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1.
Elementary K+ currents were recorded at 19 °C in cell-attached and in inside-out patches excised from neonatal rat heart myocytes. An outwardly rectifying K+ channel which prevented Na+ ions from permeating could be detected in about 10% of the patches attaining (at 5 mmol/l external K+ and between – 20 mV and + 20 mV) a unitary conductance of 66 +- 3.9 pS. K
(outw.-rect.)
+
channels have one open and at least two closed states. Open probability and open rose steeply on shifting the membrane potential in the positive direction, thereby tending to saturate. Open probability (at –7 mV) was as low as 3 ± 1% but increased several-fold on exposing the cytoplasmic surface to Mg-ATP (100 mol/l) without a concomitant change of open. No channel activation occurred in response to ATP in the absence of cytoplasmic Mg–+. The cytoplasmic administration of the catalytic subunit of protein kinase A (120–150 /ml) or GTP--S (100 mol/l) caused a similar channel activation. GDP--S (100 mol/l) was also tested and found to be ineffective in this respect. This suggests that cardiac K
(outw.-rect.)
+
channels are metabolically modulated by both cAMP-dependent phosphorylation and a G-protein.
Offprint requests to: M. Kohlhardt 相似文献
2.
Synaptosomal acetylcholine synthesis was found to be dependent on the presence of Na+-dependent HC-3 sensitive choline transport at low (5.5 mM) and high (35 mM) K+ concentrations. However, at 5, 20, and 100 M choline, choline phosphorylation was proportional to total choline uptake, in the presence or absence of high affinity transport. Only in the presence of eserine (50 M) did acetylcholine synthesis increase as the choline concentration was elevated from 20 M to 100 M, and this effect was observed at low and high K+ concentrations. Our results suggest that: 1) the synthesis of non-surplus synaptosomal ACh is dependent on high affinity choline transport; and 2) choline is equally likely to be phosphorylated after being taken up by low or high affinity transport. 相似文献
3.
Extraneuronal catecholamine uptake was investigated in isolated quiescent rat myocardial cells. By administration of (3H-)(–)noradrenaline concentration of 22 nmol/l up to 1000 mol/l the following data were obtained: (1) The KM of the uptake process amounted to 260 mol/l, the Vmax to 4.24 nmol/(10 min × mg Protein) corresponding to 179 nmol/(min × gWWt)(WWT = Wet Weight). (2) The uptake was largely inhibited by the uptake2-inhibitors corticosterone (100 mol/l), isoprenaline (IC so = 30.6 mol/l), and O-methylisoprenaline (IC50 = 2.1 pmol/l), but not by the uptake1-inhibitors cocaine (100 mol/l) and desipramine (10 mol/l). (3) The affinity-values KM and IC50 closely agreed with those already known, but the Vmax-value was higher than those obtained in whole rat hearts by a factor of at least 1.79. This is caused presumably by the voltage dependence of the uptake mechanism and the resulting inhibition of uptake 2 during the periods of depolarisation in beating hearts of other studies. 相似文献
4.
Ian Findlay 《The Journal of membrane biology》1988,101(1):83-92
Summary The effects of ADP upon the gating of ATP-sensitive K+ channels from rat ventricular myocytes have been investigated by patch-clamp single-channel current recording experiments. ADP was applied to the internal surface of excised insideout membrane patches and depending upon the experimental protocol and the concentration it was found that ADP could either inhibit or stimulate openings of ATP-sensitive K+ channels. In the absence of inactivation, ATP-sensitive K+ channels were inhibited by ADP in a dose-dependent manner. Partially inactivated channels, on the other hand, were stimulated by low (10 to 250 M) and inhibited by high (>250 M) concentrations of ADP. ATP-sensitive K+ channels which were being inhibited by ATP (<1 mM) could be opened by the simultaneous application of ADP (50 M to 1 mM). ADP had no effect upon channels inhibited by mM concentrations of ATP. The situation was further complicated when it was found that inhibition evoked by ADP was strongly attenuated by the presence of Mg2+ ions whilst channel stimulation, whether of partially inactivated channels or channels inhibited by ATP, required the presence of Mg2+ ions. The analog of ADP, ADPS, always evoked inhibition of ATP-sensitive K+ channels which was not affected by the presence or absence of Mg2+ ions. 相似文献
5.
