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1.
cDNA encoding the full-length hKv1.3 lymphocyte channel and a C-terminal truncated (Δ459-523) form that lacks the putative
PKA Ser468 phosphorylation site were stably transfected in human embryonic kidney (HEK) 293 cells. Immunostaining of the transfected
cells revealed a distribution at the plasma membrane that was uniform in the case of the full-length channel whereas clustering
was observed in the case of the truncated channel. Some staining within the cell cytoplasm was found in both instances, suggesting
an active process of biosynthesis. Analyses of the K+ current by the patch-clamp technique in the whole cell configuration showed that depolarizing steps to 40 mV from a holding
potential (HP) of −80 mV elicited an outward current of 2 to 10 nA. The current threshold was positive to −40 mV and the current
amplitude increased in a voltage-dependent manner. The parameters of activation were −5.7 and −9.9 mV (slope factor) and −35
mV (half activation, V
0.5) in the case of the full-length and truncated channels, respectively. The characteristics of the inactivation were 14.2 and
24.6 mV (slope factor) and −17.3 and −39.0 mV (V
0.5) for the full-length and truncated channels, respectively. The activation time constant of the full-length channel for potentials
ranging from −30 to 40 mV decreased from 18 to 12 msec whereas the inactivation time constant decreased from 6600 msec at
−30 mV to 1800 msec at 40 mV. The unit current amplitude measured in cells bathing in 140 mm KCl was 1.3 ± 0.1 pA at 40 mV, the unit conductance, 34.5 pS and the zero current voltage, 0 mV. Both forms of the channels
were inhibited by TEA, 4-AP, Ni2+ and charybdotoxin. In contrast to the native (Jurkat) lymphocyte Kv1.3 channel that is fully inhibited by PKA and PKC, the
addition of TPA resulted in 34.6 ± 7.3% and 38.7 ± 9.4% inhibition of the full-length and the truncated channels, respectively.
8-BrcAMP induced a 39.4 ± 5.4% inhibition of the full-length channel but had no effect (8.6 ± 8.3%) on the truncated channel.
Cell dialysis with alkaline phosphatase had no effects, suggesting that the decreased sensitivity of the transfected channels
to PKA and PKC was not due to an already phosphorylated channel. Patch extract experiments suggested that the hKv1.3 channel
was partially sensitive to PKA and PKC. Cotransfecting the Kvβ1.2 subunit resulted in a decrease in the value of the time
constant of inactivation of the full-length channel but did not modify its sensitivity to PKA and PKC. The cotransfected Kvβ2
subunit had no effects. Our results indicate that the hKv1.3 lymphocyte channel retains its electrophysiological characteristics
when transfected in the Kvβ-negative HEK 293 cell line but its sensitivity to modulation by PKA and PKC is significantly reduced.
Received: 18 June 1997/Revised: 7 October 1997 相似文献
2.
S. A. Fedulova 《Neurophysiology》1999,31(6):372-375
We studied the effect of 4-aminopyridine (4-AP) on the channels responsible for transient rapidly inactivating potassium A-type
current (I
A
) in the somatic membrane of cultured rat hippocampal neurons. External application of 4-AP in different concentrations (from
100 μM to 5mM) was made to the locus of the neuron under study using a fast local superfusion technique. TheI
A
blockade was dose-dependent and voltage-independent. Interaction of the blocker with theI
A
-conducting channels at a test potential of +40 mV could be approximated by Hill isotherm with the cooperativity coefficient
of 2 and EC50 equal to 2.1 mM. The action of 4-AP accelerated temporal inactivation ofI
A
. The intensity ofI
A
blockade became higher as the frequency of test membrane potential shifts increased. 相似文献
3.
