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1.
Summary To assess the mechanism(s) by which intraluminal chloride concentration is raised above equilibrium values, intracellular Cl– activity (
i
Cl
) was studied in the proximal tubule ofNecturus kidney. Paired measurements of cell membrane PD (V
BL) and Cl-selective electrode PD (V
BL
Cl
) were performed in single tubules, during reversible shifts of peritubular or luminal fluid composition. Steadystate
i
Cl
was estimated at 14.6±0.6 mmol/liter, a figure substantially higher than that predicted for passive distribution. To determine the site of the uphill Cl– transport into the cell, an inhibitor of anion transport (SITS) was added to the perfusion fluid. Introduction of SITS in peritubular perfusate decreased
i
Cl
, whereas addition of the drug in luminal fluid slightly increased
i
Cl
; both results are consistent with basolateral membrane uphill Cl– transport from interstitium to the cell. TMA+ for Na+ substitutions in either luminal or peritubular perfusate had no effect on
i
Cl
. Removal of bicarbonate from peritubular fluid, at constant pH (a situation increasing HCO
3
–
outflux), resulted in an increase of
i
Cl
, presumably related to enhanced Cl– cell influx: we infer that Cl– is exchanged against HCO
3
–
at the basolateral membrane. The following mechanism is suggested to account for the rise in luminal Cl– concentration above equilibrium values: intracellular CO2 hydration gives rise to cell HCO
3
–
concentrations above equilibrium. The passive exit of HCO
3
–
at the basolateral membrane energizes an uphill entry of Cl– into the cell. The resulting increase of
i
Cl
, above equilibrium, generates downhill Cl– diffusion from cell to lumen. As a result, luminal Cl– concentration also increases.C.N.R.S. Greco 24. Part of this work was presented at the 12th annual meeting of the American Society of Nephrology, Boston, Mass. (Edelman et al., 1979). 相似文献
2.
Summary Activation of protein kinase C has been shown to cause both stimulation and inhibition of transport processes in the brush-border membrane and renal tubule. This study was designed to examine the dose-response nature and time-dependent effect of 4 -phorbol-12-myristate-13-acetate (PMA) on the rates of bicarbonate absorption (J
HCO3) and fluid absorption (J
v) in the proximal convoluted tubule (PCT) of rat kidney. Bicarbonate flux was determined by total CO2 changes between the collected fluid and the original perfusate as analyzed by microcalorimetry. Luminal perfusion of PMA (10–10 10–5 M) within 10 min caused a significant increase ofJ
HCO3 andJ
v. A peaked curve of the dose response was observed with maximal effect at 10–8 M PMA on both bicarbonate and fluid reabsorption, which could be blocked completely by amiloride (10–3
m) and EIPA (10–5 M). On the other hand, with an increase of perfusion time beyond 15 min, PMA (10–8 and 10–6 M) could inhibitJ
HCO3 andJ
v. Amiloride (10–3 M) or EIPA (10–5 M) significantly inhibitsJ
HCO3 andJ
v, while there is no additive effect of PMA and amiloride or EIPA on PCT transport. An inactive phorbol-ester, 4-phorbol, that does not activate protein kinase C, had no effects onJ
HCO3 andJ
v. Capillary perfusion of PMA (10–8 M) significantly stimulate bothJ
HCO3 andJ
v; however, PMA did not affect glucose transport from either the luminal side or basolateral side of the PCT. These results indicate that activation of endogenous protein kinase C by PMA could either stimulate or inhibit both bicarbonate and fluid reabsorption in the PCT dependent on time and dose, and these effects are through the modulation of Na+/H– exchange mechanism. 相似文献
3.
