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1.
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
2.
The effect of aminooxyacetic acid (AOAA), NH4
+, phenylsuccinate (Phs), ketone bodies (KB) and glutamine (Gln), that might interfere with the biosynthesis of neurotransmitter
glutamate on the K+-evoked Ca2+-dependent release ofd-[3H]aspartate from rat cerebellar slices was studied. Therefore slices were preincubated in a Krebs-Ringer-bicarbonate-glucose
(KR) buffer, loaded withd-[3H]aspartate and superfused in the presence of Ca2+ or when Ca2+ was replaced by Mg2+ or in some cases by EGTA. AOAA, NH
4
+
and Phs increase the K+-evoked Ca2+-dependent release of radioactivity by 30%, 68% and 188% compared to the control respectively indicating that these agents
are inhibitors of the K+-evoked Ca2+-dependent release of glutamate. KB and Gln had no effect on the Ca2+-dependent release of radioactivity. AOAA., NH
4
+
, Phs and KB but not Gln increase the total release of radioactivity by 43%, 69%, 139%, and 37% respectively. AOAA, NH
4
+
and KB but not Phs or Gln increase the Ca2+-independent release (Mg2+ replacing Ca2+) of radioactivity by 71%, 71% and 108% respectively. The present results indicate that in the cerebellum: 1) Neurotransmitter
glutamate is mostly synthesized through the phosphate activated glutaminase (PAG) reaction 2) It is further supported that
glutamate released in a Ca2+-dependent manner before entering its pool in the cytosol has to move into the mitochondrial matrix. 相似文献
3.
The release of [3H]purines was investigated in a crude mitochondrial fraction (P2 fraction) from rat brain cortex pre-loaded with [3H]adenosine for 30 sec at 37°C in vitro. Potassium, veratridine and glutamate were used as depolarizing agents to evoke the release of [3H]purines. Ca2+ removal, the addition of EGTA, and treatment with organic or inorganic Ca2+ antagonists did not inhibit [3H]purine release in this preparation. On the other hand, Ca2+ removal and the addition of EGTA greatly enhanced3H-purine release induced by glutamate. D-600 and diltiazem enhanced K+-evoked [3H]purine release, and nifedipine increased veratridine evoked [3H]purine release indicating that either these Ca2+ antagonists have different sites of action, or that K+ and veratridine may release [3H]purine from different metabolic pools. Organic Ca2+ antagonists failed to enhance the [3H]purine release evoked by glutamate, further supporting the notion that various depolarizing agents may release [3H]purines from different cellular compartments. 相似文献
4.
RELEASE OF ATP FROM A SYNAPTOSOMAL PREPARATION BY ELEVATED EXTRACELLULAR K+ AND BY VERATRIDINE 总被引:6,自引:0,他引:6
T. D. White 《Journal of neurochemistry》1978,30(2):329-336
A technique was developed which permitted the release of ATP from synaptosomes by elevated extracellular K+ or by veratridine to be directly and continuously monitored. The released ATP interacted with firefly luciferin and luciferase in the incubation medium to produce light which could be detected by a photomultiplier. The assay system was specific for ATP, in that similar concentrations of adenosine, AMP or ADP did not produce chemiluminescence. Moreover, the maximum peak of light emission correlated linearly with the concentrations of ATP present in the medium, so that semiquantitative estimates of ATP release could be made. Elevating the extracellular K+ concentration produced a graded release of ATP from synaptosomes. Rb+ also released ATP but Na+, Li+ and choline did not. The response to elevated K+ was not blocked by tetrodotoxin (TTX), indicating that this effect was not mediated by the opening of Na+-channels in synaptosomal membranes. Veratridine (50 μM) caused a graded release of ATP which was larger and more prolonged than that caused by elevated K+. The release of ATP by veratridine was blocked by TTX indicating that the opening of Na+-channels was involved. Neither veratridine nor elevated K+ released ATP from microsomal or mitochondrial fractions, showing that the release of ATP probably did not originate from microsomal, vesicular or mitochondrial contaminants of the synaptosomal preparation. Release of ATP by elevated K+ was diminished in a medium lacking CaCl+ or when EGTA was added to chelate Ca2+. In contrast, release by veratridine appeared to be augmented in Ca2+-free media or in the presence of EGTA. The K+-induced release of ATP, which is Ca2+ dependent, closely resembles the exocytotic release of putative neurotransmitters from presynaptic nerve-terminals. On the other hand, the apparent lack of a Ca2+ requirement for veratridine's action suggests that this process could originate from other sites, or involve mechanisms other than conventional neurotransmitter release processes. 相似文献
5.
