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1. Voltage‐clamp studies were carried out on single rabbit myelinated nerve fibres at 14 degrees C with the method of Dodge & Frankenhaeuser (1958). 2. A method was developed to allow the ionic currents through the modal membrane to be calibrated exactly under voltage‐clamp conditions by measuring the resistance of the internode through which the current was injected. 3. The ionic currents in a rabbit node of Ranvier can be resolved into two components, a sodium current and a leak current. Potassium current is almost entirely absent. 4. The sodium currents in rabbit nodes were fitted by the Hodgkin‐Huxley model using m2h kinetics. The kinetics of sodium currents in a rabbit node differ from that in a frog node under similar experimental conditions in two respects: (a) inactivation is faster, tau h for rabbit being 2‐3 times smaller around ‐50 mV; (b) the P(Na) (E) curve for mammal is shifted 10‐15 mV in the hyperpolarizing direction. 5. From the kinetics of sodium current, the non‐propagating rabbit action potential was reconstructed at 14 degrees C. The transient inward sodium current is responsible for the fast initial depolarization phase of the action potential, while the repolarizing phase is accounted for by leak alone. The computed shape of the action potential was in good agreement with the experimentally obtained action potential. 6. At 14 degrees C, frog and rabbit nodes with similar diameters have similar measured gNa values.

SUMMARY1. A study has been made of the hyperpolarization that follows a period of electrical activity (the post-tetanic hyperpolarization) in mammalian non-myelinated nerve fibres.2. Evidence is presented that under certain circumstances this posttetanic hyperpolarization is a result of activity of an electrogenic sodium pump that normally is absolutely dependent on the external presence of potassium.3. When the external chloride is replaced by sulphate or by isethionate the post-tetanic hyperpolarization, which in normal Locke solution is only a few millivolts in amplitude, is increased usually to about 20 mV, and on occasion to 35 mV.4. This effect of removing the chloride takes several minutes to develop and is consistent with the idea that the increase in the post-tetanic response is the result of removing the short-circuiting effect of internal chloride ions (by their being washed out into the chloride-free bathing medium).5. Small anions, such as chloride, nitrate, iodide, bromide, and thiocyanate can short-circuit the electrogenic pump, whereas larger anions such as sulphate and isethionate cannot. The bicarbonate ion, which is larger than chloride, short-circuits the pump but less effectively.6. In Locke solution containing 5 mm potassium the post-tetanic hyperpolarization declines exponentially, with a time constant of about 1-3 min. The time constant is inversely related to the external potassium concentration.7. However, when the external potassium concentration is zero the hyperpolarization declines rapidly to a very small value. Subsequent * Present address: Department of Pharmacology, University of Oxford.H. P. RANG AND J. M. RITCHIE addition of potassium to the bathing medium causes a marked redevelopment of the hyperpolarization.8. This potassium-activated response declines exponentially with a time constant that is inversely related to the potassium concentration. When the added potassium concentration is 5 mm, the time constant is 1-9 min.9. The amplitude of the potassium-activated response increases with increasing concentrations of potassium.10. Other cations can produce this activated response. Thus, thallium is more effective than, rubidium as effective as, caesium and ammonium about 1/10 as effective as, and lithium ions about 1/30 as effective as potassium in producing the activated response. Choline is quite ineffective.11. The size of the post-tetanic response is little affected by changes in the duration of the period of stimulation. However, increasing the duration definitely increases the time constant of recovery.12. Reducing the external sodium concentration increases the size of the post-tetanic hyperpolarization (by about 25 %), but the effect is complex and requires further study. 13. Reducing the calcium of the Locke-solution from 2-2 to 0-2 mm has no appreciable effect on the post-tetanic response, nor has increasing the pH of the Locke from 7-2 to 9-2.14. When the membrane potential is increased or decreased, by externally applied currents, there is relatively little change i...