High density fed-batch cultures for hybridoma cells performed with the aid of a kinetic model 总被引:1,自引:1,他引:0
R. Pörtner A. Schilling I. Lüdemann H. Märkl 《Bioprocess and biosystems engineering》1996,15(3):117-124
Hybridoma fed-batch cultures with either standard medium as feed or concentrated medium as feed and removal of toxic metabolites through dialysis were performed by using model calculations for a priori determination of process parameters. In a first step a kinetic model for specific growth and death rate, respectively as well as for substrate uptake and metabolite production rates was formulated. In a bed-batch culture with standard medium as feed the appropriate time for start of the feeding pump and the increase of feed rate were determined a priori. The glutamine concentration was controlled at 0.04 mmoll–1. A priori calculation and course of the culture coincided rather well. A cell concentration of 3.2×106 cells ml–1, a MAb-concentration of 54 mg
MAb
l–1 and a MAb-time-space-yield of 0.53 mg
MAb
l–1h–1 were obtained.For further increase of the efficiency a high density fed-batch process was developed, where concentrated medium is fed to the cells and the accumulating toxic low molecular weight metabolites are removed through a dialysis membrane into a dialyizng fluid. In a membrane dialysis reactor consisting of a culture chamber and a dialyzing chamber, which are separated by a cylindrical dialysis membrane, again model calculations were used to determine feed rate and exchange rate of dialyzing fluid. A viable cell density of 1.2×107 cells ml–1 and a MAb concentration of 425 mg l–1 were reached in a culture with stepwise feeding of 10 x concentrated medium and exchange of dialyzing fluid for removal of low molecular metabolites. The course of the culture could be predicted a priori rather well. The MAb-time-space-yield was 2.47 mg
MAb
l–1h–1, appr. 5 times higher compared to fed-batch cultures with standard medium as feed.List of Symbols
A
membrane area m2
-
c
i
substrate or product concentration in culture chamber mmoll–1
-
c
a
substrate or product concentration in dialyzing chamber mmoll–1
-
c
0i
substrate or product concentration in the feed of culture chamber mmoll–1
-
c
0a
substrate or product concentration in the feed of dialyzing chamber mmoll–1
-
c
Gln
glutamine concentration mmoll–1
-
c
Amm
ammonia concentration mmoll–1
-
c
MAb
MAb concentration mmoll–1
-
D
a
dilution rate in dialyzing chamber d–1
-
F
i
feed rate during fed-batch to the culture chamber mlh–1
-
V
a
volume of dialyzing chamber l
-
V
i
volume of culture chamber l
-
P
membrane permeability coefficient cm min–1
-
q
specific substrate uptake or metabolite production rate mmol cell–1 h–1
-
q
Gln
spec. glutamine uptake rate mmol cell–1 h–1
-
q
MAb
spec. MAb production rate mmol cell–1 h–1
-
t
time h
-
X
v
viable cell concentration cells ml–1
-
MAb
MAb-time-space-yield mgl–1 h–1
-
specific growth rate h–1
-
d
specific death rate h–1
Financial support from the Volkswagen-Stiftung, Germany, grand nr. I/69 359 is gratefully acknowledged.The concentrated medium was kindly provided by SERVA, Heidelberg, Germany. The hybridoma cell line was donated by Prof. fil. dr. Volker Kasche, Technische Universität Hamburg-Harburg, Germany.We express our special thanks to Andreas Schütt, Ralf Gassner, Katja Herbers and Thomas Schäfer for their help in this project. 相似文献
4.
Summary The Na+ requirement for active, electrogenic Cl– absorption byAmphiuma small intestine was studied by tracer techniques and double-barreled Cl–-sensitive microelectrodes. Addition of Cl– to a Cl–-free medium bathingin vitro intestinal segments produced a saturable (K
m
=5.4mm) increase in shortcircuit current (I
sc) which was inhibitable by 1mm SITS. The selectivity sequence for the anion-evoked current was Cl–=Br–>SCN–>NO
3
–
>F–=I–. Current evoked by Cl– reached a maximum with increasing medium Na concentration (K
m
=12.4mm). Addition of Na+, as Na gluconate (10mm), to mucosal and serosal Na+-free media stimulated the Cl– current and simultaneously increased the absorptive Cl– flux (J
ms
Cl
) and net flux (J
net
Cl
) without changing the secretory Cl– flux (J
sm
Cl
). Addition of Na+ only to the serosal fluid stimulatedJ
ms
Cl
much more than Na+ addition only to the mucosal fluid in paired tissues. Serosal DIDS (1mm) blocked the stimulation. Serosal 10mm Tris gluconate or choline gluconate failed to stimulateJ
ms
Cl
. Intracellular Cl– activity (a
Cl
i
) in villus epithelial cells was above electrochemical equilibrium indicating active Cl– uptake. Ouabain (1mm) eliminated Cl– accumulation and reduced the mucosal membrane potential
m
over 2 to 3 hr. In contrast, SITS had no effect on Cl– accumulation and hyperpolarized the mucosal membrane. Replacement of serosal Na+ with choline eliminated Cl– accumulation while replacement of mucosal Na+ had no effect. In conclusion by two independent methods active electrogenic Cl– absorption depends on serosal rather than mucosal Na+. It is concluded that Cl– enters the cell via a primary (rheogenic) transport mechanism. At the serosal membrane the Na+ gradient most likely energizes H+ export and regulates mucosal Cl– accumulation perhaps by influencing cell pH or HCO
3
–
concentration. 相似文献
5.