Schwartz A 《Plant physiology》1985,79(4):1003-1005
Ca2+ (0.1-1.0 millimolar) accelerated dark-induced stomatal closure and reduced stomatal apertures in the light in epidermal peels of Commelina communis L. In contrast, ethyleneglycol-bis-(β-aminoethyl ether) N,N′tetraacetic acid (EGTA) (2 millimolar), a Ca2+ chelator, prevented closure in the dark and accelerated opening in the light. EGTA did not promote significant opening in the dark. It is therefore concluded that EGTA does not increase ion uptake into guard cells, but rather prevents ion efflux. Addition of EGTA to incubating solutions with 10 millimolar KCl resulted in steady state apertures of 15.6 micrometers, whereas in the absence of EGTA similar apertures required 55 millimolar KCl and 150 millimolar KCl was needed in the presence of 1 millimolar CaCl2. The results demonstrate the importance of Ca2+ in the regulation of stomatal closure and point to a role of Ca2+ in the regulation of K+ efflux from stomatal guard cells. 相似文献
6.
To characterize the effect of glutamine on the release of glutamate, aspartate, and γ-aminobutyric acid (GABA), rat hippocampal slices were superfused with different concentrations of glutamine or Ca2+. Amino acids released and retained were analyzed by HPLC. Glutamine (0.5 mmol/L) increased more than threefold the release of glutamate evoked by 50 mmol/L K+ in the presence of 2.6 mmol/L Ca2+ without a corresponding increase in glutamate content, while the release of aspartate was increased less and that of GABA not at all by glutamine. The evoked release of all three amino acids, including the enhanced release of glutamate in the presence of glutamine, was strongly dependent on Ca2+ concentrations between 0.1 and 2.6 mmol/L. The potentiation of glutamate release by glutamine reached a plateau at 0.25 mmol/L glutamine. Intermittent electrical field stimulation increased the release of only glutamate and this release was nearly doubled by glutamine. The increased release was Ca2+ dependent and tetrodotoxin (TTX) sensitive. Results suggest that extracellular glutamine promotes primarily the formation of releasable glutamate and this enhancement is dependent on extracellular Ca2+. 相似文献
7.
F R Butcher 《Life sciences》1979,24(21):1979-1982
Stimulation of 86Rb+ efflux from isolated parotid acinar cells by carbchol was biphasic. The phases of stimulated 86Rb+ efflux were separated on the basis of their relative requirements for extracellular Ca2+. If the isolated cells were incubated in Ca2+ free buffer containing 1.0 mM ethylene glycol bis (β-aminoethyl ether) N, N1 - tetra acetic acid (EGTA) for 30 min. before adding carbachol an initial phase of 86Rb+ efflux was observed. A second phase of 86Rb+ efflux was obtained upon addition of 2.0 mM Ca2+. However when cells were incubated for 60 min. in Ca2+ free buffer containing 1.0 mM EGTA the initial phase of release caused by carbachol was inhibited by 95 percent. If the EGTA was titrated with Ca2+ to give 1.0 mM Ca2+, following the 60 min. depletion regimen, the second phase was observed. Although 60 min. of Ca2+-depletion in EGTA buffer was required for complete inhibition of the effect of carbachol on the initial phase of 86Rb+ efflux, the response was fully restored within 4 min. after the readdition of Ca2+. 相似文献
8.
