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1.
The fluorescent dye chlorotetracycline was used to study the relationship between the light-induced decrease in cytosolic
free calcium concentration, [Ca2+]c, and its effect on ion transport at the plasma membrane in the giant cells of Chara corallina Klein ex Willd. A kinetic analysis of the simultaneously measured light-induced changes in membrane potential and in [Ca2+]c led to the same time constant of about 40 s. The reversal potential of the light effect on membrane potential was in agreement
with the dominant role of a K+ channel in the plasma membrane. Thus, the experiments reported here provide evidence for the following light-driven signal
transduction chain from the chloroplasts to K+ transport of the plasma membrane: (i) light causes an uptake of Ca2+ into the chloroplasts, (ii) this causes a decrease in cytosolic [Ca2+]c, (iii) this leads to a decrease in the activity of a K+ channel. The results also initiated a re-analysis of previously published data of the light effect on the velocity of cytosolic
streaming and supported the hypothesis that Ca2+ fluxes coming out of the chloroplasts upon darkening cause a Ca2+-induced phosphorylation of myosin, which slows down cytoplasmic streaming.
Received: 3 May 1997 / Accepted: 19 May 1998 相似文献
2.
The inward K+ channels (IKin) of guard cells are inhibited upon application of abscisic acid (ABA). It has been postulated that IKin inhibition requires an elevation in cytosolic free Ca2+ levels ([Ca2+]c) because: (i) experimental increases in [Ca2+] c can mimic the ABA effect, and; (ii) ABA can trigger an elevation of [Ca2+]c in guard cells. However, not all guard cells respond to ABA with a [Ca2+]c increase, and the magnitude of the increases that do occur is variable. Therefore, an obligate role for Ca2+ in the regulation of downstream effectors of ABA response, such as the IKin channels, remains in question. In this study, we developed a methodology for simultaneous patch clamping and confocal ratiometric
Ca2+ imaging of Vicia faba L. guard-cell protoplasts. This allowed us to directly assess the relationship between ABA-induced changes in [Ca2+]c and IKin inhibition. In the presence of extracellular Ca2+, the extent of [Ca2+]c elevation correlated with the extent of IKin inhibition. However, upon chelation of either extracellular Ca2+, [Ca2+]c, or both, extracellular Ca2+ and [Ca2+]c, [Ca2+]c elevation did not occur in response to ABA yet IKin currents were still strongly inhibited. These data illustrate that Ca2+-independent regulation is involved in ABA-inhibition of stomatal opening processes.
Received: 17 September 1999 / Accepted: 26 October 1999 相似文献
3.
Summary A steep, oscillating tip-focused gradient in cytosolic free calcium ([Ca2+]c) has been implicated in pollen tube growth. Further understanding of the biological causes and consequences of these processes relies on the precise imaging of [Ca2+]c during the different growth phases. In this work, the minimum technical requirements for confocal [Ca2+]c imaging ofAgapanthus umbellatus pollen tubes were examined. A range of dyes, dye forms, and loading methods were compared. Non-ratio and ratio imaging were critically analysed, in terms of the detection of the [Ca2+]c gradient and its fluctuations over time. Both ratiometric and nonratiornetric methods detected relative changes in [Ca2+]c. However, visualisation of the [Ca2+]c gradient, with an accurate spatial definition, was only possible with ratiometric methods. The gradient observed in this study ranged from 1.8 M (tip) to 180–220 nM (basal level), within the first 4–10 m. Apical [Ca2+]c fluctuations with an amplitude between 415 nM and 1.8 M showed a period of 40 to 75 s. All protocols for dye-loading proved to have strengths and weaknesses. Thus, the choice of a dye and its loading procedure should consider the required imaging period, extent of sequestration, effect on cell performance and viability, ease of loading procedure, and aim of the study. The present study constitutes an examination of the [Ca2+]c gradient in pollen tubes by these criteria.Abbreviations CLSM
confocal laser scanning microscope
- [Ca2+]c
cytosolic free calcium
- PT
pollen tube
Dedicated to Professor Walter Gustav Url on the occasion of his 70th birthday 相似文献
4.