E. I. Solntseva 《Neurophysiology》1993,25(6):354-358
A low-threshold outward current was studied in the neurons ofHelix pomatia at –70 to –30 mV using a two-electrode voltage clamp technique. In addition to the well-known A current (I
A), a slower outward current calledI
As (slow) was revealed. Activation and inactivation times ofI
As at –40 mV ranged from 90 to 120 msec and from 3 to 5 sec, respectively. The current recovered within 2 to 5 sec after inactivation at –120 mV. Analysis of changes in the reversal potential ofI
As caused by an increase in external potassium concentration suggests a potassium origin forI
As. The curves ofI
As stationary activation and inactivation fit the Boltzmann equation. Deriving from an activation curve, the activation potential for a half-maximum current,, is –40 mV, and the slope factor,k, is –9.8 mV, while those values for the inactivation curve are –84 mV (a half-maximum inactivation) and 7.5 mV.I
As is blocked by 4-aminopyridine (1–30 µM), tetraethylammonium (1 mM), and Ba2+ (1 mM), but is resistant to Cs+ (1 mM). PeakI
As is not affected either by substitution of external Ca2+ for Mg2+ or by application of Cd2+ (0.5–1.0 mM). The results suggest that activation ofI
As does not require Ca2+ entry into the cell.Neirofiziologiya/Neurophysiology, Vol. 25, No. 6, pp. 427–432, November–December, 1993. 相似文献
4.
Single-channel properties of a delayed rectifier voltage-gated K+ channel (I-type) were investigated in peripheral myelinated axons from Xenopus laevis. Channels activated between −60 and −40 mV with a potential of half-maximal activation, E50, at −47.5 mV. Averaged single-channel currents activated with a time delay at all membrane potentials tested. Time to half-maximal
activation decreased from 80 to 1.6 msec between −60 and +40 mV. The channel inactivated monoexponentially with a time constant
of 10.9 sec at −40 mV. The time constant of deactivation was 126 msec at −80 mV and 16.9 msec at −110 mV. In symmetrical 105
mm K+, the single-channel conductance (γ) was 22 and 13 pS at negative and positive membrane potentials, respectively, at 13–15°C.
In Na+-rich solution with 2.5 mm extracellular K+γ was 7 pS and the reversal potential was negative to −80 mV, indicating a high selectivity for K+ over Na+. γ depended on extracellular K+ concentration (K
D
= 19.6 mm) and temperature (Q
10= 1.45). External tetraethylammonium (TEA) reduced the apparent single-channel current amplitude at all potentials tested
with a half-maximal inhibiting concentration (IC50) of 0.6 mm. Open probability of the channel, but not single-channel current amplitude was decreased by extracellular dendrotoxin (DTX,
IC50= 6.8 nm) and mast cell degranulating peptide (MCDP, IC50= 41.9 nm). In Ringer solution the membrane potential of macroscopic I-channel patches was about −65 mV and depolarized under TEA and
DTX. It is concluded that besides their activation during action potentials, I-channels may also stabilize the resting membrane
potential.
Received: 2 June 1995/Revised: 13 October 1995 相似文献
5.
Voltage-clamp experiments were performed on single bovine adrenal fasciculata cells in short-term primary culture using either
standard (broken membrane) or perforated whole-cell patch clamp recording. The membrane current measured with the perforated
method was dominated by a very stable transient outward current. By contrast, the transient outward current recorded using
the standard method was unstable. The reversal potential of the transient outward current varied linearly with the logarithm
of [K+]
e
with a slope of 47 mV per decade. The onset of activation was sigmoidal and was fitted with a power function where n= 4. Time constants ranged from 1 to 4 msec with a maximum at −25 mV. The steady-state activation curve spanned the voltage
range −50 to +80 mV without reaching a clear maximum. During a pulse, the current decayed in a biexponential manner. Time
constants τ1 and τ2 were voltage-dependent and ranged from 50 to 200 msec respectively for a voltage step at +50 mV. The steady-state inactivation
was dependent on the conditioning pulse duration. Using short conditioning pulses (1.2 sec), the curve which spanned the voltage
range −40 to −20 mV, was 15 mV more positive than that obtained with longer conditioning pulses (60 sec). Time constants of
this ``very slow inactivation' process (τvs) determined for voltage steps at −60 and −50 mV were 15 and 10 sec respectively. A ``facilitation process' of the peak current
was observed when the duration or the amplitude of conditioning pulses were increased in the voltage range −100 to −50 mV.