Sharma Shweta Bhat T.K. Dawra R.K. 《World journal of microbiology & biotechnology》1999,15(6):673-677
Tannase (tannin acyl hydrolase E.C. 3.1.1.20) has been isolated from Aspergillus niger van Tieghem and purified 29-fold. The enzyme had a temperature optimum of 60°C, pH optimum of 6.0 with a second peak at pH 4.5, Km of 0.20mM and Vmax of 5.0mol min–1 mg–1protein. 相似文献
6.
Summary Ionic currents from freshly dissociated rabbit corneal endothelial cells were examined using patch-clamp technology and a perforated patch technique. Whole-cell current recordings revealed a transient outward K+-selective current that was blockable in a dose-dependent manner by 4-aminopyridine (4-AP) and quinidine. This current is similar to the A-type current present in many excitable cells and is the first reported instance of such a current in any epithelial cell type. In addition to the transient current, an outwardly rectifying nonselective cation current was also observed. This current is also blocked by quinidine.To examine the possible role of these currents in the stromal volume regulatory function of the endothelium, corneas were perfused under a specular microscope with a glutathionebicarbonate Ringer's solution (GBR) or GBR plus either 1 mM quinidine or 10 mM 4-AP. For quinidine perfusions, control corneas swelled at a rate of 6 m/hr, while quinidine-perfused corneas swelled at a rate of 48 m/hr. For 4-AP perfusions, control corneas deswelled at a rate of –2 m/hr, while 4-AP perfused corneas swelled at a rate of 24 m/hr. One possible mechanism of the stromal swelling induced by these K+ channel blockers may be the result of loss of the K+ recycling pathway necessary for proper Na+/K+ ATPase function.We would like to thank Dr. William Bourne for the use of his specular microscopy corneal perfusion apparatus and Helen Hendrickson for her technical assistance. This work was supported by NIH grants EY06206, EY03282, EY06005, and an unrestricted award from Research to Prevent Blindness. 相似文献
7.
In Fuji, the production of ethylene was increased with the addition of AgNO3 and inhibited with the addition of 10 M aminoethoxyvinylglycine (AVG). The addition of 80 M AgNO3 to transformed explants of Fuji cultured on selection medium resulted in increased ethylene production (20 l l–1) at 3 weeks. Under examining the effect of AgNO3 in Fuji, the 40 M AgNO3 showed with higher 33.8% and 6.5% in the efficiency of regeneration and transformation. However, ethylene production in Gala explants treated with 10M AgNO3 (3 l l–1) decreased after 2 weeks compared with the control (5 l l–1). Although the regeneration efficiency of Gala with 10 M AgNO3 was higher (41.1%) than the control (20.1%), there was no significant difference in the transformation efficiency at the same concentration. Shoot regeneration of Fuji and Gala was completely inhibited with 10 M AVG. These results suggest that the addition of AgNO3 affects the efficiency of Agrobacterium-mediated gene transfer in Fuji.Eun Soo Seong, Ill Min Chung- These two Authors Contributed equally to this work 相似文献
8.
In hippocampal neurons, 5-hydroxytryptamine (5-HT) activates an inwardly rectifying K+ current via G protein. We identified the K+ channel activated by 5-HT (K5-HT channel) and studied the effects of G protein subunits and nucleotides on the K+ channel kinetics in adult rat hippocampal neurons. In inside-out patches with 10 m 5-HT in the pipette, application of GTP (100 m) to the cytoplasmic side of the membrane activated an inwardly rectifying K+ channel with a slope conductance of 36±1 pS (symmetrical 140 mm K+) at –60 mV and a mean open time of 1.1±0.1 msec (n=5). Transducin activated the (K5-HT) channels and this was reversed by -GDP. Whether the K5-HT channel was activated endogenously (GTP, GTPS) or exogenously (), the presence of 1 mm ATP resulted in a 4-fold increase in channel activity due in large part to the prolongation of the open time duration. These effects of ATP were irreversible and not mimicked by AMPPMP, suggesting that phosphorylation might be involved. However, inhibitors of protein kinases A and C (H-7, staurosporine) and tyrosine kinase (tyrphostin 25) failed to block the effect of ATP. These results show that G activates the G protein-gated K+ channel in hippocampal neurons, and that ATP modifies the gating kinetics of the channel, resulting in increased open probability via as yet unknown pathways. 相似文献
9.
