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11.
D. R. Laver 《The Journal of membrane biology》1990,118(1):55-67
Summary The patch-clamp technique is used here to investigate the kinetics of Ca2+ block in single high-conductance Ca2+-activated K+ channels. These channels are detected in the membrane surounding cytoplasmic drops fromChara australis, a membrane which originates from the tonoplast of the parent cell. The amplitudes and durations of single channel events are measured over a wide range of membrane potential (–300 to 200 mV). Ca2+ on either side of the channel reduces its K+ conductance and alters its ion-gating characteristics in a voltage-dependent manner. This Ca2+-induced attenuation of conductance is analyzed using the theory of diffusion-limited ion flow through pores. Interaction of external Ca2+ with the channel's ion-gating mechanism is examined in terms of a kinetic model for ion-gating that includes two voltage-dependent gating mechanisms. The kinetics of channel block by external Ca2+ indicates that (i) external Ca2+ binds at two sites, a superficial site and a deep site, located at 8 and 40% along the trans-pore potential difference, (ii) the external vestibule cannot be occupied by more than one Ca2+ or K+, and (iii) the kinetics of Ca2+ binding at the deep site is coupled with that of a voltage-dependent gate on the external side of the channel. Kinetics of channel block by internal Ca2+ indicates that more than one Ca2+ is involved. 相似文献
12.
Roger A. Hoffman Peter Habeeb Gerald R. Buzzell 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1990,160(3):269-275
Summary Long-term increased or decreased circulating levels of thyroid hormones significantly modify porphyrin concentrations and morphology in the Harderian glands of male and female hamsters. Administration of T3 reduced porphyrin concentrations in females; this treatment or decreasing thyroid hormone levels with KClO4 suppressed the post-castration rise of porphyrins in males. Hypophysectomy led to increased porphyrins in the Harderian glands of males; this rise was suppressed in hypophysectomized males by T3 or T4. In females, hypophysectomy reduced porphyrins which were further reduced by daily administration of T3 or T4. These modifications in the normal females were identical in castrated males. Mitotic activity in the Harderian glands of females was stimulated by KClO4 and by hypophysectomy with or without exogenous T3. In males, castration increased mitotic activity which was suppressed by T3 and exacerbated by KClO4. Increased mitotic activity seemingly follows loss of tissue mass. The data show that thyroid hormones act directly on the Harderian glands rather than indirectly through modification of TSH synthesis/release. Female type glands in males are a consequence of loss of gonadal androgens by castration, or by suppression or loss of thyroid hormones by hypophysectomy or by treatment with KClO4. However, male type glands in females are the result of androgen treatment, and/or increased levels of thyroid hormones via reduced ambient temperatures or of photic input. We conclude that regulation of the Harderian gland appears to be different in the two sexes.Abbreviations
T
3
Triiodothyronine
-
T
4
Thyroxine
-
TSH
Thyroid Stimulating Hormone
-
KClO
4
Potassium Perchlorate
-
h
hours
-
ml
milliliter
-
mg
milligram
-
g
gram
-
male
-
female
-
castrated male
- AP
hypophysectomized
-
CON
Control
-
ALA
delta aminole-vulenic acid
-
HG
Harderian Gland 相似文献
13.
Influence of single amino acids on the development of hamster one-cell embryos in vitro 总被引:1,自引:0,他引:1
One-cell hamster embryos placed in culture have always shown a complete block to development at the two-cell stage. In a preliminary study using a chemically defined culture medium containing 20 amino acids (HECM-1), many one-cell embryos were able to escape the "two-cell block" and develop to the four-cell stage. Use of a simpler formulation containing only the amino acids hypotaurine and glutamine revealed marked inhibitory and stimulatory effects of adding the other amino acids. In the first experiment, 19 amino acids were separately examined for effects on one-cell embryo development. Six amino acids (phenylalanine, valine, isoleucine, tyrosine, tryptophan, and arginine) inhibited embryo development (reduced mean cell number; MCN), and three others (glycine, cystine, and lysine) stimulated development (increased MCN), compared with basic medium containing only glutamine and hypotaurine (low control). When the responses with the six inhibitory amino acids were totalled, only 3 of 185 (2%) one-cell embryos reached the six-or seven-cell stage compared to a total of 15 of 76 (20%) embryos that developed to these stages using the three stimulatory amino acids. When tested together in a second experiment, the six inhibitory amino acids significantly reduced the MCN, from 4.28 +/- 0.44 (low control) to 3.71 +/- 0.55. In this group, 17 of 117 (15%) of one-cell embryos reached more than four-cell and only 4 of 117 (3%) reached six- or 7-cell stages, compared with 39 of 117 (33%) and 12 of 117 (10%), respectively, for the basal medium group.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
14.
