Search PubMed⌕ Search

Biomedical subjects

J R Clay

Publications and source records attributed to J R Clay.

At least 55 records · Page 3Linked to original sources
PubMed1985

Comparison of the effects of internal TEA+ and Cs+ on potassium current in squid giant axons.

Internal tetraethylammonium (TEA) and cesium ions block outward potassium current in nerve membrane in a voltage-dependent manner. Blockade with Cs+ occurs virtually instantaneously after membrane depolarization, whereas blockade with TEA+ occurs after a delay. The latter result suggested to Armstrong (1966, J. Gen. Physiol., 50:279-293; 1969, J. Gen. Physiol., 54:553-575) that potassium channels must open before TEA+ blockade can occur, which is in contrast to Cs+ blockade, which appears to be independent of channel gating. The results in this study concerning the effect of TEA+ on inward (tail) current argue against the Armstrong model. Specifically, TEA+ (partially) blocks inward current without altering the tail current time constant. This result indicates that TEA+ can occupy its binding site within the channel whether or not the channel gates are open. This alternative hypothesis can describe both the steady-state and time-dependent components of TEA+ blockade.

Animals↗
PubMed1985

Potassium current in the squid giant axon.

The squid giant axon was the first preparation to be investigated with the voltage clamp technique over 30 years ago by Cole (1949) and Hodgkin et al. (1952). During the intervening years it has continued to serve as a useful preparation for the development of other new techniques such as internal perfusion (Baker et al., 1962), gating current measurements (Armstrong and Bezanilla, 1974), and patch clamp measurements (Conti and Neher, 1980). It also has served as a useful comparative preparation for investigations of sodium and potassium currents in other excitable membrane preparations. This article has focused on the activation kinetics and the instantaneous current-voltage relation of the potassium component. The squid axon is well suited for studies of IK, because it appears to have only a single type of potassium channel, and the leakage current is relatively small under ideal conditions. The IK component is activated in a sigmoidal manner following membrane depolarization. It deactivates with a single exponential time constant following return of the membrane potential to the holding level, although the deactivation time constant varies with changes in the external potassium concentration. There has not, as yet, appeared a self-consistent model which describes all of these results. The current-voltage relation is a nonlinear function of driving force, which is approximately described by the Goldman-Hodgkin-Katz equation, although a model of the IV based on single file diffusion of ions through a channel is more in tune with the modern view of the ion permeation process (Hodgkin and Keynes, 1955; Hille and Schwarz, 1978; Clay and Shlesinger, 1977, 1983, 1984). Further progress in this area will probably be achieved both by the traditional techniques and by the patch clamp technique. The traditional method is well suited for studying tail current kinetics and the slow inactivation process. The patch clamp technique is well suited for studying the distribution of channels in the membrane and the kinetics of channel gating in steady state conditions.

Animals↗
PubMed1984

Analysis of the effects of cesium ions on potassium channel currents in biological membranes.

Cesium ions block potassium channels in biological membranes in a voltage dependent manner. For example, external cesium blocks inward current with little or no effect on outward current. Consequently, it produces a characteristic N-shaped current-voltage relationship. We have modeled this result by single file diffusion of ions in a narrow channel spanning the membrane with a special blocking site in the channel for cesium ions. The model enables us to make detailed comparisons of the effects of cesium on potassium channels in different types of biological membranes.

Animals↗
PubMed1984

Effects of D-600 on sodium current in squid axons.

The effects of the calcium antagonist D-600 (methoxyverapamil) on the excitatory inward sodium current, INa, of internally perfused squid giant axons were studied under voltage-clamp conditions. We observed little or no effect of the drug when it was added to the external solution at concentrations of 10-200 microM. Furthermore, it did not produce a frequency, or use-dependent block of INa when repetitive voltage-clamp pulses were used at rates of 2-5 Hz. However, it did produce use-dependent blockade of INa when it was placed internally at a concentration of 200 microM. These results in conjunction with other studies suggest that D-600 is a selective blocker of calcium channels in squid axons when the drug is placed in the external solution. Its effects, when placed in the internal solution, are similar to those of permanently charged local anesthetic derivatives, which also produce use-dependent block of INa.

Animals↗
PubMed1984

Potassium channel kinetics in squid axons with elevated levels of external potassium concentration.