Summary The effects of the sulphonylurea activators of ATP-sensitive potassium channels (K+
ATP), cromakalim and pinacidil, on the evoked-release of endogenous glutamate from superfused slices of rat cerebellum was examined. K+-stimulated release was Ca2+-dependent, whereas tetrapentylammonium (TPeA)-evoked release occurred both in the presence and absence of Ca2+, but was significantly greater in Ca2+-free medium. The Ca2+-dependent TPeA and K+-evoked release of glutamate was inhibited by both cromakalim and pinacidil in a concentration-dependent fashion. However, although cromakalim markedly reduced Ca2+-independent TPeA-evoked release, pinacidil was ineffective. In addition, the vehicle for cromakalim, ethanol, markedly potentiated both Ca2+-dependent and -independent TPeA-evoked release, but not K+-evoked release. Despite a high concentration of sulphonylurea binding sites and a dense glutamatergic innervation, the concentrations of K+
ATP channel activators required to inhibit stimulus-evoked release from the cerebellum are higher than those reported to inhibit glutamate release or reduce neuronal activity in other parts of the CNS. 相似文献
9.
Acute stress is known to enhance the memory of events that are potentially threatening to the organisms. Glutamate, the most
abundant excitatory neurotransmitter in the mammalian central nervous system, plays a critical role in learning and memory
formation and calcium (Ca2+) plays an essential role in transmitter release from nerve terminals (synaptosomes). In the present study, we investigated
the effects of acute restraint stress on cytosolic free Ca2+ concentration ([Ca2+]i) and glutamate release in cerebrocortical synaptosomes from mice. Acute restraint stress caused a significant increase in
resting [Ca2+]i and significantly enhanced the ability of the depolarizing agents K+ and 4-aminopyridine (4-AP) to increase [Ca2+]i. It also brought about a significant increase in spontaneous (unstimulated) glutamate release and significantly enhanced
K+- and 4-AP-induced Ca2+-dependent glutamate release. The pretreatment of synaptosomes with a combination of ω-agatoxin IVA (a P-type Ca2+ channel blocker) and ω-conotoxin GVIA (an N-type Ca2+ channel blocker) completely suppressed the enhancements of [Ca2+]i and Ca2+-dependent glutamate release in acute restraint-stressed mice. These results indicate that acute restraint stress enhances
K+- or 4-AP-induced glutamate release by increasing [Ca2+]i via stimulation of Ca2+ entry through P- and N-type Ca2+ channels. 相似文献
10.
Richard J. H. Wojcikiewicz rew B. Tobin Stefan R. Nahorski 《Journal of neurochemistry》1994,63(1):177-185
Abstract: Stimulation of muscarinic receptors expressed in SH-SY5Y human neuroblastoma cells resulted in a complex profile of inositol 1,4,5-trisphosphate (InsP3) accumulation, with a dramatic increase (six- to eightfold) over the first 10 s (the “peak” phase) and subsequently, from ~60 s onward, maintained at a lower but sustained level (the “plateau” phase). Chelation of extracellular Ca2+ with EGTA or inhibition of Ca2+ channels with Ni2+ showed that the plateau phase was dependent upon Ca2+ entry. Furthermore, use of thapsigargin and EGTA to discharge and sequester Ca2+ from intracellular stores revealed that Ca2+ from this source was capable of supporting the peak phase of the InsP3 response. Carbachol-stimulated phosphoinositidase C activity in permeabilized SH-SY5Y cells was also shown to be highly dependent on free Ca2+ concentration (20–100 nM) and suggests that under normal conditions, InsP3 formation is enhanced by increases in cytosolic free Ca2+ concentration that accompany muscarinic receptor activation. Measurement of carbachol-stimulated total inositol phosphate accumulation in the presence of Li+ indicated that the initial rate of phosphoinositide hydrolysis (from 0 to 30 s) was about fivefold greater than that from 30 to 300 s. This rapid but partial desensitization of receptor-mediated phosphoinositide hydrolysis provides strong evidence for the mechanism underlying the changes in InsP3 accumulation over this time. Because very similar data were obtained in Chinese hamster ovary cells transfected with human m3 receptor cDNA, we suggest that although increases in cytosolic free Ca2+ concentration amplify InsP3 formation during stimulation of m3 muscarinic receptors, the primary factor that governs the profile of InsP3 accumulation is rapid, but partial, desensitization. Such desensitization does not appear to be mediated by changes in cytosolic Ca2+ or protein kinase C activity. 相似文献
11.