Estimating denitrification rates in estuarine sediments: A comparison of stoichiometric and acetylene based methods 总被引:6,自引:0,他引:6
Samantha B. Joye Stephen V. Smith James T. Hollibaugh Hans W. Paerl 《Biogeochemistry》1996,33(3):197-215
We compared denitrification rates obtained using an adaptation of the acetylene block technique to rates estimated from benthic flux nutrient stoichiometry in the subtidal sediments of Tomales Bay, California (USA). By amending whole cores with acetylene and saturating nitrate concentrations, we obtained potential denitrification rates, which ranged between 4 and 30 mmol N m–2 d–1. We determined the apparent Michaelis constant (Kapp) and the maximum potential rate (Vmp) of the denitrifying community and used these constants in a rectangular hyperbola to estimatein situ denitrification rates. Both the Kapp and Vmp of the denitrifying community exhibited significant variation over both depth in the sediment column and time of sampling.Estimates ofin situ denitrification obtained using our kinetic-fix adaptation of the acetylene block ranged between 1.8 (March) and 9 (Sept.) mmol N m–1 d–1. Denitrification rates obtained using benthic flux stoichiometry ranged between 0.7 and 4.1 mmol N m–2 d–1. Average denitrification rates obtained using the kinetic-fix acetylene block approach exceeded those obtained from net benthic flux stoichiometry; however, these differences were not significant. We conclude that our kinetic-fix adaptation of the acetylene block technique provides realistic estimates of denitrification in sediments, even when pore water nitrate concentrations are low and nitrification and denitrification are closely coupled. 相似文献
15.
In the evergreen Fagaceae forests of Japan, an ectomycorrhizal fungusTricholoma bakamatsutake forms shiros or developing mycelial blocks. To determine the physiological characteristics of the mycelial blocks, organic acids of the soil and major nutrient elements of the soil and roots were compared at three types of sites: presently colonized mycelial blocks, previously colonized sites behind the blocks, and uncolonized sites in front of the blocks. The upper part of the mycelial blocks showed the following features compared with the uncolonized site: lower pH (5.1), higher concentrations of oxalic and gluconic acids, lower content of total nitrogen, a similar amount of total carbon, reduced total and available phosphorus, higher content of total calcium and lower content of exchangeable calcium. These findings suggested that the activity of the fungus led to soil acidification by the organic acids, an increase in C/N ratio, depletion of phosphorus and accumulation of calcium. 相似文献
16.
Sybil P. Seitzinger Lars Peter Nielsen Jane Caffrey Peter Bondo Christensen 《Biogeochemistry》1993,23(3):147-167
Measurements of denitrification using the acetylene inhibition,15N isotope tracer, and N2 flux methods were carried out concurrently using sediment cores from Vilhelmsborg sø, Denmark, in an attempt to clarify some of the limitations of each technique. Three experimental treatments of overlying water were used: control, nitrate enriched, and ammonia enriched water. The N2 flux and15N tracer experiments showed high rates of coupled nitrification/denitrification in the sediments. The acetylene inhibition method did not capture any coupled nitrification/denitrification. This could be explained by acetylene inhibition of nitrification. A combined15N tracer/acetylene inhibition experiment demonstrated that acetylene inhibition of N2O reduction was incomplete and the method, therefore, only measured approximately 50% of the denitrification due to nitrate from the overlying water. Similar rates of denitrification due to nitrate in the overlying water were measured by the N2 flux method and the acetylene inhibition method, after correcting for the 50% efficiency of acetylene inhibition. Rates of denitrification due to nitrate from the overlying water measured by the15N tracer method, however, were only approximately 35% or less of those measured by the acetylene inhibition or N2 flux methods. 相似文献
17.