Potassium ion current in squid axons is usually modified by the effects of ion accumulation in the periaxonal space during voltage-clamp depolarization. The time course of potassium channel activation and ion accumulation usually overlap. A widely accepted procedure for circumventing the effects of accumulation in measurements of activation kinetics consists of measuring the difference in the current at the end of a depolarizing pulse and immediately following return of the membrane potential to the holding level. This instantaneous jump procedure is based upon the assumptions that the potassium channel current-voltage relation (IV) is a linear function of the driving force, and that the IV and the potassium channel-gating kinetics are both independent of ion accumulation. The latter assumption appears to be appropriate for activation kinetics. However, both assumptions concerning the IV are incorrect, in general. Consequently, the jump procedure provides a misleading view of gating kinetics for membrane depolarizations that produce net current flow. Jump conductance measurements for depolarizations that produce little or no net current indicate that the Hodgkin-Huxley n4 model of potassium channel kinetics is appropriate for the physiological range of membrane potentials.

Animals↗
PubMed1984

Phase resetting of the rhythmic activity of embryonic heart cell aggregates. Experiment and theory.

Injection of a current pulse of brief duration into an aggregate of spontaneously beating chick embryonic heart cells resets the phase of the activity by either advancing or delaying the time of occurrence of the spontaneous beat subsequent to current injection. This effect depends upon the polarity, amplitude, and duration of the current pulse, as well as on the time of injection of the pulse. The transition from prolongation to shortening of the interbeat interval appears experimentally to be discontinuous for some stimulus conditions. These observations are analyzed by numerical investigation of a model of the ionic currents that underlie spontaneous activity in these preparations. The model consists of: Ix, which underlies the repolarization phase of the action potential, IK2, a time-dependent potassium ion pacemaker current, Ibg, a background or time-independent current, and INa, an inward sodium ion current that underlies the upstroke of the action potential. The steady state amplitude of the sum of these currents is an N-shaped function of potential. Slight shifts in the position of this current-voltage relation along the current axis can produce either one, two, or three intersections with the voltage axis. The number of these equilibrium points and the voltage dependence of INa contribute to apparent discontinuities of phase resetting. A current-voltage relation with three equilibrium points has a saddle point in the pacemaker voltage range. Certain combinations of current-pulse parameters and timing of injection can shift the state point near this saddle point and lead to an interbeat interval that is unbounded . Activation of INa is steeply voltage dependent. This results in apparently discontinuous phase resetting behavior for sufficiently large pulse amplitudes regardless of the number of equilibrium points. However, phase resetting is fundamentally a continuous function of the time of pulse injection for these conditions. These results demonstrate the ionic basis of phase resetting and provide a framework for topological analysis of this phenomenon in chick embryonic heart cell aggregates.

Action Potentials↗
PubMed1983

Effects of external cesium and rubidium on outward potassium currents in squid axons.

We have studied the effects of external cesium and rubidium on potassium conductance of voltage clamped squid axons over a broad range of concentrations of these ions relative to the external potassium concentration. Our primary novel finding concerning cesium is that relatively large concentrations of this ion are able to block a small, but statistically significant fraction of outward potassium current for potentials less than approximately 50 mV positive to reversal potential. This effect is relieved at more positive potentials. We have also found that external rubidium blocks outward current with a qualitatively similar voltage dependence. This effect is more readily apparent than the cesium blockade, occurring even for concentrations less than that of external potassium. Rubidium also has a blocking effect on inward current, which is relieved for potentials more than 20-40 mV negative to reversal, thereby allowing both potassium and rubidium ions to cross the membrane. We have described these results with a single-file diffusion model of ion permeation through potassium channels. The model analysis suggests that both rubidium and cesium ions exert their blocking effects at the innermost site of a two-site channel, and that rubidium competes with potassium ions for entry into the channel more effectively than does cesium under comparable conditions.

Animals↗
PubMed1983

Relationship between membrane excitability and single channel open-close kinetics.

We have developed a novel technique for simulating the influence of the effects of single channel kinetics on the voltage changes associated with membrane excitability. The technique uses probability distribution functions for the durations of channel open- and closed-state lifetimes, which can be calculated for any model of the ion conductance process. To illustrate the technique, we have used the Hodgkin and Huxley model of nerve membrane ion conductances to simulate channel kinetics during predetermined voltage changes, such as a voltage jump and an action potential. We have also simulated the influence of channels on voltage changes in a free running, non-voltage-clamped patch of membrane 1 micron2 or less in area. The latter results provide a direct illustration of the relationship between fluctuations of membrane excitability and fluctuations in channel open- and closed-state lifetimes.

