首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
The association of [125I-]calmodulin with rat brain synaptosomal plasma membranes, when incubated for 1 h at 25° in the presence or in absence of 20 M Ca2+, follows a sigmoid path with a Hill coefficient h=1.79±0.12 and h=1.72±0.11, respectively. The total association of calmodulin with the membrane increased approx. 60%–80% at all the range of calmodulin concentrations used in the presence of 20 M Ca2+. A three fold increase of guanylate cyclase activity was shown in the presence of low concentrations of calmodulin (up to 10 mM); higher concentrations (up to 40 mM) however, led to a progressive inhibition of the enzyme activity with respect to maximal stimulation. Calmodulin increased the lipid fluidity of synaptosomal plasma membranes labeled with 1,6-diphenyl-1,3,5-hexatriene (DPH), as indicated by the steady-state fluorescence anisotropy [(ro/r)-1]–1. Arrhenius-type plots of [(ro/r)-1]–1 indicated that the lipid separation of the membrane at 22.7±1.2° was perturbed by calmodulin such that the temperature was reduced to 16.3±0.9° and 15.5±0.8° in the absence or in the presence of 20 M Ca2+. Arrhenius plots of guanylate cyclase and acetylcholinesterase activities exhibited brak points at 25.7±1.4° and 22.3±1.0° in control synaptosomal plasma membranes, respectively. The break point for the guanylate cyclase was reduced to 16.3±0.9° in calmodulin treated synaptosomal plasma membranes whereas that of acetylcholinesterase remained unaffected (21.1±0.9°). The allosteric properties of guanylate cyclase by Mn-GTP (as reflected by changes in the Hill coefficient) were modulated by calmodulin while those of acetylcholinesterase by fluoride (F) were not altered. We propose that calmodulin achieves these effects through asymmetric perturbations of the membrane lipid structure and that increase in membrane fluidity of the inner leaflet of the membrane induced by calmodulin may be an early key event to the process of neurotransmitter release.  相似文献   

2.
Marian  M.  Bindoli  A.  Callegarin  F.  Rigobello  M. P.  Vincenti  E.  Bragadin  M.  Scutari  G. 《Neurochemical research》1999,24(7):875-881
The effect of 2,6-diisopropylphenol (propofol) in comparison to that of the halogenated anesthetics enflurane, isoflurane, and halothane on tetrapenylphosphonium uptake by rat brain synaptosomes was studied. A direct method to separately measure the synaptosomal and the mitochondrial transmembrane potential by using the tetraphenylphosphonium cation (TPP+) was utilized. The latter is a lipophylic charged molecule which distributes between two compartments according to the transmembrane electrical potential in the presence or absence of 60 mM KCl as a synaptosomal membrane depolarizing agent. After previously reporting the damages induced by general anesthetics on isolated mitochondria, the aim of this paper was to study their possible action on the synaptosomal membrane potential and whether or not drugs concentrations damaging isolated mitochondria are also effective on synaptosomal mitochondria. The results indicated that, in the presence of glucose, mitochondria included in synaptosomes were able to maintain a transmembrane potential of 202 ± 8 mV (mean ± SD) while the synaptosomal membrane showed a potential of 78 ± 8 mV (mean ± SD). When anesthetic concentrations (0.6–1 mM propofol, 10–40 M enflurane, 30–50 M isoflurane, 8–15 M halothane) that impair mitochondrial energy metabolism were used, the synaptosomal transmembrane potential was maintained and, in addition, a slight increase of the TPP+ taken up was observed as the anesthetic concentration was increased.  相似文献   

