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11.
Summary Cellular impalements were used in combination with standard transepithelial electrical measurements to evaluate some of the determinants of the spontaneous lumen-positive voltage,V
e
, which attends net Cl– absorption,J
Cl
net
, and to assess how ADH might augment bothJ
Cl
met
andV
e
in the mouse medullary thick ascending limb of Henle microperfusedin vitro. Substituting luminal 5mm Ba++ for 5mm K+ resulted in a tenfold increase in the apical-to-basal membrane resistance ratio,R
c
/R
bl
, and increasing luminal K+ from 5 to 50mm in the presence of luminal 10–4
m furosemide resulted in a 53-mV depolarization of apical membrane voltage,V
a
. Thus K+ accounted for at least 85% of apical membrane conductance. Either with or without ADH. 10–4
m luminal furosemide reducedV
e
andJ
Cl
net
to near zero values and hyperpolarized bothV
a
andV
bl
, the voltage across basolateral membranes; however, the depolarization ofV
bl
was greater in the presence than in the absence of hormone while the hormone had no significant effect on the depolarization ofV
a
, Thus ADH-dependent increases inV
b
were referable to greater depolarizations ofV
bl
in the presence of ADH than in the absence of ADH 68% of the furosemide-induced hyperpolarization ofV
a
was referable to a decrease in the K+ current across apical membranes, but, at a minimum, only 19% of the hyperpolarization ofV
bl
could be accounted for by a furosemide-induced reduction in basolateral membrane Cl– current. Thus an increase in intracellular Cl– activity may have contributed to the depolarization ofV
bl
during net Cl– absorption, and the intracellular Cl– activity was likely greater with ADH than without hormone. Since ADH increases apical K+ conductance and since the chemical driving force for electroneutral Na+,K+,2Cl– cotransport from lumen to cell may have been less in the presence of ADH than in the absence of hormone, the cardinal effects of ADH may have been to increase the functional number of both Ba++-sensitive conductance K+ channels and electroneutral Na+,K+,2Cl– cotransport units in apical plasma membranes. 相似文献
12.
We have studied the properties of kainic acid receptor-activated channels using domoic acid as an agonist. Similarities of the electrophysiological, pharmacological and noise properties of domoic acid and kainic acid-evoked currents confirm that domoate is a potent and specific agonist of the kainate receptor. Single-channel properties of domoic acid-evoked currents were directly determined from outside-out membrane patches for the first time, and results were compared with those obtained by fluctuation analysis of macroscopic currents. Small conductance cationic-selective channels of 4 pS and a mean open time of 2 to 3 ms were detected using both methods.
Offprint requests to: O. Moran 相似文献
13.
We used whole-cell patch-clamp recording techniques to investigate G protein-activated currents in cultured rat retinal pigment
epithelial (RPE) cells. Using 140 mm KCl intracellular and 130 mm NaCl extracellular solutions, rat RPE cells possessed both inward and outward K+ currents. Upon addition of the nonhydrolyzable guanine triphosphate analogue, guanosine-5′-O-(3-thiophosphate) (GTPγS, 0.1
mm), to the recording electrode, a nonspecific cation (NSC) current was elicited. The NSC current had a mean reversal potential
of +5.7 mV in 130 mm extracellular NaCl with Cs+-aspartate in the pipette, and was not affected by alterations in the extracellular Ca2+ or Cl− concentration. The GTPγS-activated current was found to be permeable to several monovalent cations (K+, Na+, choline, TRIS, and NMDG). Addition of fluoroaluminate, an activator of large molecular weight heterotrimeric GTP-binding
proteins (G proteins), to the intracellular recording solution activated the NSC current. The G protein involved was pertussis
toxin (PTX)-sensitive, since GTPγS failed to activate the NSC current in cells pretreated with PTX. Further investigation
of second messenger molecules suggested that activation of the NSC current was not affected by alterations in intracellular
Ca2+ or ATP. From these results, we conclude that a G protein-regulated NSC current is present in rat RPE cells. Activation of
the NSC current may sufficiently depolarize RPE cells to activate outward K+ currents. This would provide a mechanism by which these cells could rid themselves of accumulated K+.
Received: 25 January 1996/Revised: 24 April 1996 相似文献
14.