Summary Isolated and homogenised Deiters' neurons from the lateral vestibular nucleus of rabbit in a Krebs-Ringer solution containing no added Mg++, 1.3 moles/ ml and 5 moles/ml Mg++, broke down ATP at the maximal rate of 0.29+-0.20, 2.40+–0.20, and 5.95+–0.63 moles/cell/hr. In 1.3 mM Mg++ solution the single cell homogenates took up phosphate at the mean rate of 2.6+–0.2 moles/cell/hr. If the rabbits were injected 1 hour before with 20 mg/kg body weight of 2-amino-1-propene-1,1,3, tricarbonitrile (triap), the breakdown of ATP in these latter media was 0.82+–0.44, 2,5+–0.60, and 6.7+– 1.1 moles/cell/hr, respectively, and the quantity of inorganic liberated did not decrease. 相似文献
10.
Multiple buds have been induced from cotyledonary nodes of cashew, Anacardium occidentale L. Maximum number of buds were initiated on Murashige & Skoog's (MS) medium supplemented with 117 mM sucrose, 14.6 mM maltose and 22.2 M BA. The buds were harvested at each subculture after elongation on MS medium supplemented with 100 ml l-1 coconut water, 14.6 mM maltose, increased sucrose (146 mM) and decreased BA (4.4 M). Excised microshoots were rooted in vitro on MS medium in the presence of 117 mM sucrose, 2.9 M IAA and 4.9 M IBA. Plantlets have been successfully transferred to soil and have been established in the field. 相似文献
11.
Teresa Piñol Javier Palazon Teresa Altabella Manuel Serrano 《Journal of Plant Growth Regulation》1987,5(4):183-189
Growth, K+ content, and alkaloid production were compared in nonorganogenetic callus cultures ofNicotiana tabacum cv. Burley 21 grown at 25°C in the dark on two different media: a basal medium with 1 M -naphthaleneacetic acid and 1 M kinetin, and one with 1 M -naphthaleneacetic acid and 1 M 4PU-30 (N-(2-chloro-4-pyridyl)-N-phenylurea). These callus tissues behaved differently not only in growth and K+ content but also in alkaloid production. In comparison to cultures grown with kinetin, those grown with 4PU-30 showed a significantly higher fresh weight and dry weight and K+ content during the growth period studied. The data clearly indicate a positive correlation between K+ uptake rate stimulated by 4PU-30 and cell enlargement rate. However, the alkaloid biosynthesis in the callus tissues was activated by the supply of kinetin and diminished by that of 4PU-30. It thus appears that cellular enlargement of meristematic tissue stimulated by 4PU-30 limited alkaloid production. 相似文献
12.
John W. Daly Yukio Nishizawa Michael W. Edwards James A. Waters Robert S. Aronstam 《Neurochemical research》1991,16(4):489-500
Nicotinic agonists stimulate22Na flux in rat pheochromocytoma PC12 cells. The stimulatory effect of carbamylcholine is maximal at 1 mM, while the stimulatory effect of nicotine and anatoxin maximize at the same level at 100 M and 10 M, respectively. The tertiary amines arecolone and isoarecolone have no effect on flux at 100 M, while the methiodides at 100 M stimulate flux to an extent similar to 1 mM carbamylcholine. Dihydro and alcohol analogues of isoarecolone methiodide have markedly smaller effects on flux. A preincubation for 2 to 20 min with carbamylcholine (2 mM), nicotine (300 M), anatoxin (30 M) or isoarecolone methiodide (100 M) causes marked desensitization to a subsequent carbamylcholine-elicited stimulation of flux. d-Tubocurarine, mecamylamine, hexamethonium, and chlorisondamine inhibit carbamylcholine-elicited flux with IC50 values of 1.0, 0.8, 43, and 0.020 M, respectively. Atropine has no effect at 1 M, but reduces the response to carbamylcholine by 50% at 8.6 M, presumably as a noncompetitive blocker. Other noncompetitive blockers of nicotinic acetylcholine-receptors, such as histrionicotoxins, gephyrotoxin, pumiliotoxin C, phencyclidine, bupivacaine and piperocaine, inhibit carbamylcholine-elicited stimulation of22Na flux with IC50 values from 0.3 to 1.8 M. In contrast to d-tubocurarine, which inhibits carbamylcholine-elicited desensitization, and mecamylamine, which has no apparent effect on desensitization, chlorisondamine and certain noncompetitive blockers appear to enhance desensitization. The effects of agonists, antagonists and noncompetitive blockers at the neuronal nicotinic acetylcholine receptor-channel of PC12 cells are compared to their effects on binding of [125I]-bungarotoxin to agonist-recognition sites and of [3H]perhydrohistrionicotoxin to noncompetitive blocker sites of the nicotinic acetylcholine receptor-channel of electric ray (Torpedo) electroplax membranes. There are marked differences in relative potencies for the two types of nicotinic acetylcholine receptor-channel. 相似文献
13.