3.
Expcsure of adult male albino rats to higher environmental temperature (HET) at 35° for 2–12 hr or at 45° for 1–2 hr increases hypothalamic synaptosomal acetylcholinesterase (AChE) activity. Synaptosomal AChE activity in cerebral cortex of rats exposed to 35° for 12 hr and in cerebral cortex and pons-medulla of rats exposed to 45° for 1–2 hr are also activated. AChE activity of synaptosomes prepared from normal rat brain regions incubated in-vitro at 39° or 41° for 0.5 hr increases significantly in cerebral cortex and hypothalamus. The activation of AChE in ponsmedulla is also observed when this brain region is incubated at 41° for 0.5 hr. Increase of (a) the duration of incubation at 41° and (b) the incubation temperature to 43° under in-vitro condition decreases the synaptosomal AChE activity. Lioneweaver-Burk plots indicate that (a) in-vivo and invitro HET-induced increases of brain regional synaptosomal AChE activity are coupled with an increase ofV max without any change inK m (b) very high temperature (43° under in-vitro condition) causes a decrease inV max with an increase inK m of AChE activity irrespective of brain regions. Arrhenius plots show that there is a decrease in transition temperature in hypothalamus of rats exposed to either 35° or 45°; whereas such a decrease in transition temperature of the pons-medulla and cerebral cortex regions are observed only after exposure to 45°. These results suggests that heat exposure increases the lipid fluidity of synaptosomal membrane depending on the brain region which may expose the catalytic site of the enzyme (AChE) and hence activate the synaptosomal membrane bound AChE activity in brain regions. Further the in-vitro higher temperature (43°C)-induced inhibition of synaptosomal AChE activity irrespective of brain regions may be the cause iof partial proteolysis/disaggregation of AChE oligomers and/or solubilization of this membrane-bound enzyme.To whom to address reprint requests:  相似文献   

4.
The use of 3–nitropropionic acid (3–NP) and other mitochondria inhibitors to effectuate animal models of Huntington's disease has been well established. 3–NP administration has been shown to lead to pathology similar to that of HD, including massive loss of striatal neurons associated with oxidative stress. Oxidative stress induced by 3–NP also extends to the cortex, an area where little neuron loss occurs. No mechanism as of yet accounts for selective loss of striatal neurons while sparing cortical neurons. In the present study, a nitroxide stearate lipid bilayer-specific spin-label was utilized to probe 3–NP-induced fluidity changes in striatal and cortical synaptosomal membranes. In cortical synaptosomes, membrane fluidity increased in animals previously treated with 3–NP when compared to controls injected with saline vehicle, while in striatal synaptosomes, membrane fluidity decreased in animals treated with 3–NP when compared to controls. The results of the present study suggest that oxidatively-induced changes in membrane fluidity may be involved in mechanisms by which selective striatal neuronal loss occurs in this animal model of Huntington's disease.  相似文献   

5.
2,6-diisopropylphenol (propofol), a general intravenous anesthetic, inhibits the glutamate-dependent Ca2+ entry in rat synaptosomes with an approximate IC50 of 3.0×10–5 M.Propofol, at concentrations above 10–6M, also inhibits the ATP-dependent uptake of glutamate in the presence of Ca2+, with an approximate IC50 of 3.5×10–5M, while it only has a slight inhibitory effect on the release of glutamate. The ouabain-insensitive synaptosomal ATPase is strongly inhibited by propofol, with an IC50 of about 2.5×10–6M, at concentrations which do not affect the luciferase system.  相似文献   