Russell F. Husted Loren H. Cohen Philip R. Steinmetz 《The Journal of membrane biology》1979,47(1):27-37
Summary Bicarbonate is transferred across the serosal (S) membrane of the epithelial cells of the turtle bladder in two directions. Cellular HCO
3
–
generated behind the H+ pump moves across this membrane into the serosal solution. This efflux of HCO
3
–
is inhibited by SITS (4-isothiocyano-4-acetamido-2,2-disulfonic stilbene). When HCO
3
–
is added to the serosal solution it is transported across the epithelium in exchange for absorbed Cl–. This secretory HCO
3
–
flow traverses the serosal cell membrane in the opposite direction. In this study the effects of serosal addition of 5×10–4
m SITS on HCO
3
–
secretion and Cl– absorption were examined. The rate of H+ secretion was brought to zero by an opposing pH gradient, and 20mm HCO
3
–
was added toS. HCO
3
–
secretion, measured by pH stat titration, was equivalent to the increase inMS Cl– flux after HCO
3
–
addition. Neither theSM flux of HCO
3
–
nor theMS flux of Cl– were affected by SITS. In the absence of electrochemical gradients, net Cl– absorption was observed only in the presence of HCO
3
–
in the media; under such conditions, unidirectional and net fluxes of Cl– were not altered by serosal or mucosal SITS. H+ secretion, however, measured simultaneously as the short-circuit current in ouabain-treated bladders decreased markedly after serosal SITS. The inhibition of the efflux of HCO
3
–
in series with the H+ pump and the failure of SITS to affect HCO
3
–
secretion and Cl– absorption suggest that the epithelium contains at least two types of transport systems for bicarbonate in the serosal membrane. 相似文献
6.
Summary In the plasmodium ofPhysarum polycephalum application of various monovalent cation salts elicited either a depolarization or a hyperpolarization of the membrane potential. The hyperpolarization was restricted to phosphates and bicarbonates of large cations (Na+, Li+, NMe4
+, NEt4
+). More than 50 other combinations of cations (K+, Rb+, Cs+, NH4
+) and anions (Cl–, NO3
–, SO4
2–, acetate, lactate, citrate, etc.) induced the depolarization. In both cases the magnitude of the deflection in membrane potential () varied linearly with logarithm of concentration above the threshold Cth (10–4 M for all monovalent cation salts examined) according to the following equation: =± R log (C/Cth).The value of R was 10–15 mV, and plus and minus signs correspond to depolarization and hyperpolarization, respectively. Depolarizing and hyperpolarizing agents competed with each other and exhibited a sharp transition between the two states of the membrane which were characterized by — R and R in the above equation or displayed a strong hysteresis, depending on which agents had first been applied to the plasmodial membrane. This transition in the membrane potential corresponded to the transition between positive and negative taxis at the behavioral level. 相似文献
7.