B. Kodadová 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1996,179(3):301-310
The ability of pheromone receptor cells of male Antheraea polyphemus (Saturniidae) to resolve stimulus pulses was determined at different temperatures (8°, 18°, 28°C). The cells were stimulated by repeated 20-ms puffs of the pheromone components (E, Z)-6, 11-hexadecadienyl acetate and (E, Z)-6,11-hexadecadienal. At higher temperatures, higher frequencies of stimulus pulses were resolved by the nerve-impulse response: about 1.25 pulses per second at 8°C, 2.5 pulses/s at 18°C and 5 pulses/s at 28°C. The decreased ability of receptor cells to resolve stimulus pulses at low temperatures may reduce the male moth's chance of reaching the pheromone source. The peak nerve-impulse frequency increased whereas the duration of nerve-impulse responses to single stimulus pulses decreased at higher temperatures. At a given temperature and stimulus intensity the peak nerveimpulse frequency decreased with shorter intervals between the stimulus pulses, but the duration of the responses remained almost constant. The time needed for recovery from adaptation caused by a single stimulus pulse was longer at lower temperatures. The aldehyde receptor cell recovered more quickly than the acetate cell. At low stimulus concentration, the resolution ability of the acetate cell was strongly decreased, whereas in the aldehyde cell it was only slightly impaired. 相似文献
15.
The temperature dependence of high voltage activated Ca2+ channels has been investigated in cultured dorsal root ganglion neurones from chick embryos, using the cell-attached patch-clamp technique. The dihydropyridine sensitive L-type Ca2+ channel had a conductance of 23 pS, with 110 mM Ba2+ as charge carrier and in the presence of 3 M Bay K 8644. When the temperature was raised from 15 to 30 °C, the unitary channel current amplitude increased, with Q10 value equal to 1.4. The rising phase of the averaged single-channel current became faster, with Q10 value 2.7, whereas the decay phase showed a lower temperature sensitivity. Channel open probability decreased according to an exponential distribution of open and closed times. A second type of Ca2+ channel was identified, which was DHP-insensitive and had a lower conductance with a mean value equal to 13 pS. For the current amplitude, the Q10 value was 1.3. Both activation and inactivation kinetics were strongly accelerated by an increase in temperature. The corresponding time constants gave Q10 values equal to 5.9 for activation, and 2.0 for inactivation. Peak channel open probability was highly sensitive to a change in temperature, with a Q10 value of 1.6. Finally, in -conotoxin GVIA pre-treated neurones, a non-inactivating DHP-insensitive Ca2+ channel with the lowest unitary conductance (10 pS) and a much lower temperature dependence was recorded. Single-channel current was increased by heating, with Q10 value 1.3, whereas the channel kinetics were almost unaffected by temperature. Our data are consistent with the assumption that the different temperature dependence of the Ca2+ channel behaviours may be explained by separate gating processes of three types of Ca2+ channels. 相似文献
16.
L. H. Clapp A. M. Gurney N. B. Standen P. D. Langton 《The Journal of membrane biology》1994,140(3):205-213
Tension and patch clamp recording techniques were used to investigate the relaxation of rabbit pulmonary artery and the properties of the K+ current activated by levcromakalim in isolated myocytes. Under whole-cell voltage clamp, holding at –60 mV in symmetrical 139 mm K+, levcromakalim (10 m) induced a noisy inward current of –116 ± 19 pA (n = 13) which developed over 1 to 2 min. This current could be blocked by either glibenclamide (10 m) or phencyclidine (5–50 M) and was unaffected when extracellular Ca2+ was removed. Both these drugs inhibited the levcromakalim-induced relaxation of muscle strips precontracted with 20 mm [K+]
o
. Application of voltage ramps in symmetrical 139 mm K+ confirmed that the levcromakalim-induced current was carried by K+ ions and was weakly voltage dependent over the potential range from –100 to +40 mV.The unitary current amplitude and density of the channels underlying the levcromakalim-activated whole-cell K+ current was estimated from the noise in the current record. We estimate that levcromakalim caused activation of around 300 channels per cell, with a single channel current of 1.1 pA, corresponding to a slope conductance of about 19 pS. Furthermore, cells dialyzed with an ATP-free pipette solution developed a large noisy inward current at –60 mV, which could subsequently be blocked by flash photolysis of caged ATP. Analysis of the noise associated with this current indicated that the single channel amplitude underlying the ATP-blocked current was 1.4 pA, a value similar to that estimated for the levcromakalim-induced current. We conclude that the conductance of this ATP-sensitive channel is likely to be small under physiological conditions and that it is present at low density.We thank SmithKline & Beecham for the gift of levcromakalim, ICI Pharmaceuticals for the gift of charybdotoxin and Prof. D. Colquhoun for the noise analysis programs. We also thank Mr. R. Davey for technical assistance with tension experiments. This work was supported by the British Heart Foundation and the Wellcome Trust. L.H.C. is a Wellcome Research Fellow and P.L. is an intermediate fellow of the BHF. 相似文献
17.
18.