6.
In the present study we demonstrated that synaptosomes isolated from rabbit brain cortex contain NO synthase and xanthine oxidase that can be activated by ultraviolet B radiation and Ca2+ accumulation to produce nitric oxide and superoxide which react together to form peroxynitrite. Irradiation of synaptosomes with ultraviolet B (up to 100 mJ/cm2), or increase the intrasynaptosomal calcium concentration using various doses (up to 100 μM) of the calcium ionophore A 23187, a gradual increase in both nitric oxide and peroxynitrite release that was inhibited by N-monomethyl-L-arginine (100 μM) was observed. The rate of nitric oxide release and cyclic GMP production by NO synthase and soluble guanylate cyclase, both located in the soluble fraction of synaptosomes (synaptosol), were increased approximately eight fold after treatment of synaptosomes with Ultraviolet B radiation (100 mJ/cm2). In reconstitution experiments, when purified NO synthase isolated from synaptosol was added to xanthine oxidase, in the presence of the appropriate cofactors and substrates, a ten fold increase in peroxynitrite production at various doses (up to 20 mJ/cm2) of UVB radiation was observed. Ultraviolet B irradiated synaptosomes promptly increased malondialdehyde production with subsequent decrease of synaptosomal plasma membrane fluidity estimated by fluorescence anisotropy of 1-4-(trimethyl-amino-phenyl)-6-phenyl-hexa-1,3,5-triene. Desferrioxamine (100 μM) tested in Ultraviolet B-irradiated synaptosomes showed a decrease (approximately 80%) in malondialdehyde production with subsequent restoration of the membrane fluidity to that of non-irradiated (control) synaptosomes. Ca2+-stimulated ATPase activity was decreased after Ultraviolet B (100 mJ/cm2) radiation of synaptosomes indicating that the subsequent increase of intrasynaptosomal calcium promoted peroxynitrite production by a calmodulin-dependent increase of NO synthase and xanthine oxidase activities. Furthermore, it was shown that UVB-irradiated synaptosomes were subjected to higher oxidative stress by exogenous peroxynitrite (100 μM) compared to non-irradiated (control) synaptosomes. In summary, the present results indicate that activation of NO synthase and xanthine oxidase of brain cells lead to the formation of peroxynitrite providing important clues in the role of peroxynitrite as a causative factor in neurotoxicity.  相似文献   

7.
The antioxidant properties of 5-aminosalicylic acid in vitro were evaluated in a synaptosomal membrane system prepared from gerbil cortical synaptosomes using EPR spin labeling and spectroscopic techniques. MAL-6 (2,2,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl) and 5-NS (5-nitroxide stearate) spin labels were used to assess changes in protein oxidation and membrane lipid fluidity, respectively. Synaptosomal membranes were subjected to oxidative stress by incubation with 1 mM azo-bis(isobutyronitrile) (AIBN) or 1 mM 2,2-azobis(amidino propane) dihydrochloride (AAPH) at 37°C for 30 minutes. The EPR analyses of the samples showed significant oxidation of synaptosomal proteins and a decrease in membrane fluidity. 5-Aminosalicylic acid also was evaluated by means of FRAP (the ferric reducing ability of plasma) test as a potential antioxidant. 5-Aminosalicylic acid also showed protection against the oxidation in gerbil cortical synaptosomes system caused by AIBN and AAPH. These results are consistent with the notion of antioxidant protection against free radical induced oxidative stress in synaptosomal membrane system by this agent.  相似文献   

8.
Cholesterol-rich, liquid-ordered (Lo) domains are believed to be biologically relevant, and yet detailed knowledge about them, especially in live cells under physiological conditions, is elusive. Although these domains have been observed in model membranes, understanding cholesterol-lipid interactions at the molecular level, under controlled lipid mixing, remains a challenge. Further, although there are a number of fluorescent lipid analogs that partition into liquid-disordered (Ld) domains, the number of such analogs with a high affinity for biologically relevant Lo domains is limited. Here, we use a new Bodipy-labeled cholesterol (Bdp-Chol) derivative to investigate membrane fluidity, lipid order, and partitioning in various lipid phases in giant unilamellar vesicles (GUVs) as a model system. GUVs were prepared from mixtures of various molar fractions of dioleoylphosphatidylcholine, cholesterol, and egg sphingomyelin. The Ld phase domains were also labeled with 1,1′-didodecyl-3,3,3′,3′-tetramethylindocarbocyanine (DiI-C12) for comparison. Two-photon fluorescence lifetime and anisotropy imaging of Bdp-Chol are sensitive to lipid phase domains in GUVs. The fluorescence lifetime of Bdp-Chol in liquid-disordered, single-phase GUVs is 5.50 ± 0.08 ns, compared with 4.1 ± 0.4 ns in the presence of DiI-C12. The observed reduction of fluorescence lifetime is attributed to Förster resonance energy transfer between Bdp-Chol (a donor) and DiI-C12 (an acceptor) with an estimated efficiency of 0.25 and donor-acceptor distance of 2.6 ± 0.2 nm. These results also indicate preferential partitioning (Kp = 1.88) of Bdp-Chol into the Lo phase. One-photon, time-resolved fluorescence anisotropy of Bdp-Chol decays as a triexponential in the lipid bilayer with an average rotational diffusion coefficient, lipid order parameter, and membrane fluidity that are sensitive to phase domains. The translational diffusion coefficient of Bdp-Chol, as measured using fluorescence correlation spectroscopy, is (7.4 ± 0.3) × 10−8 cm2/s and (5.0 ± 0.2) × 10−8 cm2/s in the Ld and Lo phases, respectively. Experimental translational/rotational diffusion coefficient ratios are compared with theoretical predictions using the hydrodynamic model (Saffman-Delbrück). The results suggest that Bdp-Chol is likely to form a complex with other lipid molecules during its macroscopic diffusion in GUV lipid bilayers at room temperature. Our integrated, multiscale results demonstrate the potential of this cholesterol analog for studying lipid-lipid interactions, lipid order, and membrane fluidity of biologically relevant Lo domains.  相似文献   