J. Nagel-Volkmann C. Plieth D. Becker H. Lüthen K. Drffling 《Journal of plant physiology》2009,166(17):1955-1960
Relatively little is known about changes in the cytosolic free calcium ion concentration ([Ca2+]c) in monocotyledonous plants. Therefore, we produced transgenic winter wheat lines stably expressing the calcium-sensitive photoprotein aequorin constitutively in the cytosol. [Ca2+]c was detected in vivo by luminometry, and [Ca2+]c elevations were imaged at video rate. Experiments with the transgenic seedlings focused on potential changes in [Ca2+]c during cold exposure. Temperature-induced changes in [Ca2+]c were found to be more dependent on the change in temperature (dT dt−1) than on the absolute value of temperature. [Ca2+]c increased only at cooling rates higher than 8°C min−1, indicating that an overall cellular [Ca2+]c increase is of minor relevance as a signal for cold acclimation in wheat under ecological conditions. The results are discussed with regard to the so-called ‘calcium signature hypothesis’. 相似文献
5.
David L. Jones Simon Gilroy Paul B. Larsen Stephen H. Howell Leon V. Kochian 《Planta》1998,206(3):378-387
Aluminum inhibition of root growth is a major world agricultural problem where the cause of toxicity has been linked to changes
in cellular calcium homeostasis. Therefore, the effect of aluminum ions (Al) on changes in cytoplasmic free calcium concentration
([Ca2+]c) was followed in root hairs of wild-type, Al-sensitive and Al-resistant mutants of Arabidopsis thaliana (L.) Heynh. Generally, Al exposure resulted in prolonged elevations in tip-localized [Ca2+]c in both wild-type and Al-sensitive root hairs. However, these Al-induced increases in [Ca2+]c were not tightly correlated with growth inhibition, occurring up to 15 min after Al had induced growth to stop. Also, in
32% of root hairs examined growth stopped without a detectable change in [Ca2+]c. In contrast, Al-resistant mutants showed little growth inhibition in response to AlCl3 exposure and in no case was a change in [Ca2+]c observed. Of the other externally applied stresses tested (oxidative and mechanical stress), both were found to inhibit root
hair growth, but only oxidative stress (H2O2, 10 μM) caused a prolonged rise in [Ca2+]c similar to that induced by Al. Again this increase occurred after growth had been inhibited. The lack of a tight correlation
between Al exposure, growth inhibition and altered [Ca2+]c dynamics suggests that although exposure of root hairs to toxic levels of Al causes an alteration in cellular Ca2+ homeostasis, this may not be a required event for Al toxicity. The elevation in [Ca2+]c induced by Al also strongly suggests that the phytotoxic action of Al in root hairs is not through blockage of Ca2+-permeable channels required for Ca2+ influx into the cytoplasm.
Received: 24 October 1997 / Accepted: 6 March 1998 相似文献
6.
David S. Williams 《BioEssays : news and reviews in molecular, cellular and developmental biology》1995,17(4):282-286
Analysis of the light-induced changes of cytosolic Ca2+ ([Ca2+]i) in photoreceptor cells has been taken a step further with two recently published studies(1,2). In one, changes in [Ca2+]i were measured in single detached rod outer segments from Gecko in response to various light intensities. The advances of the other(2) are embodied in its employment of transgenic Drosophila, whose photoreceptors express a visual pigment that is insensitive to the wavelength of light used in the fluorescence imaging of [Ca2+]i. These studies provide a better basis for understanding the regulation of Ca2+-mediated events in photoreceptor cells. 相似文献
7.
Camilla Rozanski Michelle Belton Frank S. Prato Jeffrey J.L. Carson 《Bioelectromagnetics》2009,30(3):213-221
This study investigated whether glutathione depletion affected the sensitivity of HL‐60 cells to static magnetic fields. The effect of Diethylmaleate (DEM) on static magnetic field induced changes in cytosolic free calcium concentration ([Ca2+]c) was examined. Cells were loaded with a fluorescent dye and exposed to a uniform static magnetic field at a strength of 0 mT (sham) or 100 mT. [Ca2+]c was monitored during field and sham exposure using a ratiometric fluorescence spectroscopy system. Cells were activated by the addition of ATP. Metrics extracted from the [Ca2+]c time series included: average [Ca2+]c during the Pre‐Field and Field Conditions, peak [Ca2+]c following ATP activation and the full width at half maximum (FWHM) of the peak ATP response. Comparison of each calcium metric between the sham and 100 mT experiments revealed the following results: average [Ca2+]c measured during the Field condition was 53 ± 2 nM and 58 ± 2 nM for sham and 100 mT groups, respectively. Average FWHM was 51 ± 3 s and 54 ± 3 s for sham and 100 mT groups, respectively. An effect of experimental order on the peak [Ca2+]c response to ATP in sham/sham experiments complicated the statistical analysis and did not allow pooling of the first and second order experiments. No statistically significant difference between the sham and 100 mT groups was observed for any of the calcium metrics. These data suggested that manipulation of free radical buffering capacity in HL‐60 cells did not affect the sensitivity of the cells to a 100 mT static magnetic field. Bioelectromagnetics 30:213–221, 2009. © 2008 Wiley‐Liss, Inc. 相似文献
8.
