Voltage Sensitive Ca-Channels and the Transient Inward Current in Paramecium Tetraurelia

1979 ◽  
Vol 78 (1) ◽  
pp. 149-161 ◽  
Author(s):  
YOUKO SATOW ◽  
CHING KUNG
Keyword(s):  
Voltage Clamp ◽  
Resting Potential ◽  
Ca Channel ◽  
Paramecium Tetraurelia ◽  
Ca Channels ◽  
Action Current ◽  
Inward Current ◽  
Transient Inward Current ◽  
Inorganic Cation ◽  
Inward Currents

Transient inward currents across the membrane of P. tetraurelia are recorded upon step depolarizations with a voltage clamp in solutions where Ca2+ is the only added inorganic cation. It is shown that the current is normally carried by Ca2+ through the Ca-channels which activate and inactivate in time. The transient inward current is dependent on both the size of the depolarizing step and the holding level before the step. Maximum inward current (Imax) occurs when the membrane is first held at the resting level (- 30 mV), then stepped to 0 mV in a solution containing 0.91 mM-Ca2+. The Imax is smaller when the membrane is first held at depolarized level. This is due to the depolarization-sensitive inactivation of the Ca-channels. The Imax is also smaller when the membrane is first held at a hyperpolarized level. This may be explained by the activation of hyperpolarization-sensitive K-channels known to exist in the Paramecium membrane. I max increases with concentration of Ca2+ up to 0.9 mM. Further increase in the Ca2+ concentration does not affect Imax. This apparent saturation at 0.9 mM-Ca2+ may reflect a rate-limiting step of Ca2+ permeation. The increase in Ca2+ concentration shifts the V-Ipeak curve in the direction of less sensitivity. This result is best explained as the effect of bound Ca2+ on the surface potential of the Paramecium membrane. These results provide the first detailed description of the properties of the action current through the Ca-channel in Paramecium. They also define the conditions under which future voltage-clamp studies of wild-type and mutant membranes of P. tetraurelia should be performed, i.e. to maximize the resolution of the Ca-channel activity, the membrane should be held at or near the resting potential and there should be over 0.9 mM-Ca2+ in the test solutions. The behaviour of the Paramecium Ca-channel and small Imax in the presence of K+ are discussed.

Ca-Induced K+-Outward Current in Paramecium Tetraurelia

1980 ◽  
Vol 88 (1) ◽  
pp. 293-304 ◽  
Author(s):  
YOUKO SATOW ◽  
CHING KUNG
Keyword(s):  
Steady State ◽  
Voltage Clamp ◽  
Outward Current ◽  
Paramecium Tetraurelia ◽  
Ca Channels ◽  
Outward Currents ◽  
Ciliary Reversal ◽  
Voltage Dependent ◽  
K Current ◽  
K Outward Current

Late K-outward currents upon membrane depolarization were recorded in Paramecium tetraurelia under a voltage clamp. A Ca-induced K-outward component is demonstrated by subtracting the value of the outward current in a pawn A mutant lacking functional Ca-channels (pwA500). The Ca-induced K-outward current activates slowly, reaching a peak after 100 to 1000 ms. The current then remains steady or reaches the steady state after a decline of several seconds. EGTA2- injection experiments show that the Ca-induced K-outward current is dependent on the internal Ca2+ concentration. The current is shown to depend on the voltage-dependent Ca conductance, by study of the leaky pawn A mutant (pwA132), which has a lowered Ca conductance as well as a lowered Ca-induced K-current. The Ca-induced GK is thus indirectly dependent on the voltage. The maximal GK is about 40 nmho/cell at + 7 mV in 4 mM-K+. The Ca-induced K current is sustained throughout the prolonged depolarization and the prolonged ciliary reversal.

Voltage-dependent calcium current and the effects of adrenergic modulation in rat aortic smooth muscle cells

1992 ◽  
Vol 337 (1279) ◽  
pp. 37-47 ◽  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Ca Channel ◽  
Ca Current ◽  
Inward Current ◽  
Aortic Smooth Muscle Cells ◽  
Aortic Smooth Muscle ◽  
Inward Currents ◽  
Sustained Inward Current