9.
Synaptosomes were isolated from cerebrums of rats fed standard (20% protein) or protein-free diets for 30 days. Arrhenius plots of their (Na+/K+)ATPase activities revealed a transition temperature of 25.5°C for control rats and 23.4°C for rats on protein-free diet, indicating that the latter increases synaptosomal membrane fluidity. The only change observed in the composition of the synaptosomal membranes was a 26% decrease of sialic acid. In synaptosomes from rats on protein-free diet the uptake of tyrosine was slightly reduced while that of glutamate was not affected. However, the exit of glutamate was reduced.  相似文献   

10.
Compartmentation and flux characteristics of nitrate in spruce   总被引:8,自引:0,他引:8  
The radiotracer13N was used to undertake compartmental analyses for NO 3 in intact non-mycorrhizal roots ofPicea glauca (Moench) Voss. seedlings. Three compartments were defined, with half-lives of exchange of 2.5 s, 20 s, and 7 min. These were identified as representing surface adsorption, apparent free space, and cytoplasm, respectively. Influx, efflux, and net flux as well as cytoplasmic and apparent-free-space nitrate concentrations were estimated for three different concentration regimes of external nitrate. After exposure to external NO 3 for 3 d, influx was calculated to be 0.09 mol·g–1·h–1 (at 10 M [NO 3 ]o), 0.5mol·g–1·h–1 (at 100 M [NO inf3 sup– ]o), and 1.2 mol · g–1· h–1 (at 1.5 mM [NO 3 ]o). Efflux increased with increasing [NO 3 ]o, constituting 4% of influx at 10 M, 6% at 100 M, and 21% at 1.5 mM. Cytoplasmic [NO 3 ] was estimated to be 0.3 mM at 10 uM [NO 3 ]o, 2mM at 100 M [NO 3 ]o, and 4mM at 1.5 mM [NO 3 ]o, while free-space [NO 3 ] was 16 M, 173 M, and 2.2 mM, respectively. A series of experiments was carried out to confirm the identity of the compartments resolved by efflux analysis. Pretreatment at high temperature or application of 2-chloro-ethanol, sodium dodecyl sulphate or hydrogen peroxide made it possible to distinguish the metabolic (cytoplasmic) phase from the remaining two (physical) phases. Likewise, varying [Pi] of the medium altered efflux and thereby [NO 3 ]cyt, but did not affect [NO 3 ]free space.Abbreviations and Symbols [NO 3 ]cyt cytoplasmic NO 3 concentration - [NO 3 ]free space apparent-free-space NO 3 concentration - [NO 3 ]o concentration of NO 3 in the external solution - NO 3 flux - co efflux from the cytoplasm - oc influx to the cytoplasm - net net flux - xylem flux to the xylem - red/vac combined flux to reduction and the vacuole The research was supported by a Natural Sciences and Engineering Research Council, Canada, grant to Dr. A.D.M. Glass and by a University of British Columbia Graduate Fellowship to Herbert J. Kronzucker. Our thanks go to Dr. M. Adam and Mr. P. Culbert at the particle accelerator facility TRIUMF on the University of British Columbia Campus for providing13NO 3 , Drs. R.D. Guy and S. Silim for providing plant material, and Dr. M.Y. Wang, Mr. J. Mehroke and Mr. P. Poon for assistance in experiments and for helpful discussions.  相似文献   