Action Potentials↗
PubMed1982

Delayed kinetics of squid axon potassium channels do not always superpose after time translation.

The activation of potassium ion conductance in squid axons by voltage-clamp depolarization is delayed when the depolarizing step is preceded by a conditioning hyperpolarization of the axonal membrane. Moreover, the control conductance kinetics superpose with the delayed kinetics when they are translated along the time axis by an amount equal to the delay. We have found that the degree of superposition with internally perfused axons depends upon voltage-clamp protocol. The kinetics superpose almost exactly for modest test depolarizations, whereas they clearly fail to superpose completely for more positive levels of membrane depolarization. We have modeled these results by incorporating a time dependence into the rate constant of activation of potassium channel gates in the Hodgkin and Huxley model of potassium ionic conductance.

Animals↗
PubMed1981

Analysis of subthreshold pace-maker currents in chick embryonic heart cells.

1. Small re-aggregates of cells dissociated from the ventricles of 7-day-old chick embryonic hearts beat spontaneously in low external potassium concentration (Ko = 1.3 mM) tissue culture medium. This activity was blocked by the addition of tetrodotoxin (TTX) or potassium ions to the external medium. 2. A two-micro-electrode voltage-clamp technique was used to analyse the subthreshold currents responsible for the pace-maker depolarization. 3. Voltage-clamp steps 6-10 sec in duration revealed a time-dependent current having first order kinetics. Its membrane potential range of steady-state activation was -90 to -60 mV. 4. The current kinetics were qualitatively similar to those of Hodgkin & Huxley (1952b) with a peak time constant of approximately 1 sec at V = -75 mV. The kinetics were independent of Ko. 5. The time-dependent current was attributed to gated membrane channels. The fully activated current-voltage (I-V) relation of the channels was determined from the ratio of the amplitudes of the time-dependent currents during and after voltage-clamp steps following the procedure of Noble & Tsien (1968). 6. The fully activated I-V relation displayed inward rectification with negative slope conductance at potentials more than 15 mV positive to its reversal potential. Changes of Ko shifted the I-V curve along the voltage axis like a potassium electrode. 7. The time-independent (background) current was obtained by subtracting the gated channel current from the steady-state I-V curve. This current also rectified in the inward direction. 8. The inwardly rectifying I-V relations were theoretically described by a channel having a row of ion-selective sites along which ions move in a single file (Hodgkin & Keynes, 1955), and a membrane-bound particle which blocked the channel in a voltage-dependent manner. 9. The relationship of the voltage-clamp results to spontaneous activity is discussed and comparisons are made with measurements from whole embryonic heart and other cardiac tissues.

Age Factors↗
PubMed1981

Developmental changes in subthreshold pace-maker currents in chick embryonic heart cells.

1. Small re-aggregates of cells dissociated from the ventricles of 12-or 17-day-old chick embryonic hearts have beating properties in low potassium tissue culture medium which differ from re-aggregates prepared from 7-day-old hearts; 12-day preparations either beat with a slower rate than comparable-size 7-day preparations or they are quiescent; 17-day preparations usually do not beat. 2. A two-micro-electrode voltage-clamp technique was used to analyse the membrane currents in these preparations in the -120 to -40 mV potential range. 3. A potassium ion membrane current was measured in 12-day and 17-day preparations which was qualitatively similar to the potassium ion current measured in 7-day preparations (Clay & Shrier, 1981). It differed in that its amplitude was diminished in the ratio 1:0.2:0.03 over the 7-, 12- and 17-day sequence. Furthermore, its maximum time constant was 2 sec at both 12 and 17 days, as compared to 1 sec at 7 days, and its membrane potential range of activation was -80 to -70 mV at 12 and 17 days, as compared to -90 to -60 mV at 7 days. 4. The background current also changed with development, becoming less inward over the 7-, 12- and 17-day sequence. The voltage dependence of this change suggested that it was caused by a developmental decrease of the background sodium current. 5. The change in the background component is primarily responsible for the loss of automaticity over the 7-, 12-, and 17-day sequence.