Uwe Ludewig Christoph Lorra Olaf Pongs Stefan H. Heinemann 《European biophysics journal : EBJ》1993,22(4):237-247
The members of the RCK family of cloned voltage-dependent K+ channels are quite homologous in primary structure, but they are highly diverse in functional properties. RCK4 channels differ from RCK1 and RCK2 channels in inactivation and permeation properties, the sensitivity to external TEA, and to current modulation by external K+ ions. Here we show several other interesting differences: While RCK1 and RCK2 are blocked in a voltage and concentration dependent manner by internal Mg2+ ions, RCK4 is only weakly blocked at very high potentials. The single-channel current-voltage relations of RCK4 are rather linear while RCK2 exhibits an inwardly rectifying single-channel current in symmetrical K+ solutions. The deactivation of the channels, measured by tail current protocols, is faster in RCK4 by a factor of two compared with RCK2. In a search for the structural motif responsible for these differences, point mutants creating homology between RCK2 and RCK4 in the pore region were tested. The single-point mutant K533Y in the background of RCK4 conferred the properties of Mg2+ block, tail current kinetics, and inward ion permeation of RCK2 to RCK4. This mutant was previously shown to be responsible for the alterations in external TEA sensitivity and channel regulation by external K+ ions. Thus, this residue is expected to be located at the external side of the pore entrance. The data are consistent with the idea that the mutation alters the channel occupancy by K+ and thereby indirectly affects internal Mg2+ block and channel closing.Abbreviations TEA
tetraethylammonium
- EGTA
Ethylene glycol-bis (-aminoethyl ether) N,N,N,N-tetraacetic acid
- 2S3B model
2-site 3-barrier model
Correspondence to: S. H. Heinemann 相似文献
18.
Carla Tatone Christina Grietje Van Eekelen Rosella Colonna 《Molecular reproduction and development》1994,38(2):200-208
The ability of parthenogenetically activated mouse eggs to establish a plasma membrane (PM) block to sperm penetration was studied. Zona-free eggs preloaded with Hoechst 33342 were activated by exposure to ethanol or OAG (1-oleoyl-2-acetyl-sn-glycerol) and inseminated after different periods. Eggs challenged with sperm at 30- or 60-min postactivation displayed a fertilization frequency significantly lower than that of control eggs. Conversely, when insemination was carried out at 120-min postactivation, the proportion of fertilized eggs was equivalent to that observed in the control group. Moreover, we report that when the eggs were induced to resume meiosis without any notable loss of CGs (egg exposure to OAG at 100 μM external Ca2+ or to heat shock), a normal ability to be penetrated was recorded at 30-min postactivation. Similar behaviour was exhibited by eggs that underwent a CG exocytosis close to that triggered by sperm in absence of nuclear activation (microinjection of inositol 1,4,5-trisphosphate into the egg at 1 μM cytosolic concentration). Present data support the conclusion that parthenogenetically activated mouse eggs are capable of a transitory PM block response that requires both CG exocytosis and meiosis resumption to occur. © 1994 Wiley-Liss, Inc. 相似文献
19.