Summary The kinetics of lactate dehydrogenase in situ were studied in sections of unfixed liver of the male mouse using a quantitative histochemical technique. The sections were incubated on substrate-containing gel films. The absorbance of the final reaction products deposited in a single hepatocyte was measured continuously during the incubation as a function of incubation time using a scanning microdensitometer. The absorbance increased non-linearly during the first minute of incubation, but linearly for at least the next 3 min afterwards. The initial velocity (v
i
) of the dehydrogenase was calculated from two equations proposed previously by us, v
i=2.82 °A and v
i
=v+2°A, where v and °A are, respectively, the gradient and intercept o linear regression line of absorbance on time for incubation times between 1 and 3 min.The dependence of v
i
on lactate concentration gave the following mean kinetic constants. For periportal hepatocytes, the apparent K
m
=14 mM and V
max
=80 moles hydrogen equivalents formed cm–3 hepatocyte cytoplasm min–1. For pericentral hepatocytes, K
m
=12 mM and V
max
=87 moles hydrogen equivalents cm–3 min–1. The K
m
values are very similar to those determined previously from biochemical assays. The concentrations of the enzyme in single hepatocytes calculated from the V
max
values are in good agreement with those obtained by another method. These data substantiate the validity of our equations. 相似文献
8.
An assay is described that allows the direct measurement of the enzyme activity catalyzing the transfer of the methyl group from N
5-methyltetrahydromethanopterin (CH3–H4MPT) to coenzyme M (H–S–CoM) in methanogenic archaebacteria. With this method the topology, the partial purification, and the catalytic properties of the methyltransferase in methanol- and acetate-grown Methanosarcina barkeri and in H2/CO2-grown Methanobacterium thermoautotrophicum were studied. The enzyme activity was found to be associated almost completely with the membrane fraction and to require detergents for solubilization. The transferase activity in methanol-grown M. barkeri was studied in detail. The membrane fraction exhibited a specific activity of CH3–S–CoM formation from CH3–H4MPT (apparent K
m=50 M) and H–S–CoM (apparent K
m=250 M) of approximately 0.6 mol·min-1·mg protein-1. For activity the presence of Ti(III) citrate (apparent K
m=15 M) and of ATP (apparent K
m=30 M) were required in catalytic amounts. Ti(III) could be substituted by reduced ferredoxin. ATP could not be substituted by AMP, CTP, GTP, S-adenosylmethionine, or by ATP analogues. The membrane fraction was methylated by CH3–H4MPT in the absence of H–S–CoM. This methylation was dependent on Ti(III) and ATP. The methylated membrane fraction catalyzed the methyltransfer from CH3–H4MPT to H–S–CoM in the absence of ATP and Ti(III). Demethylation in the presence of H–S–CoM also did not require Ti(III) or ATP. Based on these findings a mechanism for the methyltransfer reaction and for the activation of the enzyme is proposed.Abbreviations H4MPT
tetrahydromethanopterin
- CH3–H4MPT
N
5-methyl-H4MPT
- H–S–CoM
2-mercaptoethanesulfonate or coenzyme M
- CH3–S–CoM
2(methylthio)ethanesulfonate or methylcoenzyme M
- SDS-PAGE
sodium dodecylsulfate polyacrylamide gel electrophoresis
- DTT
dithiothreitol
- MOPS
morpholinopropanesulfonate
- CHAPS
3-[(3-cholamidopropyl)-dimethylammonio]-1-propane-sulfonate
- 1 U =
1 mol/min 相似文献
9.
A possible modulation of permeabilities of membrane vesicles to anions and cations was explored by light scattering techniques, evaluated by measuring the capacity of the vesicles to shrink and swell in response to changes of the osmolarity of the incubation medium. Membrane fractions were obtained by phase partition. Purity was evaluated by detection and quantification of membrane enzyme markers: vanadate-sensitive ATPase for the plasma membrane, nitrate-sensitive ATPase for the tonoplast and azide-sensitive ATPase for mitochondria. Membrane vesicles (250 g protein) were exposed to hypertonic solutions of salts (0.6 osmolar). Kinetics of the changes in apparent absorbance at 546 nm were observed by the addition of potassium, nitrate and chloride salts. The diffusion of ions into vesicles was induced by an osmotic gradient across the membrane and brought about volume changes of vesicles. Upon addition of vesicles to the different solutions the following ion permselectivity sequences were observed: PNO
3
– >PCl
–>PSO
4
2– and PK
+PNa
+>PNH
4
+.Abbreviations ATP
adenosine 5-triphosphate
- EDTA
ethylene diaminetetraacetic acid
- Tris-Mes
(Tris[hydroxymethyl]aminomethane, Mes-(2-[N-Morpholino]ethanesulfonic acid)
- PEG
polyethylene glycol 相似文献
10.