D. L. C. Procter 《Polar Biology》1987,7(5):303-306
Summary Respiration of an undescribed species of soil nematode of the genus Chiloplacus from the Canadian High Arctic was measured at 2°, 5°, 10°, 15°, 20° and 25°C. The corresponding metabolic rates were 0.2697×10-3 l, 0.3406×10-3 l, 0.8408×10-3 l, 0.8539×10-3 l, 1.8420×10-3 l and 2.9360×10-3 l O2 ind-1 h-1, respectively, for a nematode of 1.0 g dry weight. The relationship between respiration and dry weight for Chiloplacus sp. at 10°C is described by the function log R=-3.0693+0.8844 log W. Q10 values for the 2°–5°, 5°–10°, 10°–15°, 15°–20° and 20°–25°C temperature intervals were 2.18, 6.09, 1.03, 4.65 and 2.54, respectively. Chiloplacus sp. showed raised metabolic rates at low tempetatures compared with species from warmer environments. Metabolic rates of representative samples of the soil, nematode fauna (dominated by individuals of the genus Plectus) from the same location were 0.1593×10-3 l, 0.3603×10-3 l and 0.5332×10-3 l O2 ind-1 h-1 at 5°, 10° and 15°C for an average nematode of 0.4297 g dry weight. 相似文献
14.
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
15.
Summary The present study was designed to examine which type of adenosine receptors was involved in enhancement of high K+-evoked taurine release fromin vivo rat hippocampus using microdialysis. Perfusion with 0.5 or 5.0 mM adenosine enhanced high K+-evoked taurine release. Perfusion with 2M R(–)-N6-2-phenylisopropyladenosine (PIA), a selective adenosine A1 receptor agonist, did not modulate taurine release. Perfusion with 1M 1,3-dipropyl-8-cyclopentylxanthine (DPCPX), a selective adenosine A1 receptor antagonist, increased taurine release. On the other hand, perfusion with 20M 2-[4-(2-carboxyethyl)phenethylamino]-5-N-ethyl-carboxamideadenosine (CGS21680), a selective adenosine A2A receptor agonist, enhanced taurine release, while perfusion with 1 mM 3,7-dimethyl-propagylxanthine (DMPX), an adenosine A2 receptor antagonist, did not affect taurine release. These results demonstrate that adenosine enhances high K+-evoked taurine release via activation of adenosine A2A receptors from both neurons and glial cells ofin vivo rat hippocampus. 相似文献
16.
The ionic requirements for K+-evoked efflux of endogenous taurine from primary cerebellar astrocyte cultures were studied. The Ca2+ ionophore A23187 evoked taurine efflux in a dose-dependent fashion with a time-course identical to that of K+-induced efflux. The Ca2+-channel antagonist nifedipine had no effect upon efflux induced by 10 or 50 mM K+. In addition, verapamil did not antagonize 50 mM K+-evoked efflux except at high, non-pharmacological concentrations (>100 M), and preincubation with 2 M -conotoxin had no effect on 50 mM K+-evoked efflux. Similarly, preincubation with 1 mM ouabain had no effect on the amount of taurine released by K+ stimulation, but did accelerate the onset of efflux by 2–4 min. Although 2 M tetrodotoxin had no effect on K+-evoked release, replacing Na+ with choline abolished the taurine efflux seen in response to K+ stimulation. Together, these findings suggest that neuronal N- and L-type Ca2+- and voltage-dependent Na+-channels are not involved in the influx of Ca2+ which appears to be necessary for K+-evoked taurine efflux, and that in addition to Ca2+, extracellular Na+ is also required. 相似文献
17.