11.
The aim of this work was to examine the effects of changes in external K+ concentration (K o ) around its physiological value, of various K+ channels blockers, including internal Cs+, of vacuolar H+-ATPase inhibitors and of the protonophore CCCP on the resting potential and the voltage-dependent K+ current of differentiated neuroblastoma x glioma hybrid NG108-15 cells using the whole-cell patch-clamp technique. The results are as follows: (i) under standard conditions (K o =5 mm) the membrane potential was –60±1 mV. It was unchanged when K o was decreased to 1 mm and was depolarized by 4±1 mV when Ko was increased to 10 mm. (ii) Internal Cs+ depolarized the membrane by 21±3 mV. (iii) The internal application of the vacuolar H+-ATPase inhibitors N-ethylmaleimide (NEM), NO 3 and bafilomycin A1 (BFA) depolarized the membrane by 15±2, 18±2 and 16±2 mV, respectively, (iv) When NEM or BFA were added to the internal medium containing Cs+, the membrane was depolarized by 45±1 and 42±2 mV, respectively. (v) The external application of CCCP induced a transient depolarization followed by a prolonged hyperpolarization. This hyperpolarization was absent in BFA-treated cells. The voltage-dependent K+ current was increased at negative voltages and decreased at positive voltages by NEM, BFA and CCCP. Taken together, these results suggest that under physiological conditions, the resting potential of NG108-15 neuroblastoma cells is maintained at negative values by both voltage-dependent K+ channels and an electrogenic vacuolar type H+-ATPase.This work was supported by a grant from INSERM (CRE 91 0906).  相似文献   

12.
Summary Previous platelet studies have shown that calcium plays important roles in stimulus-secretion coupling, aggregation, and other membrane-associated functions. In addition, lanthanum induces platelet aggregation and the platelet release reaction and also influences platelet responsiveness to various stimuli. The spin-label results presented here suggest that one mechanism through which calcium and lanthanum mediate their effects on platelet functions may be by decreasing the lipid fluidity of the surface membrane.The structure of platelet membrane lipids was examined with the spin-label method. Washed human platelets were labeled with the 5-, 12- and 16-nitroxide stearic acid spin probes. Order parameters which measure the fluidity of the lipid environment of the incorporated probe may be calculated from the electron spin resonance (ESR) spectra of 5-nitroxide stearate [I(12,3)]-labeled cells. Evidence is presented which indicates that these spectra principally reflect properties of the platelet surface membrane lipids. The membrane fluidity increased with temperature for the range 17 to 37 °C. Either calcium or lanthanum additions to intact cells increased the rigidity of the platelet membranes at 37 °C, although the La3+ effect was larger and occurred at lower concentrations than that of Ca2+. For example, addition of 1mm La3+ or 4mm Ca2+ increased the order parameter of I(12,3)-labeled platelets by 4.3±1.7% or 2.1±0.5%. Preliminary studies conducted on purified platelet plasma membranes labeled with I(12,3) indicated that 1mm LaCl3 or 4mm CaCl2 additions similarly decreased the lipid fluidity at 37 °C. The above cation-induced effects on the fluidity of whole platelets were reversed by the use of the divalent cation-chelating agent ethylene glycol-bis-(-aminoethyl ether)-N,N-tetra-acetic acid (EGTA). Lastly, lanthanum (0.2–1mm) caused rapid aggregation of platelets which were suspended in a 50-mm Tris buffer pH 7.4 that did not contain adenosine.  相似文献   