Reduction of nitrate byPseudomonas putrefaciens entrapped in composite agar layer/microporous membrane structures 总被引:1,自引:0,他引:1
Summary Composite structures consisting of aPseudomonas putrefaciens immobilized-cell agar layer bounded by a microporous membrane filter were used for water denitrification. With methanol as the C-source, one litre of high nitrate water (3 mM) was completely freed from NO
3
–
and NO
2
–
ions in 11 days at a rate of 90 mol N–NO
3
–
/day/g of agar gel, while no production of ammonium ions could be detected. When acetic acid was substituted for methanol, the denitrifying activity was lower. No noticeable contamination of the treated water due to cell leakage from the biocatalytic structures occurred during the incubation periods. 相似文献
11.
Summary A stopped-flow rapid reaction apparatus was used to study the rate of pH equilibration in human red cell suspensions. Flux of OH– or H+ was determined over a wide range of extracellular pH (4–11) in CO2-free erythrocyte suspensions. In these experiments, an erythrocyte suspension at pH 7.3 is rapidly mixed with an equal volume of NaCl solution at 3.0>pH>11.5. The pH of the extracellular fluid of the mixture changes rapidly as OH– or H+ moves across the red cell membrane. Flux and velocity constants can be calculated from the initiald pH/dt using the known initial intra- and extracellular pH. It was found that the further the extracellular pH is from 7.3 (in either direction from 4–11), the greater the absolute value of total OH– and/or H+ flux. Pretreatment with SITS (4-acetamido-4-isothiocyanostilbene-2,2-disulfonic acid), a potent anion exchange inhibitor, greatly reduces flux over the entire pH range, while exposure to valinomycin, a potassium ionophore, has no measurable effect. These data suggest that (i) both H+ and OH– may be moving across the red cell membrane at all pH; (ii) the species dominating pH equilibration is probably dependent on the extracellular pH, which determines the magnitude of the driving gradient for each ion; and (iii) the rapid exchange pathway of the erythrocyte membrane may be utilized for both H+ and OH– transport during CO2-free pH equilibration. 相似文献
12.
Two maize lines differing in drought resistance were grown at different drought stress induced by polyethylene glycol (PEG 10 000) solutions with osmotic potentials of –0.20, –0.40 and –0.80 MPa in the semipermeable membrane system. During the five days soil water content decreased (from 0.43 to 0.29, 0.25 and 0.23 g cm–3 for three PEG solutions, respectively) as well as leaf water potentials (w; from – 0.54 to –0.76, –1.06 and –1.46 MPa). These values were not significantly different between the investigated lines, indicating that a controlled and consistent soil moisture stress was achieved. Soil drying induced an increase in the ABA content of leaves and xylem of both lines and the effects on stomatal conductance were greater in drought susceptible line (B-432) compared to drought resistant line (ZPBL-1304). To test possible difference in stomatal sensitivity to xylem ABA between lines and to assess any ABA vs. w interaction, roots were fed with 10, 50 and 100 mmol m–3 ABA solutions in another set of experiments. These results showed that manipulation of xylem ABA affected stomata of both lines similarly. Comparison of stomatal sensitivity to drought-induced and applied ABA demonstrated that drought treatment affected stomata of investigated lines by differentially increasing their sensitivity to xylem ABA, thus confirming an interaction between chemical signalling and hydraulic signalling. 相似文献
13.
Summary A method for removal of toxic hexavalent chromium (chlomate: CrO
inf4
sup2–
) was developed by use of dialysis-sac cultures of a chromate-reducing strain of Enterobacter cloacae (HO1). E. cloacae strain HO1 cells were put in dialysis (semipermeable membrane) sacs, and the sacs were submerged in water containing toxic CrO
inf4
sup2–
. The dialysis sacs allowed CrO
inf4
sup2–
to diffuse into the culture, and CrO
inf4
sup2–
was reduced anaerobically in the dialysis sacs by the E. cloacae cells. Because reduced chromium readily formed insoluble chromium hydroxides in the dialysis sacs, the greater part of reduced chromium was unable to diffuse out through the semipermeable membrane. Thus the dialysis culture of E. cloacae strain HO1 could successfully remove toxic chromium from the surrounding water. If the initial concentration of CrO
inf4
sup2–
was less than 4mM (208 ppm as chromium), about 90% of the total chromium could be removed from water by the described method.