The activation by abscisic acid (ABA) of current through outward-rectifying K+ channels and its dependence on cytoplasmic pH (pHi) was examined in stomatal guard cells of Vicia faba L. Intact guard cells were impaled with multibarrelled and H+-selective microelectrodes to record membrane potentials and pHi during exposures to ABA and the weak acid butyrate. Potassium channel currents were monitored under voltage clamp and, in some experiments, guard cells were loaded with pH buffers by iontophoresis to suppress changes in pHi. Following impalements, stable pHi values ranged between 7.53 and 7.81 (7.67±0.04, n = 17). On adding 20 M ABA, pHi rose over periods of 5–8 min to values 0.27±0.03 pH units above the pHi before ABA addition, and declined slowly thereafter. Concurrent voltage-clamp measurements showed a parallel rise in the outward-rectifying K+ channel current (IK, out) and, once evoked, both pHi and IK, out responses were unaffected by ABA washout. Acid loads, imposed with external butyrate, abolished the ABA-evoked rise in IK, out. Butyrate concentrations of 10 and 30 mM (pH0 6.1) caused pHi to fall to values near 7.0 and below, both before and after adding ABA, consistent with a cytoplasmic buffer capacity of 128±12 mM per pH unit (n = 10) near neutrality. Butyrate washout was characterised by an appreciable alkaline overshoot in pHi and concomitant swell in the steady-state conductance of IK, out. The rise in pHi and iK, out in ABA were also virtually eliminated when guard cells were first loaded with pH buffers to raise the cytoplasmic buffer capacity four- to sixfold; however, buffer loading was without appreciable effect on the ABA-evoked inactivation of a second, inward-rectifying class of K+ channels (IK, in). The pHi dependence of IK, out was consistent with a cooperative binding of at least 2H+ (apparent pKa = 8.3) to achieve a voltage-independent block of the channel. These results establish a causal link previously implicated between cytoplasmic alkalinisation and the activation of IK, out in ABA and, thus, affirm a role for H+ in signalling and transport control in plants distinct from its function as a substrate in H+-coupled transport. Additional evidence implicates a coordinate control of IK, in by cytoplasmic-free [Ca2+] and pHi.Abbreviations ABA
abscisic acid
- [Ca2+]i
cytoplasmic free [Ca2+]i
- EK
K+ equilibrium potential
- IK, out, IK, in
outward-, inward-rectifying K+ channel (current)
- I-V
current-voltage (relation)
- Mes
2-(N-morpholino)ethanesulfonic acid
- pHi
cytoplasmic pH
- Tes
2-{[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]-amino}ethanesulfonic acid
- Vm
membrane potential
We are grateful to G. Thiel (Pflanzenphysiologisches Institut, Universität Göttingen, Germany) for helpful discussions. This work was possible with equipment grants-in-aid from the Gatsby Charitable Foundation, the Royal Society and the University of London Central Research Fund. F.A. holds a Sainsbury Studentship. 相似文献
20.
The behavior of cell nuclei, mitochondrial nucleoids (mt-nucleoids) and plastid nucleoids (ptnucleoids) was studied in the
root apical meristem of Arabidopsis thaliana. Samples were embedded in Technovit 7100 resin, cut into thin sections and stained with 4′-6-diamidino-2-phenylindole for
light-microscopic autoradiography and microphotometry. Synthesis of cell nuclear DNA and cell division were both active in
the root apical meristem between 0 μm and 300 μm from the central cells. It is estimated that the cells generated in the lower
part of the root apical meristem enter the elongation zone after at least four divisions. Throughout the entire meristematic
zone, individual cells had mitochondria which contained 1–5 mt-nucleoids. The number of mitochondria increased gradually from
65 to 200 in the meristem of the central cylinder. Therefore, throughout the meristem, individual mitochondria divided either
once or twice per mitotic cycle. By contrast, based on the incorporation of [3H]thymidine into organelle nucleoids, syntheses of mitochondrial DNA (mtDNA) and plastid DNA (ptDNA) occurred independently
of the mitotic cycle and mainly in a restricted region (i.e., the lower part of the root apical meristem). Fluorimetry, using
a videointensified microscope photon-counting system, revealed that the amount of mtDNA per mt-nucleoid in the cells in the
lower part of the meristem, where mtDNA synthesis was active, corresponded to more than 1 Mbp. By contrast, in the meristematic
cells just below the elongation zone of the root tip, the amount of mtDNA per mt-nucleoid fell to approximately 170 kbp. These
findings strongly indicate that the amount of mtDNA per mitochondrion, which has been synthesized in the lower part of the
meristem, is gradually reduced as a result of continual mitochondrial divisions during low levels of mtDNA synthesis. This
phenomenon would explain why differentiated cells in the elongation zone have mitochondria that contain only extremely small
amounts of mtDNA.
This work was supported by a Grant-in Aid (T.K.) for Special Research on Priority Areas (Project No. 02242102, Cellular and
Molecular Basis for Reproduction Processes in Plants) from the Ministry of Education, Science and Culture of Japan and by
a Grant-in Aid (T.K.) for Original and Creative Research Project on Biotechnology from the Research Council, Ministry of Agriculture,
Forestry and Fisheries of Japan. 相似文献