Ben A. Bulthuis Gregory M. Koningstein Adriaan H. Stouthamer Henk W. van Verseveld 《Antonie van Leeuwenhoek》1993,63(1):1-16
The magnitude of the proton motive force (p) and its constituents, the electrical () and chemical potential (-ZpH), were established for chemostat cultures of a protease-producing, relaxed (rel
–) variant and a not protease-producing, stringent (rel
+) variant of an industrial strain ofBacillus licheniformis (respectively referred to as the A- and the B-type). For both types, an inverse relation of p with the specific growth rate was found. The calculated intracellular pH (pHin) was not constant but inversely related to . This change in pHin might be related to regulatory functions of metabolism but a regulatory role for pHin itself could not be envisaged. Measurement of the adenylate energy charge (EC) showed a direct relation with for glucose-limited chemostat cultures; in nitrogen-limited chemostat cultures, the EC showed an approximately constant value at low and an increased value at higher . For both limitations, the ATP/ADP ratio was directly related to .The phosphorylation potential (G'p) was invariant with . From the values for G'p and p, a variable H+/ATP-stoichiometry was inferred: H+/ATP=1.83+0.52µ, so that at a given H+/O-ratio of four (4), the apparent P/O-ratio (inferred from regression analysis) showed a decline of 2.16 to 1.87 for =0 to max (we discuss how more than half of this decline will be independent of any change in internal cell-volume). We propose that the constancy of G'p and the decrease in the efficiency of energy-conservation (P/O-value) with increasing are a way in which the cells try to cope with an apparent less than perfect coordination between anabolism and catabolism to keep up the highest possible with a minimum loss of growth-efficiency. Protease production in nitrogen-limited cultures as compared to glucose-limited cultures, and the difference between the A- and B-type, could not be explained by a different energy-status of the cells.Abbreviations CCCP
carbonylcyanide-p-trichloromethoxyphenylhydrazone
- DW
dry weight of biomass
- F
Faraday's constant, 96.6 J/(mV × mol)
- Fo
chemostat outflow-rate (ml/h)
- FCCP
carbonylcyanide-p-trifluoromethoxyphenylhydrazone
- G'p
phosphorylation potential, the Gibbs energy change for ATP-synthesis from ADP and Pi
- G'0p
standard Gibbs energy change at specified conditions
- H+/ATP
number of protons translocated through
- ATP
synthase in synthesis of one ATP
- H+/O
protons translocated during transfer of 2 electrons from substrate to oxygen
-
specific growth rate (1/h)
- H+
transmembrane electrochemical proton potential, J/mol
- Mb
molar weight (147.6 g/mol) of bacteria with general cell formula C6.0H10.8O3.0N1.2
- pHout,in
extracellular, intracellular pH
- Pi
(intracellular) inorganic phosphate
- p
proton motive force, mV
- pH
transmembrane pH-difference
-
transmembrane electrical potential, mV
- P/O
number of ADP phosphorylated to ATP upon reduction of one O2– to H2O by two electrons transferred through the electron transfer chain
- P/O
(H+/O) × (H+/ATP)–1
- P/OF, P/ON
P/O with the two electrons donated by resp. (NADH + H+) and FADH
- q
specific rate of consumption or production (mol/g DW × h)
-
rel
+,rel
–
stringent, relaxed genotype
- R
universal gas constant, 8.36 J/(mol × degree)
- T
absolute temperature
- TPMP+
triphenylmethylphosphonium ion
- TPP+
tetraphenyl phosphonium ion
- Y
growth yield, g DW/mol
- Z
conversion constant=61.8 mV for 310 K (37 °C)
- ZpH
transmembrane proton potential or chemical potential, mV 相似文献
18.