Offprint requests to: H. Ohtake 相似文献
14.
Prof. Dr. Eng. S. Furusaki Dr. Agr. T. Nozawa S. Nomura 《Bioprocess and biosystems engineering》1990,5(2):73-78
A membrane enzyme reactor with simultaneous separation was investigated. Enzymes, urease and aspartase, were immobilized by a porous polytetrafluoroethylene membrane. Electrical field was applied in the medium while the reaction was carried out. Products with electrical charge could be separated through the membrane from the reaction medium as they were formed. Reaction behavior was analyzed by a simple model considering both pore-migration and reaction in the skelton of the membrane. According to the analysis the inherent reaction rate of the immobilized enzymes decreases significantly. This is probably caused by the structural variation of enzymes. For the case of urease, the change of pH inside the membrane may also cause the decrease of the reaction rate. The model analysis showed that the enzyme content in the membrane and the residence time of the substrate in the membrane governed overall extent of reaction.List of Symbols
e g (dm3)–1
enzyme concentration in the membrane
-
L cm
membrane thickness
-
K
m
mM
Michaelis constant
-
Rate mmol · min–1 · g–1
rate of product formation per unit weight of enzyme
-
S mM
substrate concentration
-
S
in mM
inlet substrate concentration
-
S
out mM
outlet substrate concentration
-
u cm · min–1
migration rate
-
V V
voltage between the electrodes
-
V
m mmol · min–1 · g–1
maximum reaction rate
-
X
conversion
-
z cm
distance from the surface inside the membrane
-
void fraction of the porous membrane
-
tortuosity of the membrane
-
min
space time 相似文献
15.
An anion channel that only allows outward current flow (anion influx) has been identified in protoplasts derived from wheat (Triticum aestivum L., Triticum turgidum L.) roots. The anion outward rectifier (anion OR) measured by patch-clamp of whole cells activated very quickly, usually reaching a steady-state level in less than 100 ms and was easily distinguished from the cation outward rectifier (cation OR) which activated more slowly during membrane depolarisation. The anion OR is permeable to NO
3
–
and Cl–, moderately permeable to I–, and relatively impermeable to H2PO4/– and ClO4/–. An anomalous mole-fraction effect between ClO4/
– and Cl– was observed on the outward current, indicating that the channel is a multi-ion pore. The anion OR is gated by both voltage and external anion concentration such that it activates near to the equilibrium potential for the permeant anion. It activated at more negative membrane potentials when NO
3
–
was substituted for Cl– in the external medium, indicating that the channel may function to allow NO
3
–
influx under luxuriant external NO
3
–
concentrations. For most experiments, K+ and Cl– were the main cation and anion in solution, and under these conditions it appeared likely that the anion OR functioned in membrane-potential regulation by facilitating a Cl– influx at membrane potentials more positive than the chloride reversal potential (ECl). If ECl was more negative than the K+ reversal potential (EK) then the anion OR dominated but both the anion and cation ORs occurred together when the membrane potential difference (Vm) was positive of both ECl and EK. The cation OR was inhibited by increasing external Cl– concentrations, but the anion OR was not affected by external K+ or Na+ concentration. The anion-transport inhibitors, zinc and phenylglyoxal were ineffective in blocking the anion OR. 4,4-Di-isothiocyanostilbene-2, 2-disulfonic acid (DIDS) irreversibly blocked about 34% of the current when applied extracellularly at a concentration of 25 M, and about 69% at a concentration of 200 M. However, DIDS (200 M) also occasionally acted as an irreversible blocker of the cation OR. Perchlorate blocked irreversibly 75% of the current at an external concentration of 10 mM and did not block the cation OR. Whole-cell currents also indicated that the anion OR was insensitive to external pH (pH=5–7) and calcium concentration ([Ca2+]=0.1–10 mM). Increasing intracellular calcium concentration significantly increased the occurrence of the fast outward current in whole cells (P < 0.005, X2 test). With approximately 10 nM calcium inside the cell the anion outward current was observed in 64% (n = 45) of cells and with 50 nM calcium inside the cell the anion current was observed in 88% (n = 69) of cells. Single-anion OR channels observed in outside-out patches had a conductance in 300 mM KCl (external) of about 4 pS. When voltage pulses were applied to outside-out patches the average currents were similar to those observed in whole cells. The significance of the anion OR as a likely route for Cl uptake in high salinities is discussed.Abbreviations Bath
solution bathing the extracellular face of the membrane
- DIDS
(4,4-diisothiocyanostilbene-2,2-disulfonic acid)
- Ex
reversal potential for ion x
- OR
outward rectifier
-
Pip
solution inside the pipette
- TEACl
(tetraethyl-ammonium chloride)
- Vm
membrane potential difference
We thank the Australian Research Council for financial support, G.P. Findlay and A. Garrill for helpful discussions, and K. Morris and D. Mackenzie for expert technical assistance. M.S. was supported by an Australian Postgraduate Research Award. 相似文献
16.