The membrane of the myelinated axon expresses a rich repertoire of physiologically active molecules: (1) Voltage-sensitive NA+ channels are clustered at high density (approximately 1,000/microns 2) in the nodal axon membrane and are present at lower density (< 25/microns 2) in the internodal axon membrane under the myelin. Na+ channels are also present within Schwann cell processes (in peripheral nerve) and perinodal astrocyte processes (in the central nervous system) which contact the Na+ channel-rich axon membrane at the node. In some demyelinated fibers, the bared (formerly internodal) axon membrane reorganizes and expresses a higher-than-normal Na+ channel density, providing a basis for restoration of conduction. The presence of glial cell processes, adjacent to foci of Na+ channels in immature and demyelinated axons, suggests that glial cells participate in the clustering of Na+ channels in the axon membrane. (2) "Fast" K+ channels, sensitive to 4-aminopyridine, are present in the paranodal or internodal axon membrane under the myelin; these channels may function to prevent reexcitation following action potentials, or participate in the generation of an internodal resting potential. (3) "Slow" K+ channels, sensitive to tetraethylammonium, are present in the nodal axon membrane and, in lower densities, in the internodal axon membrane; their activation produces a hyperpolarizing afterpotential which modulates repetitive firing. (4) The "inward rectifier" is activated by hyperpolarization. This channel is permeable to both Na+ and K+ ions and may modulate axonal excitability or participate in ionic reuptake following activity. (5) Na+/K(+)-ATPase and (6) Ca(2+)-ATPase are also present in the axon membrane and function to maintain transmembrane gradients of Na+, K+, and Ca2+. (7) A specialized antiporter molecule, the Na+/Ca2+ exchanger, is present in myelinated axons within central nervous system white matter. Following anoxia, the Na+/Ca2+ exchanger mediates an influx of Ca2+ which damages the axon. The molecular organization of the myelinated axon has important pathophysiological implications. Blockade of fast K+ channels and Na+/K(+)-ATPase improves action potential conduction in some demyelinated axons, and block of the Na+/Ca2+ exchanger protects white matter axons from anoxic injury. Modification of ion channels, pumps, and exchangers in myelinated fibers may thus provide an important therapeutic approach for a number of neurological disorders.

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Cultured Schwann cells from sciatic nerves of newborn rabbits and rats have been examined with patchclamp techniques. In rabbit cells, single sodium and potassium channels have been detected with single channel conductances of 20 pS and 19 pS, respectively. Single sodium channels have a reversal potential within 15 mV of ENa, are blocked by tetrodotoxin, and have rapid and voltage-independent inactivation kinetics. Single potassium channels show current reversal close to EK and are blocked by 4-aminopyridine. From these results, and from comparisons of single-channel and whole-cell data, we show that these Schwann cells contain voltage-dependent sodium and potassium channels that are similar in most respects to the corresponding channels in mammalian axonal membranes. Cultured rat Schwann cells also have sodium channels, but at a density about 1/10th that of rabbit cells, a result in agreement with saxitoxin binding experiments on axon-free sectioned nerves. Saxitoxin binding to cultured cells suggests that there are up to 25,000 sodium channels in a single rabbit Schwann cell. We speculate that in vivo Schwann cells in myelinated axons might act as a local source for sodium channels at the nodal axolemma.Soon after a rat sciatic nerve is cut and allowed to degenerate, the ability of the distal degenerated stump to bind saxitoxin (STX), which has been widely used as a specific marker for the voltage-dependent sodium channels of excitable tissue, virtually completely disappears (1). However, in the rabbit sciatic nerve under the same conditions, the axon-free degenerated stump is found to bind substantial amounts of the toxin (1). This latter binding of STX increases during the several days immediately following sectioning, a time during which Schwann cells proliferate in the distal region and occupy the space vacated by the degenerating axons. Squid Schwann cells (2) and human glial cells (3) are already known to possess sodium channels that are opened by veratridine and blocked by tetrodotoxin (TTX). However, neither these latter experiments (which measured the entry of 22Na into cells) nor the STX experiments provide unequivocal evidence for the presence of voltage-dependent sodium channels of the kind necessary for excitability.Electrophysiological experiments using the patch-clamp technique have already shown that in rat cultured Schwann cells there are both anion-selective and cation-selective channels (4, 5). The latter, however, are calcium-activated, show no selectivity for sodium compared to potassium, and lack the characteristic voltage dependence of the sodium channels found in excitable tissues. This latter finding is consistent with the earlier finding of a lack of STX binding capacity of the Schwann cells in rat distal degenerated stumps (1). The present experiments show by contrast that rabbit Schwann cells do indeed possess a relatively high density of voltage-dependent sodium channels, in conformity with the findings of the earlier experiments that they bind STX avidly (1). They also pos...