Recovery from inactivation followed a biexponential time course which seemed a mixture of both inactivation processes. In
some experimental conditions, isolated cells were able to produce overshooting action potentials. These results are discussed
in relation with the membrane electrogenesis of this cell type.
Received: 14 November 1994/Revised: 24 October 1995 相似文献
6.
V. A. Bouryi 《Neurophysiology》1998,30(4-5):301-304
Barium currents through ion channels formed by α1-subunit of L-type Ca2+ channel (I
α1) were recorded from cultured chinese hamster ovary (CHO) cells. The cells were stably transfected with either a cardiac or
a smooth muscle (SM) variant of α1-subunit. TheI
α1 in both cases exhibited similar fast voltage-dependent activation kinetics and slow apparent inactivation kinetics. With
10 mM Ba2+ in the bath solution,I
α1 was activated at potentials more positive than −40 mV, peaked between 0 and +10 mV, and reversed at about +50 mV. In addition
to slow apparent inactivation of inward current, both subunits provided an extremely slow voltage-dependent inactivation at
potentials more positive than −100 mV, with half-maximum inactivation at −43.4 mV for cardiac and −41.4 mV for SM α1-subunits.
The onset of inactivation as well as recovery from this process were within a time range of minutes. The voltage dependence
of steady-state inactivation could be fitted by the sum of two Boltzmann's equations with slope factors of about 12 mV and
5 mV. A less sloped component has its midpoints at −75.6 and −63.7 mV, and a steeper component has its midpoints at −42.8
and −37.7 mV for cardiac and SM α1-subunits, respectively. Relative contribution of the steeper component was higher in both
subunits (0.86 and 0.66 for cardiac and SM subunits, respectively). For comparison, the inactivation curves for 5-sec-long
conditioning prepulses could be fitted by single Boltzmann's distribution with a 20 mV more positive midpoint and a slope
factor of about 13 mV. In contrast to the steady-state inactivation curves, they showed considerable overlap with the steady-state
activation curve. Our results reflect functional consequences of known sequence differences between α1-subunits of the cardiac
and SM L-type Ca2+ channels and could be used in structural modeling of Ca2+ channel gating. In addition, they show that depolarization-induced window current has a transient nature and decays with
the development of extremely slow inactivation. This is the first demonstration that slow inactivation of the L-type Ca2+ channel is an intrinsic property of its α1-subunits. 相似文献
7.
Guyot A Dupré-Aucouturier S Ojeda C Rougier O Bilbaut A 《The Journal of membrane biology》2000,173(2):149-163
Voltage-activated Ca2+ currents, in zona fasciculata cells isolated from calf adrenal gland, were characterized using perforated patch-clamp recording.
In control solution (Ca2+: 2.5 mm) a transient inward current was followed, in 40% of the cells, by a sustained one. In 20 mm Ba2+, 61% of the cells displayed an inward current, which consisted of transient and sustained components. The other cells produced
either a sustained or a transient inward current. These different patterns were dependent upon time in culture. Current-voltage
relationships show that both the transient and sustained components activated, peaked and reversed at similar potentials:
−40, 0 and +60 mV, respectively. The two components, fully inactivated at −10 mV, were separated by double-pulse protocols
from different holding potentials where the transient component could be inactivated or reactivated. The decaying phase of
the sustained component was fitted by a double exponential (time constants: 1.9 and 20 sec at +10 mV); that of the transient
component was fitted by a single exponential (time constant: 19 msec at +10 mV). Steady-state activation and inactivation
curves of the two components were superimposed. Their half activation and inactivation potentials were similar, about −15
and −34 mV, respectively. The sustained component was larger in Ba2+ than in Sr2+ and Ca2+. Ni2+ (20 μm) selectively blocked the transient component while Cd2+ (10 μm) selectively blocked the sustained one. (±)Bay K 8644 (0.5 μm) increased the sustained component and nitrendipine (0.5–1 μm) blocked it selectively. The sustained component was inhibited by calciseptine (1 μm). Both components were unaffected by ω-conotoxin GVIA and MVIIC (0.5 μm). These results show that two distinct populations of Ca2+ channels coexist in this cell type. Although the voltage dependence of their activation and inactivation are comparable,
these two components of the inward current are similar to T- and L-type currents described in other cells.