Summary The electrical properties of the vacuolar membrane of the primitive green algaEremosphaera virdis were investigated using the patch-clamp technique. In whole vacuole measurements two types of transport systems with long activation time-constants were identified. The first, showing marked outward rectification, was activated by an increase in the cytosolic calcium concentration. Furthermore, it displayed sensitivity to micromolar concentrations of the anion channel blocker Zn2+ and to acidification of the cytosol. In contrast, the second time-activated current component was almost insensitive to changes in cytosolic pH and was blocked by the potassium channel inhibitor TEA. In addition to these slowly activating current components, the vacuolar membrane contained at least two further transport systems, responsible for an instantaneous current. These two current components were distinguished by their different sensitivity to protons, cytosolic calcium, and TEA. Comparing these electrical properties to those observed in vacuoles of higher plants or in cytoplasmic droplets from characean algae, respectively, it seems thatEremosphaera is intermediate, corresponding to the systematic position of this simple green alga.Abbreviations [Ca2+]cyt
cytosolic free calcium concentration
- EGTA
ethyleneglycol-bis(-aminoethylether)N,N,N,N-tetraacetic acid
- HEPES
N-[2-hydroxyethyl]piperazine-N-[2-ethanesulfonic acid]
- I
electric current
- IRC
inward rectifying current
- MES
2-[N-morpholino]ethanesulfonic acid
- ORC
outward rectifying current
- pHcyt
cytosolic pH
- pHvac
vacuolar pH
- Po
open probability
- Px
permeability coefficient of ion species X
- TEA
tetraethylammonium chloride
- Tris
tris[hydroxymethyl]aminomethane
- V
voltage 相似文献
19.
Summary Plasmalemmal ionic currents from enzymatically-isolated protoplasts of suspension-cultured carrot cells were investigated by patch-clamp techniques. Among other currents, a novel hyperpolarization-activated, inwardly-rectifying, whole-cell current was observed. The activation of this current was fast in onset, and for large hyperpolarizations a characteristic, rapid voltage-dependent inactivation was seen. Ion substitution experiments indicate that this inward current was due mainly to efflux of chloride ions. No dependence on either internal or external calcium was found, and internal MgATP was not necessary. Surprisingly, zinc did not block this current. In hyperpolarized outside-out patches, inward single-channel chloride currents having an elementary conductance of ca. 100 pS were observed. The open probability increased with hyperpolarization. Similar single-channel currents were activated by slight negative pressure applied to the pipette. These chloride currents could contribute both to the control of membrane potential and in the regulation of osmotic balance in carrot cells.Abbreviations BAPTA
1,2-bis (2-aminophenoxy)ethane-N,N,N,N-tetraacetic acid
- 2,4-D
2,4-dichlorophenoxyacetic acid
- Ex
Nernst equilibrium potential for ion x
- NMDG
N-methyl-D-glucamine
- PMSF
phenylmethylsulfonyl fluoride 相似文献
20.
G. Horseman F. Huber 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1994,175(4):389-398
Intracellular recordings were made in the brain of the cricket Gryllus bimaculatus from an ascending auditory interneuron (AN1). Acoustic stimuli with calling song temporal pattern were delivered via earphones in a preparation with the acoustic trachea cut (attenuation of crossing sound > 30 dB). The input-output function of this cell was then determined by recording its responses to stimulation of the ipsilateral ear alone, of the contralateral ear alone and to stimulation of both ears simultaneously with the same or different carrier frequencies and intensities.This interneuron was excited by the ear ipsilateral to its axon and dendritic field and unresponsive to stimuli presented to the axon-contralateral ear alone. However, in binaural stimulation experiments, the response to a constant ipsilateral stimulus was progressively reduced as the intensity of a simultaneous contralateral stimulus was increased, above a threshold intensity.Tuning curves for threshold of this inhibition, determined in binaural stimulation experiments, indicated significant inhibition in the range 3–20 kHz with lowest threshold at 4–5 kHz. The inhibition was unaffected by sectioning of the contralateral circumoesophageal or neck connective, indicating that the inhibitory influence crosses the midline at the level of the prothoracic ganglion. Intracellular recordings from AN1 in the prothoracic ganglion confirmed that it was indeed neurally inhibited by inputs from the contralateral ear.Tuning curves for excitation of an omega neuron (ON1) by the ear ipsilateral to its soma and also the tuning of inhibition of ON1 by its contralateral ON1 partner, closely match the tuning of inhibition of AN1 and to a lesser extent, of AN2. This was taken as evidence that each AN1 is inhibited by the contralateral ON1. The significance of this interaction for directional hearing and phonotaxis is discussed.Abbreviations AP/CHP
action potentials per chirp
- AN1, AN2
ascending auditory interneurons 1, 2
- ON1
omega neuron 1
- ipsi
ipsilateral contra contralateral
- PTG
prothoracic ganglion loc lateral ocellar nerve
- On
optic nerve an antennal nerve
- coc
circum-oesophageal connective so sound off 相似文献