13.
Outwardly rectifying chloride channels in lymphocytes   总被引:5,自引:0,他引:5  
Summary Outwardly rectifying Cl channels in cultured human Jurkat T-lymphocytes were activated by excising a patch of membrane using the inside-out (i/o) patch-clamp configuration and holding at depolarized voltages for prolonged periods of time (1–6 min at +80 mV, 20°C). The single-channel current at +80 mV was 4.5 ± 0.3 pA and at –80 mV, it was 1.0 ± 0.4 pA. After activation, the probability of being open (P 0)for the lymphocyte channel was voltage independent. Activation of the Cl channel in lymphocytes was temperature dependent. Nineteen percent of i/o recordings from lymphocytes made at 20°C exhibited Cl channel activity. In contrast, 49% of recordings made at 30°C showed channel activity. The number of channels in an active patch was not significantly different at the two temperatures. Channel activation in excised, depolarized patches also occurred 20-fold faster at 30°C than at 20°C. There was no marked change in the single-channel conductance at 30°C. Open-channel conductance was blocked by 200 m indanyloxyacetic acid (IAA) or 1 mm SITS when applied to the intracellular side of the patch. The characteristics of this channel are similar to epithelial outwardly rectifying Cl channels thought to be involved in fluid secretion  相似文献   

14.
The mechanism of nitrate transport across the tonoplast of barley root cells   总被引:14,自引:0,他引:14  
Nitrate-selective microelectrodes were used to measure not only nitrate activity in the cytoplasm and vacuole of barley (Hordeum vulgare L.) root cells, but also the tonoplast electrical membrane potential. For epidermal cells, the mean cytoplasmic and vacuolar pNO3 (-log10 [NO3]) values were 2.3±0.04 (n=19) and 1.41±0.03 (n=35), respectively, while for cortical cells, the mean cytoplasmic and vacuolar nitrate values were 2.58±0.18 (n=4) and 1.17±0.06 (n=13), respectively. These results indicate that the accumulation of nitrate in the vacuole must be an active process. Proton-selective microelectrodes were used to measure the proton gradient across the tonoplast to assess the possibility that nitrate transport into the vacuole is mediated by an H+/NO 3 antiport mechanism. For epidermal cells, the mean cytoplasmic and vacuolar pH values were 7.12±0.06 (n=10) and 4.93±0.11 (n=22), respectively, while for cortical cells, the mean cytoplasmic and vacuolar pH values were 7.24±0.07 (n=3) and 5.09±0.17 (n=7), respectively. Calculations of the energetics for this mechanism indicate that the observed gradient of nitrate across the tonoplast of both epidermal and cortical cells could be achieved by an H+/NO 3 antiport with a 11 stoichiometry.Abbreviations and Symbols G/F free-energy change for H+/NO 3 antiport - F Faraday constant - pHc cytoplasmic pH - pHv vacuolar pH - p[NO3]c log10 (cytoplasmic [NO 3 ]) - P[NO3]v -log10 (vacuolar [NO3]) We wish to thank Dr. K. Moore for assistance with statistical analysis.  相似文献   

15.
The effect of oxidative stress, induced by Fe2+-EDTA system, on Na+,K+-ATPase, Na+/Ca2+ exchanger and membrane fluidity of synaptosomes was investigated. Synaptosomes isolated from gerbil whole forebrain were incubated in the presence of 200 M FeSO4-EDTA per mg of protein at 37°C for 30 min. The oxidative insult reduced Na+,K+-ATPase activity by 50.7 ± 5.0 % and Na+/Ca2+ exchanger activity measured in potassium and choline media by 47.1 ± 7.2 % and 46.7 ± 8.6 %, respectively. Membrane fluidity was also significantly reduced as observed with the 1,6-diphenyl-1,3,5-hexatriene probe. Stobadine, a pyridoindole derivative, prevented the decrease in membrane fluidity and in Na+/Ca2+ exchanger activity. The Na+,K+-ATPase activity was only partially protected by this lipid antioxidant, indicating a more complex mechanism of inhibition of this protein. The results of the present study suggest that the Na+/Ca2+ exchanger and the Na+,K+-ATPase are involved in oxidation stress-mediated disturbances of intracellular ion homeostasis and may contribute to cell injury.  相似文献   