Received: 12 July 1999/Revised: 5 October 1999 相似文献
8.
Whole-cell patch-clamp techniques were used to measure anion currents through the plasma membrane of protoplasts of mesophyll
cells of expanding pea (Pisum sativum L.) leaves. Voltage-induced changes of the currents could be modelled with single exponential activation and deactivation
kinetics. The anion currents were activated at negative membrane potentials. The time constant of activation, τact, increased from 145 ms at −140 mV to 380 ms at −20 mV. A Boltzmann fit to the activation curve, n∞ (ΔGVm/ΔGmax), yielded a half-activation voltage of +27 mV. Opening and closing rate constants, α and β respectively, were calculated
from the values of τ and n∞. The currents depended on the presence of cytoplasmic Ca2+ concentrations higher than 10−6 M. Including 3 mM MgATP in the intracellular solution resulted in a voltage-dependent inactivation of the anion current.
The conductance-voltage relation resulting from the voltage-dependent activation and inactivation had a maximum at about −25
mV. The relations of the current in pea are discussed with respect to the anion currents in guard cells and suspension-cultured
tobacco cells, and its possible role in growing leaf cells.
Received: 1 March 1996 / Accepted: 16 September 1996 相似文献
9.
The responses of freshly isolated hippocampal pyramidal neurons to rapid, elevations of the external potassium concentration
([K+]
out
) were investigated using the whole-cell variation of a patch-clamp technique. An elevation of [K+]
out
induced a two-phase inward current at the membrane potentials more negative than the reversal potential for K ions. This
current consisted of a leakage, current and a time-dependent current (τ=40–50 msec at 21°C), the latter designated below asI
ΔK. It displayed first-order activation kinetics that showed neither voltage, nor concentration dependence. The amplitude of
this current was determined by the external K+ concentration and increased with hyperpolarization. Voltage dependence ofI
ΔK measured within the range from −20 to −120 mV was similar to that for inward rectifier. Activation ofI
ΔK was utterly dependent on Na+; substitution of extracellular Na+ with choline chloride almost completely depressedI
ΔK.I
ΔK was absent in the cells freshly dissociated from the nodosal and dorsal root ganglia. This suggests that this earlier unrecognized
current is instrumental in preserving densely packed hippocampal pyramidal neurons from sudden increases in [K+]
out
and following spontaneous over-excitation. It prevents the neurons from responding to K+-induced depolarizations by slowing down potassium influx. 相似文献
10.