The activities and kinetics of the enzymes G6PDH (glucose-6-phosphate dehydrogenase) and 6PGDH (6-phosphogluconate dehydrogenase) from the mesophilic cyanobacterium Synechococcus 6307 and the thermophilic cyanobacterium Synechococcus 6716 are studied in relation to temperature. In Synechococcus 6307 the apparent K
m's are for G6PDH: 80M (substrate) and 20M (NADP+); for 6PGDH: 90M (substrate) and 25M (NADP+). In Synechococcus 6716 the apparent K
m's are for G6PDH: 550M (substrate) and 30M (NADP+); for 6PGDH: 40M (substrate) and 10M (NADP+). None of the K
m's is influenced by the growth temperature and only the K
m's of G6PDH for G6P are influenced by the assay temperature in both organisms. The idea that, in general, thermophilic enzymes possess a lower affinity for their substrates and co-enzymes than mesophilic enzymes is challenged.Although ATP, ribulose-1,5-bisphosphate, NADPH and pH can all influence the activities of G6PDH and 6PGDH to a certain extent (without any difference between the mesophilic and the thermophilic strain), they cannot be responsible for the total deactivation of the enzyme activities observed in the light, thus blocking the pentose phosphate pathway.Abbreviations G6PDH
glucose-6-phosphate, dehydrogenase
- 6PGDH
6-phosphogluconate dehydrogenase
- G6P
glucose-6-phosphate
- 6PG
6-phosphogluconate
- RUDP
ribulose-1,5-bisphosphate
- Tricine
N-Tris (hydroxymethyl)-methylglycine 相似文献
19.
Microdissected Deiters' neuron plasma membranes have been used for studying the passage of GABA through the membrane both in the inward and outward direction. Working with 0.2 mM GABA in the compartment simulating the outside of the neurone and with 2.0 mM GABA in the one simulating the inside we found a net transport of GABA towards the inside. This mechanism does not require a Na+ ion gradient across the membrane. The nature of the transport process involved was studied by determining the rate of [3H]-GABA inward passage as a function of GABA concentration (1 nM–800 M) on the outward side of the membrane. The results have shown that until 50 M a diffusion process (v=D1×C, where D1=3.1×10–11 1/m2×sec) is the sole mechanism involved. Above 50 M a second diffusion process is activated v=D2×(C–50×10–6), where D2=2.8×10–11 1/m2×sec. Taking in account both inward and outward directed diffusion, one can calculate 16 M as the equilibrium concentration of GABA on the outward side of the membrane. From a kinetic point of view, these diffusion processes are able to reduce GABA concentration in a synaptic cleft from 3 mM to 20 M within 3 sec. These diffusion systems are discussed as extremely efficient in removing the excess of released GABA in the synaptic cleft. 相似文献
20.
The microsomal fraction isolated from dog mesenteric nerve fibres was found to contain ATPase activity stimulated by micromolar concentrations of Ca ions. Such a high-affinity Ca2+-ATPase (hereafter referred to as HA Ca-ATPase) followed a Michaelis-Menten kinetics with Km for Ca ions of 0.4 M and Vmax=12.5±2.4 mol Pi.mg–1h–1. The examination of the subcellular origin of HA Ca-ATPase revealed that this enzyme is associated with axonal plasma membranes as documented by its co-purification with several plasma membrane marker enzymes and with tetrodotoxin-sensitive3H-saxitoxin binding. The addition of exogenous magnesium ions (Mg) resulted in a non-competitive inhibition of HA Ca-ATPase with Ki=0.5 mM. The reaction velocity of HA Ca-ATPase was also inhibited by other divalent ions with the order of potency Mg>Mn >ZnCo>Ni. In contrast to low affinity (high Km) Mg- and Ca-ATPase, the HA Ca-ATPase was insensitive to the inhibition by sodium azide (10 mM) and sodium fluoride (10 mM). Similarly, the specific activity of HA Ca-ATPase was unaffected by vanadate (100 M) and N-ethylmaleinimide (100 M). It is concluded that axonal plasma membranes of dog mesenteric nerves contain HA Ca-ATPase which seems to be unrelated to calcium-transporting Mg-dependent, Ca-stimulated ATPase.Abbreviations used BSA
bovine serum albumin
- HA Ca-ATPase
high-affinity Ca2+-ATPase
- K-pNPPase
onabain-sensitive, K+-stimulated p-nitrophenyl phosphatase
- NEM
N-ethylmaleinimide
- SIM
250 mM sucrose, 10 mM imidazole-HCl pH 7.4
- TRIS
tris (hydroxymethyl) aminomethane 相似文献