M. J. Beilby 《The Journal of membrane biology》1981,62(3):207-218
Summary The changes in the cytoplasmic Cl– concentration, [Cl–]
c
, are monitored at the time of withdrawal (starvation) and subsequent replacement of Cl– in the outside medium. The measurement technique exploits the involvement of Cl– inChara excitation. The transient clamp current due to Cl–,I
Cl, is separated from other excitation transients through Hodgkin-Huxley (HH) equations, which have been adjusted toChara. TheI
Cl amplitude depends on HH parameters, [Cl–]
c
and the maximum membrane conductance to Cl–,
. The results are discussed in terms of these quantities.I
Cl and
were found to fall after 6–10 hr of Cl– starvation, thus supporting the hypothesis that [Cl–
c
decreases in Cl–-free medium. The best HH fit to starved data was obtained with [Cl–
c
=3.5mm. The time-course forI
Cl decline is considerably slower than the time-course of the rise of the starvation-stimulated influx. As cells starved for periods longer than 24 hr are re-exposed to Cl–, it is revealed that while [Cl–]
c
remains low during long starvation,
increases to values greater than those of the normal cells. Such differences among cells starved for various lengths of time have not been detected previously. 相似文献
17.
The effects of sunlight and the indigenous microbiota onEscherichia coli survival were examined with membrane diffusion chambers deployed in Chesapeake Bay shellfish growing waters. Chambers, fitted with an upper UV and visible light-transmitting copolymer film and lower semipermeable polycarbonate membrane, were deployed parallel to the water surface to maximize light exposure. Maximum values of a coefficient, kdens, describing changes in culturable cell densities after exposure to sunlight, were 1.7 h–1 and 0.7 h–1 in preliminary tank and in situ experiments, respectively. Mortality and sublethal stress, the latter measured with an electrochemical detection technique, were largest during the first 4 h of exposure. Owing to the light-attenuating properties of Chesapeake Bay water, light-induced cell mortality was significantly reduced at 0.25 m compared with surface exposed cells, and was undetected at 0.5–1.0 m except during seasons of maximal light penetration. Combined exposure to both sunlight and the autochthonous microbiota yielded significantly greater mortality than for either factor alone. 相似文献
18.
Summary Diffusion of auxin (indole-3-acetic acid) through planar lipid bilayer membranes was studied as a function of pH and auxin concentration. Membranes were made of egg or soybean lecithin or phosphatidyl serine inn-decane (25–35 mg/ml). Tracer and electrical techniques were used to estimate the permeabilities to nonionized (HA) and ionized (A–) auxin. The auxin tracer flux is unstirred layer limited at low pH and membrane limited at high pH, i.e., when [A–][HA]. The tracer flux is not affected by the transmembrane voltage and is much higher than the flux predicted from the membrane conductance. Thus, only nonionized auxin crosses the membrane at a significant rate. Auxin transport shows saturation kinetics, but this is due entirely to unstirred layer effects rather than to the existence of an auxin carrier in the membrane. A rapid interconversion of A– and HA at the membrane surface allows A– to facilitate the auxin flux through the unstirred layer. Thus, the total flux is higher than that expected for the simple diffusion of HA alone. The relation between flux (J
A), concentrations and permeabilities is: 1/J
A=1/P
UL([A–]+[HA])+1/P
HA
M
[HA]. By fitting this equation to our data we find thatP
UL=6.9×10–4 cm/sec andP
HA
M
=3.3×10–3 cm/sec for egg lecithin-decane bilayers. Similar membrane permeabilities were observed with phosphatidyl serine or soybean lipids. Thus, auxin permeability is not affected by a net surface charge on the membrane. Our model describing diffusion and reaction in the unstirred layers can explain the anomolous relationship between pH and weak acid (or weak base) uptake observed in many plant cells. 相似文献
19.