Cook GM 《Extremophiles : life under extreme conditions》2000,4(5):279-284
The thermophilic glycolytic anaerobe Thermoanaerobacter wiegelii grows over the pH range 5.1–7.7, and no growth is observed below pH 5.0 or above 7.7. When T. wiegelii was grown in pH-uncontrolled batch culture, glucose was fermented to ethanol, acetate, and lactate. Small amounts of lactic
acid were produced once the external pH reached 6.0, and a fructose-1,6-diphosphate (FDP) activated lactate dehydrogenase
(LDH) was detected in cell-free crude extracts. Maximal activation of LDH by FDP was observed at pH 6.2. As the pH of the
medium declined from 6.7 to 5.1 due to the production of acetate and lactate, the total protonmotive force (Δp) remained between
−110 and −130 mV, and the membrane potential (ΔΨ) decreased from −104 to −65 mV. This decrease in ΔΨ was paralleled by an
increase in the chemical gradient of protons (ZΔpH) from −31 to −62 mV at pH 5.1. Based on these results, T. wiegelii maintained a small ΔpH (0.3–0.9 units, inside alkaline) as the medium pH declined and interconverted ΔΨ to ZΔpH to maintain
the total Δp relatively constant. Intracellular potassium decreased from 150 mM at pH 6.70 to 50 mM at pH 5.1, and this represented
a 33-mV decline in the transmembrane chemical potential of potassium. The ability to synthesize ATP remained constant as the
external pH declined, and therefore metabolic energy per se was not a critical aspect of pH sensitivity.
Received: March 16, 2000 / Accepted: May 8, 2000 相似文献
11.
Pyrethroid modulation of sodium channels is unique in the sense that it is highly dependent on temperature, the potency being
augmented by lowering the temperature. To elucidate the mechanisms underlying the negative temperature dependence of pyrethroid
action, single sodium channel currents were recorded from cultured rat hippocampal neurons using the inside-out configuration
of patch-clamp technique, and the effects of the pyrethroid tetramethrin were compared at 22 and 12°C. Tetramethrin-modified
sodium channels opened with short closures and/or transitions to subconductance levels at 22 and 12°C. The time constants
of the burst length histograms for tetramethrin-modified channels upon depolarization to −60 mV were 7.69 and 14.46 msec at
22 and 12°C, respectively (Q10= 0.53). Tetramethrin at 10 μm modified 17 and 23% of channels at 22 and 12°C, respectively, indicating that the sensitivity of the sodium channel of rat
hippocampal neurons to tetramethrin was almost the same as that of tetrodotoxin-sensitive sodium channels of rat dorsal root
ganglion neurons and rat cerebellar Purkinje neurons. The time constants for burst length in tetramethrin-modified sodium
channels upon repolarization to −100 mV from −30 mV were 8.26 and 68.80 msec at 22 and 12°C (Q10= 0.12), respectively. The prolongation of tetramethrin-modified whole-cell sodium tail currents upon repolarization at lower
temperature was ascribed to a prolongation of opening of each channel. Simple state models were introduced to interpret behaviors
of tetramethrin-modified sodium channels. The Q10 values for transition rate constants upon repolarization were extremely large, indicating that temperature had a profound
effect on tetramethrin-modified sodium channels.
Received: 31 January 2000/Revised: 18 May 2000 相似文献
12.
成年蜜蜂脑神经细胞的培养和电生理特征 总被引:1,自引:0,他引:1
为了研究杀虫剂等对蜜蜂毒性作用的神经机制,需在体外建立成年蜜蜂脑神经细胞的分离培养和电生理记录技术并研究其正常电生理特征,而对成年蜜蜂脑神经细胞的分离培养和电生理特性的研究报道甚少。我们采用酶解和机械吹打相结合的方法获得了数量较多且活力较好的成年意大利蜜蜂Apis mellifera脑神经细胞,并用全细胞膜片钳技术研究了成年意大利蜜蜂脑神经细胞对电流和电压刺激的反应,获得了成年意蜂脑神经细胞的基本电生理特征以及钠电流和钾电流的特性。全细胞电流钳的记录结果表明,在体外培养条件下,细胞无自发放电发生,注射电流后仅引起细胞单次放电,引起细胞放电的阈电流平均为60.8±63 pA; 细胞动作电位产生的阈电位平均为−27.4±2.3 mV。用全细胞电压钳记录了神经细胞的钠电流和钾电流。钠电流的分离是在电压刺激下通过阻断钾通道和钙通道实现。细胞的内向钠电流在指令电压为−40~−30 mV左右激活,−10 mV达峰值,钠通道的稳态失活电压V1/2为−58.4 mV; 外向钾电流成份至少包括较小的快速失活钾电流和和较大的缓慢失活钾电流(占总钾电流的80%),其半激活膜电位V1/2为3.86 mV,无明显的稳态失活。结果提示缓慢失活钾电流的特征可能是细胞单次放电的机制之一。 相似文献
13.