T. Takeo S. Suga J. Wu Y. Dobashi T. Kanno M. Wakui 《Journal of cellular physiology》1998,174(3):387-397
Quantitative time-resolved measurements of cytosolic Ca2+ release by photolysis of caged InsP3 have been made in single rat submandibular cells using patch clamp whole-cell recording to measure the Ca2+-activated Cl− and K+ currents. Photolytic release of InsP3 from caged InsP3 at 100 Joules caused transient inward (VH = 60 mV) and outward (VH = 0 mV) currents, which were nearly symmetric in their time course. The inward current was reduced when pipette Cl− concentration was decreased, and the outward current was suppressed by K+ channel blockers, indicating that they were carried by Cl− and K+, respectively. Intracellular pre-loading of the InsP3 receptor antagonist heparin or the Ca2+ chelator EGTA clearly prevented both inward and outward currents, indicating that activation of Ca2+-dependent Cl− and K+ currents underlies the inward and the outward currents. At low flash intensities, InsP3 caused Ca2+ release which normally activated the K+ and Cl− currents in a mono-transient manner. At higher intensities, however, InsP3 induced an additional delayed outward K+ current (IK(delay)). IK(delay) was independent of the initial K+ current, independent of extracellular Ca2+, inhibited by TEA, and gradually prolongated by repeated flashes. The photolytic release of Ca2+ from caged Ca2+ did not mimic the IK(delay). It is suggested that Ca2+ releases from the InsP3-sensitive pools in an InsP3 concentration-dependent manner. Low concentrations of InsP3 induce the transient Ca2+-dependent Cl− and K+ currents, which reflects the local Ca2+ release, whereas high concentrations of InsP3 induce a delayed Ca2+-dependent K+ current, which may reflect the Ca2+ wave propagation. J. Cell. Physiol. 174:387–397, 1998. © 1998 Wiley-Liss, Inc. 相似文献
12.
Danica Stanimirovic Paul Morley Rita Ball Edith Hamel Geoff Mealing Jon P. Durkin 《Journal of cellular physiology》1996,169(3):455-467
The involvement of the early signaling messengers, inositol tris-phosphate (IP3), intracellular calcium, [Ca2+]i, and protein kinase C (PKC), in angiotensin II (AII)-induced fluid phase endocytosis was investigated in human brain capillary and microvascular endothelial cells (HCEC). AII (0.01–10 μM) stimulated the uptake of Lucifer yellow CH, an inert dye used as a marker for fluid phase endocytosis, in HCEC by 50–230%. AII also triggered a fast accumulation of IP3 and a rapid increase in [Ca2+]i in cells loaded with the Ca2+-responsive fluorescent dye fura-2. The prompt AII-induced [Ca2+]i spike was not affected by incubating HCEC in Ca2+-free medium containing 2 mM EGTA or by pretreating the cultures with the Ca2+ channel blockers, methoxyverapamil (D600; 50 μM), nickel (1 mM), or lanthanum (1 mM), suggesting that the activation of AII receptors on HCEC triggers the release of Ca2+ from intracellular stores. The AII-triggered increases in IP3, [Ca2+]i, and Lucifer yellow uptake were inhibited by the nonselective AII receptor antagonist, Sar1, Val5, Ala8-AII (SVA-AII), and by the phospholipase C (PLC) inhibitors, neomycin and U-73122. By contrast, the protein kinase C (PKC) inhibitors, staurosporine and calphostin C, failed to affect any of these AII-induced events. This study demonstrates that increased fluid phase endocytotosis induced by AII in human brain capillary endothelium, an event thought to be linked to the observed increases in blood-brain barrier permeability in acute hypertension, is likely dependent on PLC-mediated changes in [Ca2+]i and independent of PKC. © 1996 Wiley-Liss, Inc. 相似文献
13.
Elena Bossi Elena Centinaio Andrea Moriondo Antonio Peres 《Journal of cellular physiology》1998,174(2):154-159
The role of Ca2+ on the depolarization-induced appearance of a Na+ current in Xenopus oocytes was studied. Oocytes were voltage-clamped and the induction of the Na+ current was tested under various conditions. In oocytes pre-injected with 400 pmol EGTA to increase the intracellular Ca2+ buffering power, the current was significantly reduced. Conversely, when intracellular Ca2+ was made to increase by injecting an analogue of inositol 1,4,5-trisphosphate (3-F InsP3), to cause Ca2+ release from internal stores, the induction of the Na+ current was potentiated. The depolarization-inducible Na+ channels of the Xenopus oocyte membrane appear, therefore, to be Ca2+ sensitive, as well as depolarization-activated. J. Cell. Physiol. 174:154–159, 1998. © 1998 Wiley-Liss, Inc. 相似文献
14.
