Biomedical subjects
Publications and source records attributed to C Ojeda.
1.20 mM Cs+ ions reduce background current and decrease drastically the K+ depletion process, probably as a consequence of the reduction of iK1. The background current-voltage relationship becomes linear. 2. 20 mM Cs+ ions completely abolish the current induced by acetylcholine. 3. The possibility that the K+ current induced by acetylcholine is due to an increase of iK1 is discussed.
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1. Properties of the 'pace-maker' current if in rabbit sino-atrial node have been investigated by voltage clamp of small preparations and compared with those of the iK2 current in the Purkinje fibre. Besides having a similar voltage range of activation and responding in a similar way to adrenaline, if resembles iK2 in other respects. 2. When external Na is reduced, if decreases proportionally. In 25% Na the time-dependent current change due to if disappears. 3. 20 mM-Cs completely abolishes it. 4. The time constant of if during a hyperpolarizing voltage-clamp pulse displays a relatively high temperature dependence. 5. In spite of the similarities between the two current systems, experiments in high K solutions (48 mM) rule out the possibility that the current change seen on a hyperpolarization reflects the decay of a pure K current. 6. From conductance measurements during onset of if it is deduced that if behaves as an inward current activated by hyperpolarizations.
Voltage clamp experiments on isolated sheep Purkinje fibres showed an increase of the steady state outward membrane current, over the potential range -65mV to -15 mV, in the presence of tetrodotoxin (TTX, 3.10(-5 M). This "window" current is considered to be the steady state component of the fast sodium current (INa), resulting from the crossover of the activation and inactivation curves which govern the opening of the sodium channel. TTX had no significant effect on the reversal potential, activation curve, kinetics or instantaneous I-V relationship of the pacemaker current IK2. The window found in these experiments extends to potentials well into the range of the action potential plateau. Consequently small changes of the steady state INa might have large effects on the action potential duration. The effects of TTX and local anaesthetics are discussed in this context.
1. The effects of salicylate on the membrane currents in the frog node of Ranvier were investigated. 2. External salicylate slows the falling phase of the action potential, with little effect on the action potential amplitude. 3. External salicylate has no effect on the leak current. 4. Most of the actions of external salicylate can be attributed to a simple incorporation of negative charge into the membrane surface. The h infinity, tau h, tau m and m infinity curves for the sodium current are shifted to more negative potentials, as are the n infinity and k infinity curves for the potassium current. Potassium ion accumulation prevented analysis of the action of salicylate on the time constant of the potassium current kinetics. 5. In addition to the h infinity and tau h curves being shifted to more negative potentials, the shapes of the curves are also changed by salicylate. These shape changes cannot be explained by conventional homogeneous surface charge theory. Possible explanations for these changes are discussed. 6. Internal salicylate has similar effects to external salicylate: the gating variable curves for the sodium current are shifted in the negative direction on the voltage axis, rather than in the positive direction expected if negative charge were added to the inner surface of the nodal membrane. This may be due to salicylate crossing the membrane and binding preferentially to a receptor at the external surface, or might be due to a rise in intracellular calcium concentration following inhibition of oxidative phosphorylation.
1. The apparent reversal potential (Erev) of the pace-maker current (iK2) is found to depend on the experimental protocol used for its measurement. Evidence is presented showing that depolarizing (hyperpolarizing) pulses given before a test hyperpolarization used to determine Erev, shift Erev to more negative (positive) values. These shifts are opposite to those expected if the only effect of pre-pulses were to change the concentration of potassium in extracellular clefts ([K]c) via accumulation and depletion processes. 2. This effect is shown to be due to the fact that Erev is dependent on s0, the degree of activation of iK2 at the start of the test hyperpolarization. 3. When a suitable protocol is used, depletion of cleft K can be demonstrated to take place during a large hyperpolarization. Changes in the level of [K]c induced by pre-pulses must therefore also affect the Erev determination. 4. A simplified three-compartment model has been used to investigate how K accumulation and depletion can affect the time course of iK2, with particular reference to the problem of Erev determination. Computed examples show that the model is able to reproduce the main features of the time course of iK2 recorded near its reversal potential and the changes induced by pre-pulses on Erev measuremnet. By contrast, simulation on a linear cable model rules out the possibility that such results are due to voltage non-uniformity. 5. The three-compartment model predicts that the measured value of Erev differs from EK2 for two reasons: (1) when the recorded current trace is flat iK2 is still outward and decaying, and (2) the K equilibrium potential shifts to more negative values while the test hyperpolarization is applied. 6. The finding that Erev is directly affected by changes in s at the beginning of the test pulse is discussed in relation to the action of agents (such as Ca2+, H+, salicylate, adrenaline and ouabain) which are found to shift both the s00 curve and Erev.