Age Factors↗
PubMed1980

Pacemaker currents in chick embryonic heart cells change with development.

The initial heartbeat of the chick embryo occurs shortly after the first day of incubation. The pacemaker of this beat originates in the region of the primitive heart destined to become sinoatrial tissue in the adult. Individual cells isolated from the atrial and ventricular portions of the embryonic heart are also capable of beating spontaneously at this stage. However, the intrinsic activity of these cells gradually diminishes from about day 7 until day 21 when the chick hatches. We have investigated these changes in automaticity by measuring membrane pacemaker currents with the voltage-clamp technique from aggregates of cells prepared from 7-, 12- and 17-day-old chick embryo cardiac ventricle. We report that there is a voltage and time dependent conductance at 7 days which is markedly reduced by 17 days. The reduction of this pacemaker current parallels the decrease of spontaneous activity in these preparations.

Age Factors↗
PubMed1979

Current noise parameters derived from voltage noise and impedance in embryonic heart cell aggregates.

We have recorded membrane impedance and voltage noise in the pacemaker range of potentials (-70 to -59 mV) from spheroidal aggregates of 7-d embryonic chick ventricle cells made quiescent by exposure to tetrodotoxin in medium containing 4.5 mM K+. The input capacitance is proportional to aggregate volume and therefore to total membrane area. The specific membrane capacitance is 1.24 microF/cm2. The input resistance at constant potential is inversely proportional to aggregate volume and therefore to total membrane area. The specific membrane resistance in 18 k omega . cm2 at -70 mV and increases to 81 k omega . cm2 at -59 mV. The RC time constant is 22 ms at -70 mV and increases to 146 ms at -59 mV. The aggregate transmembrane small-signal impedance can be represented by a parallel RC circuit itself in parallel with an inductive branch consisting of a resistor (rL) and an inductor (L) in series. The time constant of the inductive branch (L/rL) is 340 ms, and is only weakly dependent on potential. Correlation functions of aggregate voltage noise and the impedance data were modeled by a population of channels with simple open-close kinetics. The time constant of a channel (tau s) derived from the noise analysis is 300 ms. The low frequency limit of the pacemaker current noise (SI[0]), derived from the voltage noise and impedance, increases from 10(-20) A2/Hz . cm2 at -67 mV to 10(-19) A2/Hz . cm2 at -61 mV.

Animals↗
PubMed1979

Fluctuations in interbeat interval in rhythmic heart-cell clusters. Role of membrane voltage noise.

Small clusters of ventricular cells prepared from 7-d chick heart maintain spontaneous, stationary, rhythmic beating in culture for many hours. For clusters containing I-125 cells, mean interbeat interval (IBI) is 0.45 +/- 0.08 s and is independent of cell number (N), whereas, the coefficient of variation of IBI (C) is proportional to N-1/2. Because membrane voltage noise in such clusters would also be expected to vary as N-1/2, we propose a model relating fluctuation in IBI (sigma IBI) to voltage noise (sigma v). A simplified model consisting of random voltage fluctuations superimposed upon a linear pacemaker depolarization of slope a is used to analyze the N-dependent shape of the IBI histogram. Values of sigma v derived from the relation sigma IBI = sigma v/a, or calculated from the skewness of the measured IBI histograms, both agree well with those extrapolated from steady-state noise recorded from resting heart-cell aggregates.

Animals↗
PubMed1978

Comparison of ion current noise predicted from different models of the sodium channel gating mechanism in nerve membrane.

The theoretical power density spectrum S(f) of ion current noise is calculated from several models of the sodium channel gating mechanism in nerve membrane. Sodium ion noise experimental data from the frog node of Ranvier [Conti, F., et al. (1976), J. Physiol. (London) 262:699] is used as a test of the theoretical results. The motivation for recent modeling has been evidence for a coupling between sodium activation and inactivation from voltage clamp data. The two processes are independent of one another in the Hodgkin and Huxley (HH) model [Hodgkin A.L., Huxley, A.F. (1952), J. Physiol. (London) 117:500]. The noise data is consistent with HH, as noted by Conti et al. (1976). The theoretical results given here appear to indicate that only one case of coupling models is also consistent with the noise data.

Animals↗