Density of sodium channels in mammalian myelinated nerve fibers and nature of the axonal membrane under the myelin sheath Communicated by Alfred Gilman, October 26, 1976 ABSTRACT The density of sodium channels in mammalian myelinated fibers has been estimated from measurements of the binding of [3Hjsaxitoxin to rabbit sciatic nerve. Binding both to intact and to homogenized nerve consists of a linear, nonspecific, component and a saturable component that represents binding to the sodium channel. The maxim um saturable binding capacity in intact nerve is 19.9 ± 1.9 fmolPmg wetl; the equilibrium dissociation constant, K,, is 3.4 + 2.0 nM. Homogenization makes little difference, the maximum binding capacity being 19.9 i 1.5 fmol-mg wet-l with Kt = 1.3 + 0.7 nM. These values correspond to a density of about 700,000 sodium channels per node-i.e., about 12,000 per tm2 of nodal membrane. From

SUMMARY1. A study has been made of the temperature changes associated with the passage of a single impulse in rabbit desheathed vagus nerves.2. The initial changes consist of an evolution of positive heat followed by a reabsorption of most of it; i.e. there is a phase of positive and a phase of negative heat production.3. The size of the positive heat, its time of onset, and the time of onset of the negative heat have been measured by an analogue method of analysis. In addition, these parameters, together with the size of the negative heat and the duration of both phases of initial heat, have been studied with the aid of a computer, and also by conventional heat block analysis.4. At about 50 C the measured positive heat is 7-2 ,tcal/g. impulse. It starts as soon as the compound action potential reaches the thermopile and lasts for about 107 msec.5. This positive heat decreases with increasing temperature, the ratio of heat at 40 C to that at 140 C being 1*86.6. The measured negative heat at about 50 C is 4-9 /ucal/g. impulse. It starts 102 msec after the onset of positive heat, and lasts for about 240 msec.7. When the sodium of Locke solution is replaced by lithium the positive heat is reduced by 19 %, but the negative heat is increased by 22 %.8. In potassium-free solutions the positive heat is hardly affected (increase of 5 %), but the negative heat is more than doubled. As a result the nerve may become briefly colder than its initial temperature by about 2 #0 C.9. The effect of sodium-deficient solutions on the positive heat is somewhat variable, but the negative heat is consistently diminished. 11. Replacement of most of the sodium of Locke solution by barium reduces or abolishes the negative heat, and increases the measured size of the positive heat nearly threefold.12. Veratrine (10-5 g/ml.) produces a nearly tenfold increase in the net positive heat.13. Ouabain (1 mM) and antimycin A (1 ,ug/ml.) applied for 30-60 min have little effect on the initial heat production.14. Over the temperature range 5-15°C, and for the ionic solution changes described above, there is close agreement in timing between the positive heat and the rising phase of the action potential, and between the negative heat and the falling phase.15. Because of the inevitable temporal dispersion of the action potential over the face of the thermopile, the observed temperature changes are smaller than those which occur at a single point in the nerve close to a stimulating electrode. The corrected value of the positive heat at 50 C is 24*5 ,ucal/g. impulse, while that of the negative heat is 22-2 ,lcal/g. impulse.16. The heats of mixing of the ions in solution that interchange during the action potential are much too small to account for the observed initial heats, but an exchange of ions associated with fixed charges might make a significant contribution to the heats.17. The condenser theory, according to which the positive heat represents the dissipation of electrical energy stored in the membrane capacity, while the negative heat results f...