Dagmar Siliprandi A. Toninello F. Zoccarato Michela Rugolo N. iliprandi 《Journal of bioenergetics and biomembranes》1978,10(1-2):1-11
Addition to rat liver mitochondria of 2 mM inorganic phosphate or 0.15 mM diamide, a thiol-oxidizing agent, induced an efflux of endogenous Mg2+ linear with time and dependent on coupled respiration. No net Ca2+ release occurred under these conditions, while a concomitant release of K+ was observed. Mg2+ efflux mediated either by Pi or low concentrations of diamide was completely prevented by EGTA, Ruthenium red, and NEM. These reagents also inhibited the increased rate of state 4 respiration induced both by Pi and diamide. At higher concentrations (0.4 mM), diamide induced an efflux of Mg2+ which was associated also with a release of endogenous Ca2+. Under these conditions EGTA completely prevented Mg2+ and K+ effluxes, while they were only partially inhibited by Ruthenium red and NEM. It is assumed that Mg2+ efflux, occurring at low diamide concentrations or in the presence of phosphate, is dependent on a cyclic in-and-out movement of Ca2+ across the inner mitochondrial membrane, in which the passive efflux is compensated by a continuous energy linked reuptake. This explains the dependence of Mg2+ efflux on coupled respiration, as well as the increased rate of state 4 respiration. The dependence of Mg2+ efflux on phosphate transport is explained by the phosphate requirement for Ca2+ movement.Abbreviations Diamide
diazenedicarboxylic acidbis-dimethylamide
- FCCP
p-trifluoromethoxyphenylhydrazone
- EGTA
ethylene glycol-bis-(2-amino ethyl ether)-N,N-tetracetic acid
- Pi
inorganic phosphate
- Ruthenium red
Ru2(OH)2Cl4 · 7NH3 · 3H2O
- state 4
controlled state of respiration in the presence of substrate
- RCI
respiratory control index
- NEM
N-ethyl maleimide
A partial and preliminary report of these results has been published inBiochem. Biophys. Res. Comm.,78 (1977) 23. 相似文献
15.
Ádám Fekete Laura Franklin Takeshi Ikemoto Balázs Rózsa Balázs Lendvai E. Sylvester Vizi Tibor Zelles 《Journal of neurochemistry》2009,111(3):745-756
Although the role of Na+ in several aspects of Ca2+ regulation has already been shown, the exact mechanism of intracellular Ca2+ concentration ([Ca2+]i) increase resulting from an enhancement in the persistent, non‐inactivating Na+ current (INa,P), a decisive factor in certain forms of epilepsy, has yet to be resolved. Persistent Na+ current, evoked by veratridine, induced bursts of action potentials and sustained membrane depolarization with monophasic intracellular Na+ concentration ([Na+]i) and biphasic [Ca2+]i increase in CA1 pyramidal cells in acute hippocampal slices. The Ca2+ response was tetrodotoxin‐ and extracellular Ca2+‐dependent and ionotropic glutamate receptor‐independent. The first phase of [Ca2+]i rise was the net result of Ca2+ influx through voltage‐gated Ca2+ channels and mitochondrial Ca2+ sequestration. The robust second phase in addition involved reverse operation of the Na+–Ca2+ exchanger and mitochondrial Ca2+ release. We excluded contribution of the endoplasmic reticulum. These results demonstrate a complex interaction between persistent, non‐inactivating Na+ current and [Ca2+]i regulation in CA1 pyramidal cells. The described cellular mechanisms are most likely part of the pathomechanism of certain forms of epilepsy that are associated with INa,P. Describing the magnitude, temporal pattern and sources of Ca2+ increase induced by INa,P may provide novel targets for antiepileptic drug therapy. 相似文献
16.