Isolated characean internodal cells of Nitellopsis obtusa can be stored in artificial pond water for many days, but they cannot survive in 100mol m?3 NaCl solution unless more than several mol m?3 Ca2+ is added. Short-term effects of NaCl stress on the cytosolic concentration of Ca2+ ([Ca2+]c), cytosolic pH (pHc) and vacuolar pH (pHv) were studied in relation to the external concentration of Ca2+ ([Ca2+]e). Changes in [Ca2+]c were measured with light emission from a Ca2+-sensitive photoprotein, semisynthetic fch-aequorin which had been injected into the cytosol. Both pHc and pHv were measured with double-barrelled pH-sensitive microelectrodes. When internodal cells were treated with 100 mol m?3 NaCl (0–1 mol m?3 NaCl (0.1 mol m?3 [Ca2+]e), [Ca2+]c increased and then recovered to the original level within 60 min. The time course of the transient change in [Ca2+]c was not influenced by the level of [Ca2+]c (0.1 and 10 mol m?3). In some cases, the transient increase in [Ca2+]c was induced only by increasing external osmotic pressure with sorbitol. In response to treatment with 100 mol m?3 NaCl (0.1 mol m?3 [Ca2+]c), pHc decreased by 0.1–0.2 units after 10min but recovered after 30–60 min, while pHv increased by 0.4–0.5 units after 2–50 min and tended to recover after 60 min. The initial changes in both pHc and pHv were suppressed when [Ca2+]e was raised from 0.1 to 10mol m?3. These results show that the charophyte alga Nitellopsis can regulate [Ca2+]c, pHc and pHv under NaCl stress in the short term and that the protective effect of Ca2+ on salinity stress is apparently unrelated to perturbation of Ca2+ and pH homeostasis. 相似文献
9.
Using roots from Arabidopsis thaliana expressing the recombinant calcium indicator aequorin, we show that NH3 uptake and alkalisation of plant cells act as a stimulus which induces transient elevations of the cytoplasmic free calcium concentration ([Ca2+]c). The magnitudes of these [Ca2+]c elevations are dependent on the concentration of the membrane permeable form, NH3, and hence, particularly dependent on the pH in the external medium. EGTA and La3+ are able to significantly suppress the [Ca2+]c transients showing that Ca2+ influx through the plasma membrane is likely to be involved. Verapamil and nifedipine had no inhibitory effects, which suggests that Ca2+ release from internal stores might not contribute significantly to the NH3‐triggered [Ca2+]c response. Pre‐incubation in l ‐methionine‐dl ‐sulphoximine – an inhibitor of the glutamine synthetase – did not alter the NH3‐induced [Ca2+]c responses at all. These results are consistent with previous studies where NH3‐induced changes of cytoplasmic and vacuolar pH were investigated in maize roots. Furthermore, the similarity between the kinetics of NH3‐driven cellular pH changes demonstrated in previous studies and the [Ca2+]c transients shown here suggests a direct relationship between [Ca2+]c and cellular alkalisation (cytoplasmic pH and/or vacuolar pH). However, the mechanism behind this possible causal relation remains to be elucidated. 相似文献
10.