Transient outward currents were characterized with twin electrode voltage clamp techniques in isolated F76 and D1 neuronal
membranes (soma only) of Helix aspersa subesophageal ganglia. In this study, in addition to the transient outward current (A-current, I
A
) described by Connor and Stevens (1971b), another fast outward current, referred to as I
Adepol
here, is described for the first time. This is similar to the current component characterized in Aplysia (Furukawa, Kandel & Pfaffinger, 1992). The separation of these two current components was based on activation and steady-state
inactivation curves, holding potentials and sensitivity to 4-aminopyridine (4-AP). In contrast to I
A
, I
Adepol
did not require hyperpolarizing conditioning pulses to remove inactivation; it was evoked from a holding potential of −40
mV, at which I
A
is completely inactivated. I
Adepol
shows noticeable activation at around −5 mV, whereas I
A
activates at around −50 mV. The time courses of I
Adepol
activation and inactivation were similar but slower than I
A
. It was found that I
Adepol
was more sensitive than I
A
to 4-AP. 4-AP at a concentration of 1 mm blocked I
Adepol
completely, whereas 5–6 mm 4-AP was needed to block I
A
completely. This current is potentially very important because it may, like other A currents, regulate firing frequency but
notably, it does not require a period of hyperpolarization to be active.
Received: 12 May 2000/Revised: 12 October 2000 相似文献
14.
Voltage-dependent calcium and calcium-activated potassium currents of a molluscan photoreceptor 总被引:5,自引:1,他引:4
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Two-microelectrode voltage clamp studies were performed on the somata of Hermissenda Type B photoreceptors that had been isolated by axotomy from all synaptic interaction as well as any impulse-generating (i.e., active) membrane. In the presence of 2-10 mM 4-aminopyridine (4-AP) and 100 mM tetraethylammonium ion (TEA), which eliminated two previously described voltage-dependent potassium currents (IA and the delayed rectifier), a voltage-dependent outward current was apparent in the steady state responses to command voltage steps more positive than -40 mV (absolute). This current increased with increasing external Ca++. The magnitude of the outward current decreased and an inward current became apparent following EGTA injection. Substitution of external Ba++ for Ca++ also made the inward current more apparent. This inward current, which was almost eliminated after being exposed for approximately 5 min to a solution in which external Ca++ was replaced with Cd++, was maximally activated at approximately 0 mV. Elevation of external potassium allowed the calcium (ICa++) and calcium-dependent K+ (IC) currents to be substantially separated. Command pulses to 0 mV elicited maximal ICa++ but no IC because no K+ currents flowed at their new reversal potential (0 mV) in 300 mM K+. At a holding potential of -60 mV, which was now more negative than the potassium equilibrium potential, EK+, in 300 mM K+, IC appeared as an inward tail current after positive command steps. The voltage dependence of ICa++ was demonstrated with positive steps in 100 mM Ba++, 4-AP, and TEA. Other data indicated that in 10 mM Ca++, IC underwent pronounced and prolonged inactivation whereas ICa++ did not. When the photoreceptor was stimulated with a light step (with the membrane potential held at -60 mV), there was also a prolonged inactivation of IC. In elevated external Ca++, ICa++ also showed similar inactivation. These data suggest that IC may undergo prolonged inactivation due to a direct effect of elevated intracellular Ca++, as was previously shown for a voltage-dependent potassium current, IA. These results are discussed in relation to the production of training-induced changes of membrane currents on retention days of associative learning. 相似文献
15.