Persistent Enhancement of Sustained Calcium-Dependent Glutamate Release by Phorbol Esters: Requirement for Localized Calcium Entry 总被引:3,自引:0,他引:3
David M. Terrian 《Journal of neurochemistry》1995,64(1):172-180
Abstract: Sustained activation of protein kinase C significantly enhanced a secondary (slow) phase in the depolarization-induced release of glutamate from isolated hippocampal nerve endings. The phorbol ester, 4β-phorbol 12,13-dibutyrate, was used to sustain the activation of presynaptic protein kinase C for a prolonged (10-min) period, and then this relatively water-soluble phorbol ester was removed by superfusion before a 2-min stimulus of continuous membrane depolarization. These conditions were used to investigate the persistent effects of sustained protein kinase C activation on the magnitude of the slow phase of evoked glutamate release, in which the efficiency of synaptic vesicle mobilization and recycling may be primary determinants of response magnitude. It is reported here that sustained protein kinase C activation selectively increased the Ca2+-dependent component of glutamate release during a prolonged phase of K+-induced depolarization. The magnitude of this persistent effect on Ca2+-dependent glutamate release was directly related to the dose of 4β-phorbol 12,13-dibutyrate and the duration of exposure that was used to prime the release apparatus, was observed using two alternative synaptosomal preparations, and was evident regardless of the depolarizing stimulus used (elevated [KCl] or 4-aminopyridine). However, 4β-phorbol 12,13-dibutyrate did not alter the release induced by the Ca2+ ionophore ionomycin. Thus, the persistent effects of protein kinase C activation on a prolonged phase of glutamate release were dependent on the route of Ca2+ influx. The finding that voltage-regulated Ca2+ channel blockers were able to neutralize completely the 4β-phorbol 12,13-dibutyrate-dependent facilitation of K+-evoked glutamate release provided further support for this conclusion. Thus, 4β-phorbol 12,13-dibutyrate significantly potentiated the sustained release of glutamate without altering the strict requirement that is normally displayed by synaptosomes for localized and voltage-regulated Ca2+ entry. 相似文献
17.
Craig S. Malcolm Lyndsay Ritchie Angus Grieve Roger Griffiths 《Journal of neurochemistry》1996,66(6):2350-2360
Abstract: The effect(s) of a prototypic intracellular Ca2+ antagonist, 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8), on glutamate-induced neurotoxicity was investigated in primary cultures of mouse cerebellar granule cells. Glutamate evoked an increase in cytosolic free-Ca2+ levels ([Ca2+]i) that was dependent on the extracellular concentration of Ca2+ ([Ca2+]o). In addition, this increase in [Ca2+]i correlated with a decrease in cell viability that was also dependent on [Ca2+]o. Glutamate-induced toxicity, quantified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) staining, was shown to comprise two distinct components, an “early” Na+/Cl?-dependent component observed within minutes of glutamate exposure, and a “delayed” Ca2+-dependent component (ED50~50 µM) that coincided with progressive degeneration of granule cells 4–24 h after a brief (5–15 min) exposure to 100 µM glutamate. Quantitative analysis of cell viability and morphological observations identify a “window” in which TMB-8 (at >100 µM) protects granule cells from the Ca2+-dependent, but not the Na+/Cl?-dependent, component of glutamate-induced neurotoxic damage, and furthermore, where TMB-8 inhibits glutamate-evoked increases in [Ca2+]i. These findings suggest that Ca2+ release from a TMB-8-sensitive intracellular store may be a necessary step in the onset of glutamate-induced excitotoxicity in granule cells. However, these conclusions are compromised by additional observations that show that TMB-8 (1) exhibits intrinsic toxicity and (2) is able to reverse its initial inhibitory action on glutamate-evoked increases in [Ca2+]i and subsequently effect a pronounced time-dependent potentiation of glutamate responses. Dantrolene, another putative intracellular Ca2+ antagonist, was completely without effect in this system with regard to both glutamate-evoked increases in [Ca2+]i and glutamate-induced neurotoxicity. 相似文献
18.