Smooth muscle cells from rat aorta were cultured in defined, serum-free medium and studied using whole-cell patch-clamp techniques. Under conditions designed to isolate currents through Ca channels, step depolarizations produced inward currents which were fast in onset and inactivated rapidly, with little sustained inward current being observed. Both Ni and Cd blocked these currents, with Ni being effective at 50 μM. Removal of external Na or addition of 1 μM tetrodotoxin had no effect. Peak inward currents were attained at about —15 mV, with half-maximal activation at —41 mV using —80 mV holding potentials. The transient inward currents were reduced by depolarized holding potentials, with half-maximal steady-state inactivation at —48 mV. In three of the 98 cells studied, small maintained inward currents were observed with a —40 mV holding potential. The Ca channel antagonist nicardipine (5 μM ) blocked the transient inward current while neither of the dihydropyridine Ca channel agonists S( + )202 791 and ( — )BAY K 8644 produced a significant augmentation of sustained inward current. At 10 μM, both noradrenaline and adrenaline but not phenylephrine decreased the peak inward current. This inhibition was unaffected by a variety of adrenoceptor antagonists and was also observed when internal solutions having high Ca buffering capacity were used, but was absent when GDP-β-S instead of GTP was included in the pipette solution. The main conclusions from this study are that under our cell culture conditions, rat aortic smooth muscle cells possess predominately a transient, low-threshold-activated inward Ca current and that this Ca current is inhibited by certain adrenoceptor agonists but with a quite atypical adrenoceptor antagonist pharmacology.

Contribution of Ca and Ca-activated Cl channels to regenerative depolarization and membrane bistability of cone photoreceptors

1992 ◽  
Vol 68 (3) ◽  
pp. 745-755 ◽  
Author(s):  
S. Barnes ◽  
M. C. Deschenes
Keyword(s):  
Voltage Clamp ◽  
Current Injection ◽  
Resting Potential ◽  
Negative Slope ◽  
Equilibrium Potential ◽  
Ca Channels ◽  
Cone Photoreceptors ◽  
Membrane Hyperpolarization ◽  
Cl Channels

1. Cone photoreceptors in several vertebrate species generate Ca-dependent regenerative depolarizations (e.g., Ca spikes lasting up to 2 s) in response to current injection or surround illumination and may remain in a state of prolonged depolarization (e.g., a permanent plateau near 0 mV) after these stimuli. This paper, while confirming the role of Ca channels in the regenerative depolarization, demonstrates that Ca-activated Cl channels either enhance or hinder prolonged depolarization, depending on the value of the chloride equilibrium potential (ECl). 2. Current- and voltage-clamp recordings obtained with the whole-cell patch-clamp technique were compared in 158 isolated tiger salamander cones to determine the contribution of specific ion channel types to the two forms of depolarizing response. Cones dialyzed with CsCl or KCl intracellular solution (such that ECl = 0 mV) that had sustained negative slope regions in their current-voltage (I-V) relations recorded under voltage clamp, were, under current clamp, bistable with respect to their resting potential. Injection of approximately 20-pA steps of depolarizing current resulted in transitions from the negative stable membrane potential (near -50 mV) to a long lasting plateau around 0 mV. Injection of 200–300 pA of hyperpolarizing current could then force a return to the negative stable resting potential, although once repolarization occurred, current injection had to be reduced or terminated to prevent damaging hyperpolarization of the cell. 3. The inward currents accounting for the negative slope region of the I-V relation were carried in Ca and Ca-activated Cl channels. Specific block of Ca-activated Cl current (ICl(Ca)) by 100 microM niflumic acid (NFA) eliminated the prolonged depolarization, even though the negative slope conductance region in the I-V persisted and the cone could still produce the briefer Ca-dependent regenerative depolarizations. Application of 100 microM Cd2+ blocked both forms of depolarization. 4. Substitution of Ba2+, which among other actions did not activate ICl(Ca), usually supported regenerative depolarizations of shortened duration, demonstrating the role of Ca channels in the initial phase of these responses. 5. A difference was observed in the regenerative depolarization when ECl was shifted away from 0 mV, where it had been in the experiments described above. With ECl set to -40 or -60 mV by reduction of [Cl-] in the pipette, steady-state membrane bistability was eliminated and prolonged depolarization did not occur. Under these conditions, application of the Cl channel blocker NFA showed that ICl(Ca) contributes to membrane hyperpolarization.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Relationship between the transient inward current and slow inward currents in the sino-atrial node of the rabbit.

1986 ◽  
Vol 370 (1) ◽  
pp. 299-315 ◽  
Author(s):  
H F Brown ◽  
D Noble ◽  
S J Noble ◽  
A I Taupignon
Keyword(s):  
Inward Current ◽  
Transient Inward Current ◽  
Inward Currents

Involvement of ryanodine receptors in muscarinic receptor-mediated membrane current oscillation in urinary bladder smooth muscle

2005 ◽  
Vol 288 (1) ◽  
pp. C100-C108 ◽  
Author(s):  
Shunichi Kajioka ◽  
Shinsuke Nakayama ◽  
Haruhiko Asano ◽  
Alison F. Brading
Keyword(s):  
Urinary Bladder ◽  
Extracellular Space ◽  
Ryanodine Receptors ◽  
Current Oscillation ◽  
Bladder Pressure ◽  
Bathing Solution ◽  
Inward Current ◽  
Transient Inward Current ◽  
Patch Pipette ◽  
Inward Currents