Summary The transepithelial water permeability in frog urinary bladder is believed to be essentially dependent on the ADH-regulated apical water permeability. To get a better understanding of the transmural water movement, the diffusional water permeability (P
d) of the basolateral membrane of urinary bladder was studied. Access to this post-luminal barrier was made possible by perforating the apical membrane with amphotericin B. The addition of this antibiotic increasedP
d from 1.12±0.10×10–4 cm/sec (n=7) to 4.08±0.33×10–4 cm/sec (n=7). The effect of mercuric sulfhydryl reagents, which are commonly used to characterize water channels, was tested on amphotericin B-treated bladders. HgCl2 (10–3
m) decreasedP
d by 52% andpara-chloromercuribenzoic acid (pCMB) (1.4×10–4
m) by 34%. The activation energy for the diffusional water transport was found to increase from 4.52±0.23 kcal/mol (n=3), in the control situation, to 9.99±0.91 kcal/mol (n=4) in the presence of 1.4×10–4
m
pCMB. Our second approach was to measure the kinetics of water efflux, by stop-flow light scattering, on isolated epithelial cells from urinary bladders.pCMB (0.5 or 1.4×10–4
m) was found to inhibit water exit by 91±2%. These data strongly support the existence of proteins responsible for water transport across the basolateral membrane, which are permanently present. 相似文献
20.
Horst Onken Kai Graszynski 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1989,159(1):21-28
Summary Isolated posterior gills from Chinese crabs (Eriocheir sinensis) acclimated to tap-water were perfused and bathed with full, physiological saline (containing Na+ and Cl–). Under these conditions they developed an outside positive transepithelial potential difference (PDte). Substitution of Na+ by choline on both sides of the epithelium resulted in a substantial hyperpolarization of the PDte, while substitution of Cl– by gluconate reverses PDte to outside negative values.The magnitudes of the potential differences were strongly related to the adaptation media (artificial seawater or tap-water).The KCN-sensitive, outside positive PDte was shown to be strongly dependent on Cl–. Application of thiocyanate and 4-acetamido-4-isothiocyanato-stilbene-2,2 disulfonic acid (SITS) to the bath solution resulted in a reduction of the PDte, while the Cl–-channel blocker, diphenylamine-2-carboxylic acid (DPC), showed no effect. The PDte was markedly reduced by acetazolamide, an inhibitor of carbonic anhydrase (CA), and these results are discussed with reference to the presence of a Cl–/HCO
3
–
-exchanger in the apical membrane.Chloride was shown to pass the basolateral membrane via Cl–-channels: Diphenylamine-2-carboxylic acid (DPC) reduced the PDte with an IC50 of 3.7×10–5 mol/l when added to the perfusion saline. A basolateral K+-channel and its linkage to Cl– uptake could be demonstrated by using the K+-channel blocker, Ba2+, or increased K+ concentrations in the perfusion saline (PDte decrease). Ouabain did not reduce the PDte under nominally Na+-free conditions, indicating that the Cl– transport is independent of the Na+/K+-ATPase. In this paper we shall discuss the possible energy sources and linkages between pH regulation and active Cl– absorption under these experimental conditions.Abbreviations
A9C
anthracene-9-carboxylic acid
-
CA
carbonic anhydrase
-
DMSO
dimethylsulfoxide
-
DPC
diphenylamino-2-carboxylic acid
-
PD
te
transepithelial potential difference
-
SITS
4-acetamido-4-isothiocyanato-stilbene-2,2-disulfonic acid
-
TEA
tetraethyl-ammoniumchloride 相似文献