Abstract: Bovine chromaffin secretory vesicle ghosts loaded with Na+ were found to take up Ca2+ when incubated in K+ media or in sucrose media containing micromolar concentrations of free Ca2+. Li+- or choline+loaded ghosts did not take up Ca2+. The Ca2+ accumulated by Na+-loaded ghosts could be released by the Ca2+ ionophore A23187, but not by EGTA. Ca2+ uptake was inhibited by external Sr2+, Na +, Li +, or choline +. All the 45Ca2+ accumulated by Na+-dependent Ca2+ uptake could be released by external Na +, indicating that both Ca2+ influx and efflux occur in a Na+-dependent manner. Na + -dependent Ca2+ uptake and release were only slightly inhibited by Mg2+. In the presence of the Na+ ionophore Monensin the Ca2+ uptake by Na +-loaded ghosts was reduced. Ca2+ sequestered by the Na+-dependent mechanism could also be released by external Ca2+ or Sr2+ but not by Mg2+, indicating the presence of a Ca2+/Ca2+ exchange activity in secretory membrane vesicles. This Ca2+/Ca2+ exchange system is inhibited by Mg2+, but not by Sr2+. The Na + -dependent Ca2+ uptake system in the presence of Mg2+ is a saturable process with an apparent Km of 0.28 μM and a Vmax= 14.5 nmol min?1 mg protein?1. Ruthenium red inhibited neither the Na+/Ca2+ nor the Ca2+/Ca2+ exchange, even at high concentrations. 相似文献
19.
To study the effect of agents interfering with the biosynthesis and/or the K+-evoked Ca2+-dependent release of neurotransmitter glutamate, rat cerebral slices were preincubated with Krebs-Ringer-HEPES-glucose-glutamine buffer (KRH buffer), loaded withd-[3H]aspartate and superfused with the preincubation medium in the presence or in the absence of Ca2+. The difference in radioactivity release divided by the basal release per min under the two conditions represented the K+-evoked Ca2+-dependent release. The agents used were: 1) Aminooxyacetic acid (AOAA), the inhibitor of transaminases, 2) Leucine (Leu), the inhibitor of phosphate activated glutaminase (PAG), 3) NH4
+, the inhibitor of PAG, 4) Phenylsuccinic acid (Phs), the inhibitor of the mitochondrial ketodicarboxylate carrier, 5) ketone bodies, the inhibitors of glycolysis, 6) the absence of glutamine, the substrate of PAG. The results show that Leu, NH4
+, Phs and the absence of Gln significantly increase the K+-evoked Ca2+-dependent release of radioactivity by 64%, 200%, 95% and 147% respectively, indicating that these agents are inhibitors of the K+-evoked Ca2+-dependent release of glutamate. Ketone bodies and AOAA had no effect. These results indicate that the major if not the exclusive biosynthetic pathway of neurotransmitter glutamate in rat cerebral cortex is through the PAG reaction and support a model for the pathway followed by neurotransmitter glutamate i.e. glutamate formed outside the inner mitochondrial membrane has to enter the mitochondrial matrix or is formed within it from where it can be extruded to supply the transmitter pool in exchange of GABA. 相似文献
20.
Summary Microsomal fractions were isolated from gastric antrum and fundus smooth muscle of guinea pigs. Ca2+ uptake into and Ca2+ release from the membrane vesicles were studied by a rapid filtration method, and Ca2+ transport properties of the different regions of the stomach were compared. ATP-dependent Ca2+ uptake was similar in microsomes isolated from both regions. This uptake was increased by oxalate and was not affected by NaN3. Oxalate affected Ca2+ permeability of both antrum and fundus microsome vesicles similarly. Fundus microsome vesicles preincubated in 100mm NaCl and then diluted to 1/20 concentration with Na+-free medium had significantly higher ATP-independent Ca2+ uptake than vesicles preincubated in 100mm KCl and treated the same way. This was not true for antrum vesicles. Monensin abolished Na+-dependent Ca2+ uptake, and NaCl enhanced Ca2+ efflux from fundus microsome vesicles. The halflife values of Ca2+ loss from fundus vesicles in the presence of NaCl were significantly smaller than those in the presence of KCl. The release of Ca2+ from the vesicles within the first 3 min was accelerated by NaCl to three times that by KCl. However, NaCl had ro effect on Ca2+ release from antrum microsome vesicles.Results suggest two distinct mechanisms of stomach membrane Ca2+ transport: (1) ATP-dependent Ca2+ uptake and (2) Na+–Ca2+ exchange; the latter in the fundus only. 相似文献