16.
Hydroxide, bicarbonate and buffer anion permeabilities in semitendinosus muscle fibers of Rana pipiens were measured. In all experiments, the fibers were initially equilibrated in isotonic, high K2SO4 solutions at pH o =7.2 buffered with phosphate. Two different methods were used to estimate permeabilities: (i) membrane potential changes were recorded in response to changes in external ion concentrations, and (ii) intracellular pH changes were recorded in response to changes in external concentrations of ions that alter intracellular pH. Constant field equations were used to calculate relative or absolute permeabilities.In the first method, to increase the size of the membrane potential change produced by a sudden change in anion entry, external K+ was replaced by Cs+ prior to changes of the anion under study. At constant external Cs+ activity, a hyperpolarization results from increasing external pH from 7.2 to 10.0 or higher, using either CAPS (3-[cyclohexylamino]-1-propanesulfonic acid) or CHES (2-[N-cyclohexylamino]-ethanesulfonic acid) as buffer. For each buffer, the protonated form is a zwitterion of zero net charge and the nonprotonated form is an anion. Using reported values of H+ permeability, calculations show that the reduction in [H+] o cannot account for the hyperpolarizations produced by alkaline solutions. Membrane hyperpolarization increases with increasing total external buffer concentration at constant external pH, and with increasing external pH at constant external buffer anion concentration. Taken together, these observations indicate that both OH and buffer anions permeate the surface membrane. The following relative permeabilities were obtained at pHo, 10.0± 0.3: (POH/PK) = 890 ± 150, (PCAPS/PK) = 12 ± 2 (PCHIES/PK) = 5.3 ± 0.9, and (PNO3/PK) = 4.7 ± 0.5 PNO/PK was independent of pH o up to 10.75. At pHo = 9.6, (PHCO3/PK) = 0.49 ± 0.03; at pH o = 8.9, (PCl/PK) = 18± 2 and at pH o = 7.1, (PHEPES/PK) = 20 ± 2.In the second method, on increasing external pH from 7.2 to 10.0, using 2.5 mm CAPS (total buffer concentration), the internal pH increases linearly with time over the next 10 min. This alkalinization is due to the entry of OH and the absorption of internal H+ by entering CAPS anion. The rate of CAPS entry was determined in experiments in which the external CAPS concentration was increased at constant external pH. Such increases invariably produced an increase in the rate of internal alkalinization, which was reversed when the CAPS concentration was reduced to its initial value. From the internal buffer power, the diameter of the fiber under study and the rates of change of internal pH, the absolute permeability for both OH and CAPS were calculated. At external pH = 10.0, the average (±sem) permeabilities were: POH=1.68±0.19×10–4 cm/sec and PCAPS=2.10±0.74×10–6cm/sec.We conclude that OH is about 50 times more permeable than Cl at alkaline pH and that the anionic forms of commonly used buffers have significant permeabilities.This research was supported by a grant from the National Institutes of Health (AR 31814). The authors wish to thank Dr. Peter G. Shrager and Dr. Bruce C. Spalding for reading an early draft of this report and for providing helpful suggestions.  相似文献   

17.
The binding of [14C]cortisol into dog brain synaptosomal plasma membranes (SPM) follows an exponential path described by the general formula y=a.ebx. The specific activity of the SPM-bound (Na++K+)-stimulated ATPase was linearly increased at different concentrations of cortisol. Changes in the allosteric properties of (Na++K+)-stimulated ATPase by fluoride (F) (i. e. changes of Hill coefficients) indicate that cortisol increases the membrane fluidity. The fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene-labeled SPM decreased in cortisol treated SPM compared to untreated (control) SPM, which is consistent with a general increase in membrane fluidity. This increase of fluidity by cortisol may play a role in the physiological effects of this hormone in the brain.  相似文献   