A. Verkhratsky 《Neurophysiology》1997,29(4-5):205-212
Various electrical, mechanical, and chemical stimuli, including the influences of neurotrasmitters, neuromodulators, and hormones,
trigger complex changes in [Ca2+]
i
in all types of glial cells. Glial [Ca2+]
i
responses are controlled by coordinated activity of several molecular cascades. The initiation of [Ca2+]
i
signal in glial cells results from activation of either plasmalemmal, or intracellular Ca2+ permeable channels. The interplay of different molecular cascades enables the development of agonist-specific patterns of
Ca2+ responses. Such agonist specificity may provide the means for intracellular and intercellular information coding. Furthermore,
glial [Ca2+]
i
signals can travel with no decrement within glial networks. These intercellular Ca2+ waves can be regarded as a substrate for information exchange between the glial cells. Neuronal activity can trigger [Ca2+]
i
signals in neighboring glial cells and, moreover, there is some evidence that glial [Ca2+]
i
waves can activate neuronal electrical and/or [Ca2+]
i
, responses. Glial Ca2+ signalling can be regarded as a form of glial excitability. 相似文献
11.
Andreas Ziegler Bernd Walz 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1989,165(5):697-709
Summary Superfused slices of drone retina were used for a quantitative analysis of light-induced changes in extracellular Ca2+ concentration ([Ca2+]o) and extracellular space (ECS) volume. 20-ms light flashes elicited biphasic changes in [Ca2+]o. For a saturating flash a brief, initial decrease was followed by a transient increase of 120±34 M. Long, dim steps of light (5 min) produced either a decrease or an increase in [Ca2+]o depending strongly on the previous illumination. Brighter continuous lights caused the [Ca2+]o to increase transiently by 1.4 mM to a peak from which it decayed to a plateau, up to 0.6 mM above the dark concentration.Light flashes (20 ms) caused a shrinkage in ECS volume not exceeding 4%. Thus, changes in [Ca2+]o were almost completely due to Ca2+ fluxes between the ECS and adjacent cells. Continuous lights caused a shrinkage in ECS volume rarely exceeding 16%–20%. Thus, less than 15% of the measured Ca2+ changes could be attributed to shrinkage of the ECS. These data confirm that the ECS functions as a source and a sink for Ca2+ mobilized by light. For comparison, we also made a few measurements of changes in [Ca2+]o in the retina ofCalliphora.Abbreviations [Ca
2+]i
intracellular free Ca2+ concentration
- [Ca
2+]o
extracellular free Ca2+ concentration
-
ECS
extracellular space
-
ER
endoplasmic reticulum
-
TMA
+
tetramethylammonium ion 相似文献
12.
Rui Malhó Nick D. Read M. Salomé Pais Anthony J. Trewavas 《The Plant journal : for cell and molecular biology》1994,5(3):331-341
Growing pollen tubes of Agapanthus umbellatus exhibited a tip-to-base gradient in cytosolic free calcium ([Ca2+]c). Although this gradient is believed to be involved in pollen tube growth, its role in specifying reorientation is unknown. The direction of pollen tube growth could be modified by iontophoretic micro-injection, electrical fields (EFs) or photolysis of caged Ca2+. Iontophoretic injection resulted in a temporary cessation of growth, an increase in [Ca2+]c and, upon recovery, reoriented growth. Weak EFs also elevated [Ca2+]c, reduced growth rates and resulted in the reorientation of pollen tubes towards the cathode. Treatment with very low concentrations of the Ca2+-channel blocker lanthanum chloride, prior to exposure to an EF, inhibited both the increase in [Ca2+]c and reorientation whilst only slightly affecting growth rates. The responses of growth inhibition and reorientation were mimicked when [Ca2+]c was artificially elevated by photoactivating caged Ca2+ (Nitr-5). Our data suggest that [Ca2+]c is part of a transduction mechanism which enables growing pollen tubes to successfully reorient to directional signals in the style and thus accomplish eventual fertilization of the egg. 相似文献
13.
Structural and physiological properties of leech giant glial cells as studied by confocal microscopy
We have studied the architecture of giant neuropile glial cells of the medicinal leech Hirudo medicinalis L. using confocal laser scanning microscopy. We also measured changes in the intracellular Ca2+ concentration ([Ca2+]i) induced by activation of glutamate receptors or voltage-gated Ca2+ channels in different glial cell compartments. Glial cells of isolated segmental ganglia were filled iontophoretically with the Ca2+ indicator dye Fluo-3. The three-dimensional structure, calculated from serial sections, showed that numerous fine glial branches extend within the whole neuropile, where most of the synapses between neurones are established. Activation of glial glutamate receptors by glutamate or kainate, or depolarizing the cell membrane by elevating the external K+ concentration resulted in a transient increase in [Ca2+]i, as measured by Fluo-3 fluorescence. The comparison of [Ca2+]i changes in glial cell branches with changes in the cell body demonstrated that transients in the branches were 2–3 times larger than those in the cell body. The results suggest that glutamate receptors and voltage-gated Ca2+ channels are located in the membrane not only of the glial cell body but also of the cellular branches, which may extend close to synaptic domains. 相似文献
14.