The urinary bladder pressure during micturition consists of two components: an initial, phasic component and a subsequent, sustained component. To investigate the excitation mechanisms underlying the sustained pressure, we recorded from membranes of isolated detrusor cells from the pig, which can be used as a model for human micturition. Parasympathomimetic agents promptly evoke a large transient inward current, and subsequently during its continuous presence, oscillating inward currents of relatively small amplitudes are observed. The two types of inward current are considered to cause the phasic and sustained pressure rises, respectively. Ionic substitution and applications of channel blockers revealed that Ca2+-activated Cl− channels were responsible for the large transient and oscillating inward currents. Furthermore, the inclusion of guanosine 5′- O-(2-thiodiphosphate) in the patch pipette indicates that both inward currents involve G proteins. However, applications of heparin in the patch pipette and of xestospongin C in the bathing solution suggest a signaling pathway other than inositol 1,4,5-trisphosphate (IP3) operating in the inward current oscillations, unlike the initial transient inward current. This IP3-independent inward current oscillation system required both sustained Ca2+ influx from the extracellular space and Ca2+ release from the intracellular stores. These two requirements are presumably SKF-96365-sensitive cation channels and ryanodine receptors, respectively. Experiments with various Ca2+ concentrations suggested that Ca2+ influx from the extracellular space plays a major role in pacing the oscillatory rhythm. The fact that distinct mechanisms underlie the two types of inward current may help in development of clinical treatments of, for example, urinary incontinence and residual urine volume control.

Membrane currents of pawn mutants of the pwA group in Paramecium tetraurelia

1980 ◽  
Vol 84 (1) ◽  
pp. 57-71 ◽  
Author(s):  
Y. Satow ◽  
C. Kung
Keyword(s):  
Time Course ◽  
Membrane Currents ◽  
Voltage Sensitivity ◽  
Paramecium Tetraurelia ◽  
Ca Channels ◽  
Wild Type ◽  
Outward Currents ◽  
In The Wild ◽  
Inward Currents ◽  
K Current

Membrane currents were recorded from the wild type and two pawn mutants of the pwA complementation group in Paramecium tetraurelia under a voltage clamp. Most currents are not changed by the mutations. Transient inward currents of a leaky mutant, pwA132, upon step depolarizations are less than those in the wild type. The inward transient is completely lacking in a non-leaky mutant, pwA500. The time course of the residual inward currents in the leaky mutant is not significantly different from that of wild type. The voltage sensitivity of the Ca channels in the leaky mutant is also similar to that of wild type. The inward currents upon membrane hyperpolarizations in the mutants show normal characteristics in the presence or absence of external K+. With sufficiently large, prolonged depolarization, outward currents progressively develop in the wild type but decay in the mutants. The simplest conclusion we can draw is that the pwA mutations reduce the number of functional Ca channels but do not change the channel characteristics. From the conductance measurements, 45% of the Ca channels remain in the leaky mutant pwA132, and none remain in the non-leaky mutant pwA500. By subtracting the outward currents of pwA500 from the slow and prolonged outward currents of the wild type, we have tentatively separated a Ca-induced K+ current from the voltage-dependent K+ current. The time courses of these two currents differ by two orders of magnitude.

A Ca-Induced Na-Current in Paramecium

1980 ◽  
Vol 88 (1) ◽  
pp. 305-326
Author(s):  
YOSHIRO SAIMI ◽  
CHING KUNG
Keyword(s):  
Voltage Clamp ◽  
Time Constant ◽  
Negative Resistance ◽  
Paramecium Tetraurelia ◽  
Wild Type ◽  
Na Current ◽  
Inward Current ◽  
In The Wild ◽  
Current Flows ◽  
Sustained Inward Current

Under a voltage clamp, step depolarization and repolarization can induce a sustained inward current and a tail inward current in Paramecium tetraurelia bathed in a solution containing 8 mM-Na+. These currents are best seen in the ‘paranoiac’ mutant. The I-V plot of the sustained inward current can have a region of negative resistance around −20 mV. This current is absent when Na+ is excluded from the bath solution, and it increases as the Na+ concentration increases from 2 to 8 mM. Injection of Na+ into the cell suppresses this inward current. This current develops very slowly, reaching its maximum seconds after the step depolarization and decays with a time constant of hundreds of milliseconds after the repolarization. This slow current is dependent on Ca2+. It can be suppressed by reduction or deletion of external Ca2+ or by iontophoretic injection of EGTA. ‘Pawn’ mutants with defective Caconductance also lack this current. We conclude that Paramecium has a Ca-induced conductance through which the Na-current flows. Although more prominent in the ‘paranoiac’ mutant, this Ca-induced Na-current is also seen in the wild type. This conductance may function in generating plateau depolarizations lasting seconds or even minutes and the corresponding prolonged backward swimming away from sources of irritation and stress.

scholarly journals Voltage clamp analysis of embryonic heart cell aggregates.