18.
Summary Literature data suggest that water accumulation by the human fetus is driven by osmotic gradients of small solutes. However, the existence of such gradients has not been supported by prior measurements. Attempts to estimate the size of the gradient necessary to drive net water movement have been seriously hampered by the lack of permeability data for the syncytiotrophoblast membranes. Stopped-flow light scattering techniques were employed to measure the osmotic water permeability (P f )of microvillous (MVM) and basal membrane (BM) vesicles isolated from human term placenta. At 37°C, the P f was determined to be 1.9±0.06 × 10+–3 cm/sec for MVM and 3.1±0.20 × 10+–3 cm/sec for BM (mean ±SD, n = 6). At 23°C, P f was reduced to 0.7±0.04 × 10+–3 cm/sec in MVM and 1.6±0.05 × 10+–3 cm/sec in BM. These P f values are comparable to those observed in membranes where water has been shown to permeate via a lipid diffusive mechanism. Arrhenius plots of P f over the range 20–40°C were linear, with activation energies of 13.6 ± 0.6 kcal/mol for MVM and 12.9±1.0 kcal/mol for BM. Water permeation was not affected by mercurial sulfhydryl agents and glucose transport inhibitors. These data clearly suggest that water movement across human syncytiotrophoblast membranes occurs by a lipid diffusion pathway. As noted in several other epithelial tissues, the basal membrane has a higher water permeability than the microvillous membrane. It is speculated that water accumulation by the human fetus could be driven by a solute gradient small enough to be within the error of osmolarity measurements.We thank the staff of the labor and delivery ward at University of San Francisco Medical Center for help in obtaining placental tissue. This work was supported by NIH grant HD 26392. Dr. Jansson was supported by the Sweden-America Foundation, The Swedish Society of Medicine, The Swedish Society for Medical Research, and the Swedish Medical Research Council.  相似文献   

19.
We monitored soil emissions of NO, NO2, N2O, and CO2 throughout the summer dry season at a remote North American sagebrush-steppe ecosystem following application of several resources, including water, NH 4 + , NO 3 and sucrose. Despite low levels of soil NH 4 (5.60±0.95 mg NH 4 -N per kg soil, mean ± S.E.), and NO 3 -N (1.34±0.20 mg NO 3 -N per kg soil), NO emissions ranged from about 0.2 to 2.8 ng NO-N m–2 s–1, comparable to rates measured from many agricultural, tropical, and other undisturbed ecosystems. Soil wetting increased NO emissions as much as 400-fold when initial gravimetric soil moisture contents were less than about 50 mg kg soil –1 and soil temperature was greater than or equal to 20 °C. Wetting treatments with 20 mg NH 4 + -N kg soil –1 raised NO emission rates to a level that was nearly an order of magnitude higher than that observed after water addition alone. Wetting treatments with 20 mg NO 3 -N kg soil –1 , 240 mg sucrose-C kg soil –1 , or NO 3 plus sucrose had no statistically significant effect upon NO emissions. Soil denitrifying enzyme activity was low at this site, and N2O emissions in the field were below detection limits. Soil nitrifying enzyme activity was extremely high at this site, indicating that the NH 4 + released by ammonification would be consumed at least once every 1.7 days. These observations indicate that NO emissions from this undisturbed ecosystem were likely a consequence of high nitrification activity, and that sagebrush-steppe ecosystems may be a more important NO source than has been previously assumed.  相似文献   

20.
The NO donor 3-Morpholinosydnonimine (SIN-1) releases NO in the presence of molecular oxygen. In this study, we evaluated the effect of SIN-1 on mitochondria of rat cortical synaptosomes. We demonstrated in vitro that the amount of ONOO generated and H2O2 formation directly correlated with SIN-1 concentration. The mean oxygen consumption by synaptosomal mitochondria was approximately 3.8 nmol of O2 min−1 mg−1 protein, which decreased significantly in the presence of SIN-1 1 mM to 2.5 nmol O2 min−1 mg−1. This decrease was not modified by catalase or Trolox, demonstrating that ONOO was responsible for the effect. The same concentration of SIN-1 caused a significant decrease of ATP production by synaptosomal mitochondria and depolarized the mitochondrial membrane. Moreover, ROS production increased progressively and was completely inhibited by pre-incubation of synaptosomes with Trolox. Finally, phosphatidylserine was externalized and, at the same time, intrasynaptosomal lactate dehydrogenase decreased confirming both, the external membrane breakdown after the addition of SIN-1 and the damage to the synaptosomes.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号