Transmembrane ion currents were studied in the somatic membrane of freshly isolated neurons from the spinal ganglia of early
postnatal (younger than 15-day-old) rats. According to their dissimilar voltage dependence and different sensitivity to external
application of tetraethylammonium (TEA) and 4-aminopyridine (4-AP), three types of outward potassium currents were identified.
Fast-inactivating K+ current was activated at the most negative values of the membrane potential and showed the highest sensitivity to external
application of 4-AP. The threshold for activation of slow-inactivating K+ current was within a −40 ... −30 mV range. Non-inactivating delay-rectified current showed the highest sensitivity to TEA.
All three types of K+ currents could be found in all studied neurons of animals of three age groups: 1, 5 to 6, and 14 to 15 postnatal days. The
mean density of fast-inactivating K+ current significantly increased during the first two weeks of postnatal ontogenesis. Within the studied period, the mode
of a normal (Gaussian) distribution of fast K+ current shifted toward higher current density values. The mean density of slow-inactivating K+ current also increased with the age. Yet, the mean density of non-inactivating delay-rectified K+ current significantly dropped during the first five days of the postnatal development and remained stable during the following
time interval. 相似文献
16.
《The Journal of general physiology》1993,101(4):603-626
Block of the calcium-independent transient outward K+ current, I(to), by 4-aminopyridine (4-AP) was studied in ferret right ventricular myocytes using the whole cell patch clamp technique. 4-AP reduces I(to) through a closed state blocking mechanism displaying "reverse use- dependent" behavior that was inferred from: (a) development of tonic block at hyperpolarized potentials; (b) inhibition of development of tonic block at depolarized potentials; (c) appearance of "crossover phenomena" in which the peak current is delayed in the presence of 4-AP at depolarized potentials; (d) relief of block at depolarized potentials which is concentration dependent and parallels steady-state inactivation for low 4-AP concentrations (V1/2 approximately -10 mV in 0.1 mM 4-AP) and steady-state activation at higher concentrations (V1/2 = +7 mV in 1 mM 4-AP, +15 mV in 10 mM 4-AP); and (e) reassociation of 4- AP at hyperpolarized potentials. No evidence for interaction of 4-AP with either the open or inactivated state of the I(to) channel was obtained from measurements of kinetics of recovery and deactivation in the presence of 0.5-1.0 mM 4-AP. At hyperpolarized potentials (-30 to - 90 mV) 10 mM 4-AP associates slowly (time constants ranging from approximately 800 to 1,300 ms) with the closed states of the channel (apparent Kd approximately 0.2 mM). From -90 to -20 mV the affinity of the I(to) channel for 4-AP appears to be voltage insensitive; however, at depolarized potentials (+20 to +100 mV) 4-AP dissociates with time constants ranging from approximately 350 to 150 ms. Consequently, the properties of 4-AP binding to the I(to) channel undergo a transition in the range of potentials over which channel activation and inactivation occurs (-30 to +20 mV). We propose a closed state model of I(to) channel gating and 4-AP binding kinetics, in which 4-AP binds to three closed states. In this model 4-AP has a progressively lower affinity as the channel approaches the open state, but has no intrinsic voltage dependence of binding. 相似文献
17.
A study of the effects of (+)-methysticin on sodium channels of isolated neurons from theCA1 hippocampal area of rats showed that this drug considerably accelerabed the inactivation processes providing that the membrane
potential was within the −80 to −20 mV range. When depolarization was further increased, the influence of methysticin on the
rate of inactivation became much smaller. Hyperpolarization of the membrane removed inactivation. Methysticin (100 μM) intensively
slowed down deinactivation and decreased the dependence of the rate of this process on the membrane potential. The effect
of methysticin on inactivation kinetics results in considerable shifts of the neurons' excitability; this probably underlies
neuroprotective effects of this drug manifested in the model of focal cerebral ischemia in rats and mice. 相似文献
18.