1979 ◽  
Vol 73 (2) ◽  
pp. 175-198 ◽  
Author(s):  
R D Nathan ◽  
R L DeHaan
Keyword(s):  
Voltage Clamp ◽  
Voltage Dependence ◽  
Slow Component ◽  
Outward Current ◽  
Heart Cell ◽  
Heart Cells ◽  
Spatial Homogeneity ◽  
Inward Current ◽  
Maximal Activation ◽  
Inward Currents

The double-microelectrode voltage clamp technique was applied to small spheroidal aggregates of heart cells from 7-d chick embryos. A third intracellular electrode was sometimes used to monitor spatial homogeneity. On average, aggregates were found to deviate from isopotentiality by 12% during the first 3--5 ms of large depolarizing voltage steps, when inward current was maximal, and by less than 3% thereafter. Two components of inward current were recorded: (a) a fast, transient current associated with the rapid upstroke of the action potential, which was abolished by tetrodotoxin (TTX); and (b) a slower inward current related to the plateau, which was not affected by TTX but was blocked by D600. The magnitudes, kinetics, and voltage dependence of these two inward currents and a delayed outward current were similar to those reported for adult cardiac preparations. From a holding potential of -60 mV, the peak fast component at the point of maximal activation (-20 mV) was -185 microA/cm2. This value was about seven times greater than the maximal slow component which peaked at 0 mV. The ratio of rate constants for the decay of the two currents was between 10:1 and 30:1.

Voltage-Dependent Conductances in Cephalopod Primary Sensory Hair Cells

1997 ◽  
Vol 78 (6) ◽  
pp. 3125-3132 ◽  
Author(s):  
Abdesslam Chrachri ◽  
Roddy Williamson
Keyword(s):  
Hair Cells ◽  
Outward Current ◽  
Sensory Hair ◽  
Inward Current ◽  
Transient Inward Current ◽  
Outward Currents ◽  
Sensory Hair Cells ◽  
Ionic Conductances ◽  
Voltage Dependent ◽  
Inward Currents

Chrachri, Abdesslam and Roddy Williamson. Voltage-dependent conductances in primary sensory hair cells. J. Neurophysiol. 78: 3125–3132, 1997. Cephalopods, such as sepia, squid, and octopus, show a well-developed and sophisticated control of balance particularly during prey capture and escape behaviors. There are two separate areas of sensory epithelium in cephalopod statocysts, a macula/statolith system, which detects linear accelerations (gravity), and a crista/cupula system, which detects rotational movements. The aim of this study is to characterize the ionic conductances in the basolateral membrane of primary sensory hair cells. These were studied using a whole cell patch-clamp technique, which allowed us to identify five ionic conductances in the isolated primary hair cells; an inward sodium current, an inward calcium current, and three potassium outward currents. These outward currents were distinguishable on the basis of their voltage-dependence and pharmacological sensitivities. First, a transient outward current ( I A) was elicited by depolarizing voltage steps from a holding potential of −60 mV, was inactivated by holding the cell at −40 mV, and was blocked by 4-aminopyridine. A second, voltage-sensitive, outward current with a sustained time course was identified. This current was not blocked by 4-aminopyridine nor inactivated at a holding potential of −40 mV and hence could be separated from I A using these protocols. A third outward current that depended on Ca2+ entry for its activation was detected, this current was identified by its sensitivity to Ca2+ channel blockers such as Co2+ and Cd2+ and by the N-shaped profile of its current-voltage curve. Inward currents were studied using cesium aspartate solution in the pipette to block the outward currents. Two inward currents were observed in the primary sensory hair cells. A fast transient inward current, which is presumably responsible for spike generation. This inward current appeared as a rapidly activating inward current; this was strongly voltage dependent. Three lines of evidence suggest that this fast transient inward current is a Na+ current ( I Na). First, it was blocked by tetrodotoxin (TTX); second, it also was blocked by Na+-free saline; and third, it was inactivated when primary hair cells were held at a potential more than −40 mV. The sustained inward current was not affected by TTX and was increased in amplitude 5 min after equimolar Ba2+ replaced Ca2+ as a charge carrier. This inward current also was blocked after external application of 2 mmol/l Co2+ or Cd2+. Furthermore, this current was reduced significantly in a dose-dependent manner by nifedipine, suggesting that it is an L-type Ca2+ current ( I Ca).

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