To determine whether lipid-secreting cells have cytosolic Ca2+ concentration ([Ca2+]c)-related secretory mechanisms, morphological changes and intracellular calcium dynamics of Harderian glands of guinea pigs stimulated by secretagogs were studied by electron microspy and Fura-2/AM digital image analysis. Control glandular cells contained large lipid vacuoles that were bordered by multi-layered membranes. Rough-surfaced endoplasmic reticulum, mitochondria, and smooth-surfaced endoplasmic reticulum may be involved in lipid vacuole formation. Myoepithelial cells surrounded alveoli. After carbamylcholine (CCh, 10–6, 10–5, and 10–3 M) stimulation, lipid materials within the membranous structures were frequently discharged by an exocytotic mechanism. Conspicuous deformation of glandular cells caused by vigorous contraction of myoepithelial cells was observed in isolated alveoli after 10–6M CCh stimulation, whereas the deformaties of glandular tissues perfused via vessels were small even after 10–3M CCh stimulation. Connective tissue between glandular alveoli inhibited unbridled myoepithelial-cell contraction. Fura-2/AM digital imaging analysis revealed that CCh stimulation caused an increase in [Ca2+]c in isolated alveoli. The morphological reactions and changes in [Ca2+]c were prevented by atropine. When extracellular calcium ions were absent, enhanced extrusion of lipid vacuoles, myoepithelial-cell contraction, and a rise in [Ca2+]c after CCh stimulation were not observed. Nicotine and catecholamines had no effect on the secretion or on the dynamics of [Ca2+]c. It can be concluded that acetylcholine elicits exocytosis in glandular cells and contraction of the myoepithelial cells of Harderian glands, accompanied by an increase in [Ca2+]c. The dynamics of [Ca2+]c of the gland alveoli are mostly dependent on extracellular Ca2+. 相似文献
15.
A. Champigneulle E. Siga G. Vassent M. Imbert-Teboul 《The Journal of membrane biology》1997,156(2):117-129
The effect of extracellular calcium ([Ca2+]
e
) on cytosolic calcium ([Ca2+]
i
) was investigated in thick ascending limbs and collecting ducts from the rat kidney, using the fluorescent dye fura-2. In
cortical collecting ducts, basolateral but not apical changes in [Ca2+]
e
were associated with parallel changes in [Ca2+]
i
. Basal [Ca2+]
i
was hardly modified by nifedipine and verapamil but was decreased by 60% by basolateral La3+. Increasing peritubular [Ca2+]
e
triggered Ca2+ release from intracellular stores. This effect was not reproduced by agonists of the renal Ca2+-receptor RaKCaR, e.g., Ba2+, Mg2+, Gd3+, and neomycin, but was reproduced by Ni2+. Ni2+-induced mobilization of intracellular Ca2+ was larger in the inner medullary collecting duct, a segment which poorly responds to increasing [Ca2+]
e
.
In the cortical thick ascending limb, removing basolateral Ca2+ hardly altered [Ca2+]
i
but increasing [Ca2+]
e
or adding Ba2+, Mg2+, Gd3+ and neomycin released intracellular calcium.
These data demonstrate that (1) basolateral influx of calcium occurs in cortical collecting ducts, under basal conditions;
(2) this influx occurs through nonvoltage gated channels, permeable to Ba2+, insensitive to verapamil and nifedipine, and blocked by La3+; (3) increasing [Ca2+]
e
stimulates the influx and triggers intracellular calcium release, independently of the phospholipase C-coupled receptor RaKCaR;
(4) RaKCaR is functionally expressed in thick ascending limbs; (5) another membrane receptor, sensitive to Ni2+ but not to Ca2+ is present in the collecting duct.
Received: 12 July 1996/Revised: 28 October 1996 相似文献
16.