Properties of transient K+ currents and underlying single K+ channels in rat olfactory receptor neurons 总被引:5,自引:2,他引:3
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The transient potassium current, IK(t), of enzymatically dissociated rat olfactory receptor neurons was studied using patch-clamp techniques. Upon depolarization from negative holding potentials, IK(t) activated rapidly and then inactivated with a time course described by the sum of two exponential components with time constants of 22.4 and 143 ms. Single-channel analysis revealed a further small component with a time constant of several seconds. Steady-state inactivation was complete at -20 mV and completely removed at -80 mV (midpoint -45 mV). Activation was significant at -40 mV and appeared to reach a maximum conductance at +40 mV (midpoint -13 mV). Deactivation was described by the sum of two voltage-dependent exponential components. Recovery from inactivation was extraordinarily slow (50 s at -100 mV) and the underlying processes appeared complex. IK(t) was reduced by 4-aminopyridine and tetraethylammonium applied externally. Increasing the external K+ concentration ([K+]o) from 5 to 25 mM partially removed IK(t) inactivation, usually without affecting activation kinetics. The elevated [K+]o also hyperpolarized the steady-state inactivation curve by 9 mV and significantly depolarized the voltage dependence of activation. Single transient K+ channels, with conductances of 17 and 26 pS, were observed in excised patches and often appeared to be localized into large clusters. These channels were similar to IK(t) in their kinetic, pharmacological, and voltage-dependent properties and their inactivation was also subject to modulation by [K+]o. The properties of IK(t) imply a role in action potential repolarization and suggest it may also be important in modulating spike parameters during neuronal burst firing. A simple method is also presented to correct for errors in the measurement of whole-cell resistance (Ro) that can result when patch-clamping very small cells. The analysis revealed a mean corrected Ro of 26 G omega for these cells. 相似文献
19.
The effects of blockers of voltage-gated potassium channels, tetraethylammonium (TEA) and 4-aminopyridine (4-AP), on inhibitory
postsynaptic currents (IPSC) evoked by local electrical stimulation of zones of unitary synaptic terminals on hippocampal
neurons were studied using a voltage-clamp technique under conditions of low density cell culture. At activation of the transmitter
release in the absence of action potentials (when the terminals are in a tetrodotoxin-containing medium), external application
of 5 mM 4-AP reversibly increased the averaged IPSC amplitude by 90±30%, while a similar effect of 10 mM TEA reached only
20±7%. The amplitudes of individual evoked IPSC varied between 10 and more than 150 pA. Amplitude histograms of IPSC in all
studied neurons (n=14) were of a polymodal nature and could not described by a Gaussian law. An increase in the averaged IPSC amplitude under
the influence of potassium channel blockers cannot be described as resulting only from modification of the number of trials
without transmitter release (blank events). The mechanism of potassium channel blocker-induced facilitation of IPSC evoked
by single synaptic terminals is discussed. 相似文献
20.
Two new types of calcium channels were discovered during research in ionic currents in the somatic membrane ofHelix pomatia neurons, using an intracellular perfusion technique. Apart from the principal calcium current described in the literature with a holding potential of about –110 mV, an additional calcium current was observed activated at depolarizations of –40 to –80 mV and was not reduced when the cell was perfused with solutions containing fluoride anions. The kinetics of this current were well described in the context of the Hodgkin and Huxley model with a time constant of activation of 6–8 msec and of inactivation of 300–600 msec. It increased in amplitude as the Ca++ rose in the cellular environment but was reduced by extracellular addition of the Ca++ antagonists Co++, Ni++, and Cd++, and the organic blockers nifedipine and verapamil. The association constants of these substances with corresponding channels determined from the maximum of the current-voltage relationship were 2 (Ca++), 3 (Co++), 0.06 (nifedipine), and 0.2 mM (verapamil). The properties detected in this component of calcium conductance are compared with those of calcium channels in other excitatory formations and its possible functional role is discussed.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 17, No. 5, pp. 627–633, September–October, 1985. 相似文献