1. In sheep Purkinje fibre preparations, salicylate ions produce reversible changes in resting potential and in action potential duration. In most preparations these effects resemble those produced by -ow extracellular K concentration: the resting potential first increases and then decreases, the action potential is prolonged and eventually, low potential oscillations occur in the plateau range. In a few preparations, action potential shortening occurs. 2. The threshold current for initiating action potentials by an intracellular electrode is reversibly increased by salicylate. 3. The activation curve, soo(Em), for the pace-maker K current, iK2, shifted in a hyperpolarizing direction. The magnitude of the shift is about -5 mV in 5 mM-salicylate and -30 mV in 50 mM-salicylate. 4. The apparent reversal potential for iK2 is shifted in a negative direction. The magnitude of this shift at a given salicylate concent;ation varies with the K concentration. In an extracellular K concentration of 2.7 mM an average shift of -18 mV occurs in 10 mM-salicylate; in 8 mM, the average shift is only -1 mV. 5. It is proposed that most of these effects may be produced by an increase in surface negative potential produced by the binding of salicylate to the cell membrane. This would produce the hyperpolarizing shift of activation curves for ionic current and, by increasing surface K activity, may lead to stimulation of the Na-K pump to produce an increase in the K gradient across the cell membrane.
1. The action potential duration in sheep ventricular fibres is rapidly diminished on exposure to 10(-6) M-ouabain. However, if 10--20 mM-sodium salicylate is added to the ouabain solution, glycoside-induced shortening is prevented, and a substantial increase in duration then occurs. Sodium salicylate also reverses the shortening effect of ouabain if applied after the glycoside has been allowed to act alone. 2. Sodium salicylate alone produces a much smaller prolongation than in the presence of ouabain. Alone and in the presence of ouabain it eventually increases the threshold and produces inexcitability. 3. Three other surface charge agents have been compared with salicylate: aminonaphthalene sulphonate, sodium dodecylsulphate and salicylamide, were unable to counter the actions of ouabain at the concentrations used. Since they also produced no changes in excitability it is likely that they did not bind significantly to the cell membrane. 4. In Purkinje fibres the reversal potential for the pacemaker current, iK2, is initially shifted in a negative direction in the presence of 10(-6) M-ouabain and 10 mM-salicylate instead of the positive shift expected with ouabain alone at this concentration. 5. In guinea-pig ventricle, salicylate alone reduces the duration of the action potential. This effect is rapidly reversible. Toxic levels of ouabain also reduce the action potential duration but this effect takes several hours to reverse. By contrast, the effects of salicylate and ouabain applied together are readily reversible. 6. It is suggested that the mechanism of these effects may depend on the ability of a surface negative charge agent like salicylate to increase the surface K+ concentration at the membrane and so protect the sodium-potassium pump from inhibition by large doses of ouabain.
The action of acetylcholine (ACh) on membrane potential and currents in frog atrial muscle has been studied with a double sucrose gap technique. The results show the following. 1. ACh induces the development of an extra current, outward at the resting potential, which is dependent on the ACh concentration. 2. The preparation does not show any sign of desensitization. 3. The reversal potential of the current induced by ACh is between 0 and 20 mV more negative than the resting potential and behaves as a K electrode. 4. The mechanism of ACh-induced K conductance presents inward going rectification properties. 5. The delayed outward current is not affected by ACh. However the evolution of its tail current seems to indicate a process of K accumulation related to the ACh-induced current.
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A double sucrose gap voltage clamp technic has been used to study the extra-current induced by acetylcholine (Iach) on the myocardial membrane on frog atrial trabeculae. I) No desensitization of the Iach current is noted for repeated perfusions of Ach. II) The Iach current is suppressed by atropine. III) The reversal potential Each is more negative than the resting potential --20 mV less than or equal to Each less than or equal to OmV.IV) The relationship Iach/Holding potential for various [K+]o shows a) That Each behaved as a potassium electrode, b) an inward going rectification. These results indicate that the cholinergic receptor might be related with the gk1 channel.
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1. The outward currents underlying delayed rectification have been studied in frog auricular trabeculae with the double sucrose gap method.2. The results obtained in our experimental conditions allow us to discard the hypothesis of an accumulation of ions during the flow of the outward current.3. In most of the cases the delayed current is composed of two components similar to those previously described by Noble & Tsien (1969a, b).4. The first component I(1) is activated with a time constant of about 0.3 sec at the resting potential (E = - 70 mV) and of about 0.75 sec at E = - 5 mV. The steady-state value of the degree of activation (x(1))(infinity) varies from 0 at E = - 90 mV to 1 at E = + 20 mV. The reversal potential is near E = - 80 mV; this current seems to be mainly carried by potassium ions.5. The second component I(2) is very slowly activated, with a time constant of 3.5 sec at E = - 70 mV and of 5 sec at E = - 5 mV. The steady-state value of the degree of activation (x(2))(infinity) varies from 0 at E = - 40 mV to 1 at E = + 25 mV. The reversal potential is near E = - 60 mV; this current is less specific than the first component.6. In some preparations the delayed current is composed of only one component. This current is activated with a time constant of 0.6 sec at the resting potential and of 1.5 sec at E = - 5 mV. The steady-state value of the degree of activation is 0 at E = - 90 mV and 1 at E = + 30 mV. The reversal potential is between E = - 70 and E = - 60 mV.7. The role of these currents in the processes of repolarization of normal and prolonged action potentials, and in the pace-making activity is discussed.
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