Mohammed A. Kader Sylvia Lindberg Thorsten Seidel Dortje Golldack Vladislav Yemelyanov 《Physiologia plantarum》2007,130(1):99-111
Perception of salt stress in plant cells induces a change in the free cytosolic Ca2+, [Ca2+]cyt, which transfers downstream reactions toward salt tolerance. Changes in cytosolic H+ concentration, [H+]cyt, are closely linked to the [Ca2+]cyt dynamics under various stress signals. In this study, salt‐induced changes in [Ca2+]cyt, and [H+]cyt and vacuolar [H+] concentrations were monitored in single protoplasts of rice (Oryza sativa L. indica cvs. Pokkali and BRRI Dhan29) by fluorescence microscopy. Changes in cytosolic [Ca2+] and [H+] were detected by use of the fluorescent dyes acetoxy methyl ester of calcium‐binding benzofuran and acetoxy methyl ester of 2′, 7′‐bis‐(2‐carboxyethyl)‐5‐(and‐6) carboxyfluorescein, respectively, and for vacuolar pH, fluorescent 6‐carboxyfluorescein and confocal microscopy were used. Addition of NaCl induced a higher increase in [Ca2+]cyt in the salt‐tolerant cv. Pokkali than in the salt‐sensitive cv. BRRI Dhan29. From inhibitor studies, we conclude that the internal stores appear to be the major source for [Ca2+]cyt increase in Pokkali, although the apoplast is more important in BRRI Dhan29. The [Ca2+]cyt measurements in rice also suggest that Na+ should be sensed inside the cytosol, before any increase in [Ca2+]cyt occurs. Moreover, our results with individual mesophyll protoplasts suggest that ionic stress causes an increase in [Ca2+]cyt and that osmotic stress sharply decreases [Ca2+]cyt in rice. The [pH]cyt was differently shifted in the two rice cultivars in response to salt stress and may be coupled to different activities of the H+‐ATPases. The changes in vacuolar pH were correlated with the expressional analysis of rice vacuolar H+‐ATPase in these two rice cultivars. 相似文献
17.
《Journal of receptor and signal transduction research》2013,33(2):89-95
AbstractClotrimazole is an antimycotic imidazole derivative that interferes with cellular Ca2+ homeostasis. This study examined the effect of clotrimazole on cytosolic Ca2+ concentrations ([Ca2+]i) and viability in HA59T human hepatoma cells. The Ca2+-sensitive fluorescent dye fura-2 was applied to measure [Ca2+]i. Clotrimazole induced [Ca2+]i rises in a concentration-dependent manner. The response was reduced by removing extracellular Ca2+. Clotrimazole-evoked Ca2+ entry was suppressed by store-operated channel inhibitors (nifedipine, econazole and SK&F96365) and protein kinase C modulators (GF109203X and phorbol, 12-myristate, 13-acetate). In Ca2+-free medium, incubation with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di-tert-butylhydroquinone abolished clotrimazole-induced [Ca2+]i rise. Inhibition of phospholipase C with U73122 abolished clotrimazole-induced [Ca2+]i rise. At 10–40?µM, clotrimazole inhibited cell viability, which was not reversed by chelating cytosolic Ca2+. Clotrimazole at 10 and 30?µM also induced apoptosis. Collectively, in HA59T cells, clotrimazole-induced [Ca2+]i rises by evoking phospholipase C-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via store-operated Ca2+ channels. Clotrimazole also caused apoptosis. 相似文献
18.
Cytoplasmic free calcium distributions during the development of root hairs of Arabidopsis thaliana 总被引:14,自引:8,他引:6
Carol L. Wymer Tatiana N. Bibikova Simon Gilroy 《The Plant journal : for cell and molecular biology》1997,12(2):427-439
In this study, confocal ratio analysis was used to image the relationship between cytoplasmic free calcium concentration ([Ca2+]c) and the development of root hairs of Arabidopsis thaliana. Although a localized change in [Ca2+]c that preceded or predicted the site of root hair initiation could not be detected, once initiated the majority of emerging root hairs showed an elevated [Ca2+]c (>1 μM) in their apical cytoplasm, compared with 100– 200 nM in the rest of the cell. These emerging root hairs then moved into a 3–5 h phase of sustained elongation during which they showed variable growth rates. Root hairs that were rapidly elongating exhibited a highly localized, elevated [Ca2+]c at the tip. Non-growing root hairs did not exhibit the [Ca2+]c gradient. The rhd-2 mutant, which is defective in sustained root hair growth, showed an altered [Ca2+]c distribution compared with wild-type. These results implicate [Ca2+]c in regulating the tip growth process. Treatment of elongating wild-type root hairs with the Ca2+ channel blocker verapamil (50 μM) caused dissipation of the elevated [Ca2+]c at the tip and cessation of growth, suggesting a requirement for Ca2+ channel activity at the root hair tip to maintain growth. Manganese treatment also preferentially quenched Indo-1 fluorescence in the apical cytoplasm of the root hair. As manganese is thought to enter cells through Ca2+-permeable channels, this result also suggests increased Ca2+ channel activity at the tip of the growing hair. Taken together, these data suggest that although Ca2+ does not trigger the initiation of root hairs, Ca2+ influx at the tip of the root hair leads to an elevated [Ca2+]c that may be required to sustain root hair elongation. 相似文献
19.
