetraethylammonium, nonyltriethylammonium, Ba2+and Cs+, as well as the Ca2+channel blocker verapamil, inhibited the initiation of sperm motility at doses at which these reagents inhibit chnnel-related functions in other cells. However, Na+channel blocker, tetrodotoxin and anion channel blocker 4, 4-diisothiocyatatostilbene-2, 2'-disulfonic acid inhibited the motility only at extremely high doses. These results suggest that transport of K+and Ca2+through ion channels at the plasma membrane of spermatozoa is the first event that triggers the initiation of sperm motility in rainbow trout.  相似文献   

19.
Passive fluxes of K+ (86Rb) into roots of sunflower ( Helianthus annuus L. cv. Uniflorus) were determined at low K+ concentration (0.1 and 1.0 mM K+) in the ambient solution. Metabolic uptake of K+ was inhibited by 10−4M 2,4-dinitrophenol (DNP). K+ (86Rb) fluxes were studied both continuously and by time differentiation of uptake. In high K+ roots passive uptake was directly proportional to the K+ concentration of the uptake solution, indicating free diffusion. This assumption was supported by the fact that passive Rb+ uptake was not affected by high K+ concentrations. In low K+ roots the passive uptake of K+ was higher than in high K+ roots. The increase was possibly due to carrier-mediated K+ transport. As K+ effluxes were quantitatively similar to influxes, it is suggested that passive K+ fluxes represent exchange diffusion without relation to net K+ transport.  相似文献   

20.
Using excised roots of Atriplex hortensis L., cv. Gelbe Gartenmelde, the uptake, accumulation and xylem transport of K+ and Na+ have been measured. Influx as well as xylem transport proved to discriminate little between K+ and Na+, when considered in relation to the external solution. Both K+ and Na+ inhibited the uptake and xylem transport of each other to about the same degree. Measurements of intracel-lular Na+ fluxes by means of compartment analysis indicated that the low degree of K/Na discrimination during uptake was due to low influx selectivity. Moreover, K+/Na+ exchange at the plasmalemma was not very efficient in Atriplex roots. In order to establish the basis of the low K/Na discrimination in xylem transport, the rates of K+ and Na+ transport were related to the cytoplasmic K+ and Na+ concentrations to yield the selectivity ratio of transport, S(transport) = (φcx(K) × [Na+]c)/(φcx(Na) × [K+]c). Under all conditions this ratio was far below one indicating that Na+ was favoured during xylem release in excised roots of Atriplex at low external concentrations. The implications of this discrimination in favour of Na+ are discussed with respect to salt tolerance of A. hortensis .  相似文献   

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