Background
Maitotoxin, a potent cytolytic agent, causes an increase in cytosolic free Ca2+ concentration ([Ca2+]i) via activation of Ca2+-permeable, non-selective cation channels (CaNSC). Channel activation is followed by formation of large endogenous pores that allow ethidium and propidium-based vital dyes to enter the cell. Although activation of these cytolytic/oncotic pores, or COP, precedes release of lactate dehydrogenase, an indication of oncotic cell death, the relationship between CaNSC, COP, membrane lysis, and the associated changes in cell morphology has not been clearly defined. In the present study, the effect maitotoxin on [Ca2+]i, vital dye uptake, lactate dehydrogenase release, and membrane blebbing was examined in bovine aortic endothelial cells.Results
Maitotoxin produced a concentration-dependent increase in [Ca2+]i followed by a biphasic uptake of ethidium. Comparison of ethidium (Mw 314 Da), YO-PRO-1 (Mw 375 Da), and POPO-3 (Mw 715 Da) showed that the rate of dye uptake during the first phase was inversely proportional to molecular weight, whereas the second phase appeared to be all-or-nothing. The second phase of dye uptake correlated in time with the release of lactate dehydrogenase. Uptake of vital dyes at the single cell level, determined by time-lapse videomicroscopy, was also biphasic. The first phase was associated with formation of small membrane blebs, whereas the second phase was associated with dramatic bleb dilation.Conclusions
These results suggest that maitotoxin-induced Ca2+ influx in bovine aortic endothelial cells is followed by activation of COP. COP formation is associated with controlled membrane blebbing which ultimately gives rise to uncontrolled bleb dilation, lactate dehydrogenase release, and oncotic cell death. 相似文献20.
Osmotin is a pathogenesis-related protein exhibiting cryoprotective functions. Our aim was to understand whether it is involved
in the cold acclimation of the olive tree (Olea europaea L.), a frost-sensitive species lacking dormancy. We exposed olive trees expressing tobacco osmotin gene under the 35S promoter (35S:osm) [in the same manner as wild type (wt) plants] to cold shocks in the presence/absence of cold acclimation, and monitored
changes in programmed cell death (PCD), cytoskeleton, and calcium ([Ca2+]c) signalling. In the wt, osmotin was immunolocalized only in cold-acclimated plants, and in the tissues showing PCD. In the
35S:osm clones, the protein was detected also in the non-acclimated plants, and always in the tissues exhibiting PCD. In the non-acclimated
wt protoplasts exposed to cold shock, a transient decrease in phallotoxin signal suggests a temporary disassembly of F-actin,
a transient increase occurred instead in 35S:osm protoplasts exposed to the same shock. Transient increases in [Ca2+]c were observed only in the wt protoplasts. However, when F-actin was depolymerized by cytochalasin or latrunculin, and microtubules
by colchicine, increase in [Ca2+]c also occurred in the 35S:osm protoplasts. Successive cold shocks caused transient rises in [Ca2+]c and transient decreases in the phallotoxin signal in wt protoplasts. No change occurred in [Ca2+]c occurred in the 35S:osm protoplasts. The phallotoxin signal transiently increased at the first shock, but did not change after the subsequent shocks,
and an overall signal reduction occurred with shock repetition. Following acclimation, no cold shock-induced change in [Ca2+]c levels and F-actin signal occurred either in wt or 35S:osm protoplasts. The results show that osmotin is positively involved in the acclimation-related PCD, in blocking the cold-induced
calcium signalling, and in affecting cytoskeleton in response to cold stimuli. 相似文献