Frog Sartorius Muscle Fibres Research Articles

Detubulation experiments localise delayed rectifier currents to the surface membrane of amphibian skeletal muscle fibres.

Ionic currents in intact and detubulated frog sartorius muscle fibres were compared at room temperature using a loose-patch voltage clamp configuration in four experimental groups. The test fibres (i) were detubulated by a previously established osmotic shock protocol that involved the introduction and withdrawal of extracellular glycerol followed by exposure to Ca2+/Mg2+-Ringer solution and cooling. The control fibres were spared osmotic shock and (ii) simply studied in normal Ringer solution, (iii) exposed to 30 min of steady cooling to 9-10 degrees C before electrophysiological study or (iv) exposed to and studied in glycerol-Ringer solution. The presence or absence of detubulation was confirmed for all the experimental groups through assessing for the abolition or otherwise of the delayed after-depolarisation normally associated with action potential propagation into the transverse (T) tubules. All fibre groups showed similar resting potentials (-80 to -90 mV) thus ensuring consistent baseline voltages from which the voltage clamp steps were imposed. The intact muscle fibres in the three control groups (ii)-(iv) spared osmotic shock showed both inward Na+ and delayed rectifier outward (K+) currents. In contrast, patches from detubulated muscle fibres in the test group (i) showed only delayed outward currents, consistent with contrasting contributions to Na+ and K+ currents from regions of membrane affected or spared by the detubulation procedure. Nevertheless, the voltage dependence, maximum steady state amplitudes and timecourses of the delayed outward currents were conserved through all the experimental groups. These findings suggest that the surface as opposed to the tubular membrane contributes the greater part of the delayed rectifier current in amphibian skeletal muscle.

Time-resolved equatorial X-ray diffraction studies of skinned muscle fibres during stretch and release

Equatorial X-ray diffraction patterns were recorded from small bundles of one to three chemically skinned frog sartorius muscle fibres (time resolution 250 μs) during rapid stretch and subsequent release. In the relaxed state, the dynamic A-band lattice spacing change as a result of a 2 % step stretch (determined from the positions of the 10 and 11 reflections) resulted in a 21 % increase in lattice volume, while static studies of spacing and sarcomere length indicated than an increase in volume of ⩾50 % for the same length change. In rigor, stretch caused a lattice volume decrease which was reversed by a subsequent release. In activated fibres (pCa 4.5) exposed to 10 mM 2,3-butanedione 2-monoxime (BDM), stretch was accompanied by a lattice compression exceeding that of constant volume behaviour, but during tension recovery, compression was partially reversed to leave a net spacing change close to that observed in the relaxed fibre. In the relaxed state, spacing changes were correlated with the amplitude of the length step, while in rigor and BDM states, spacing changes correlated more closely with axial force. This behaviour is explicable in terms of two components of radial force, one due to structural constraints as seen in the relaxed state, and an additional component arising from cross-bridge formation. The ratio of axial to radial force for a single thick filament resulting from a length step was four in rigor and BDM, but close to unity for the relaxed state.

Mechanism of neuromuscular blockade induced by phenthonium, a quaternary derivative of (-)-hyoscyamine, in skeletal muscles.

1. The mechanisms underlying the postjunctional blockade induced by phenthonium [N-(4-phenyl) phenacyl 1-hyoscyamine] were investigated in mammalian and amphibian muscles. This muscarinic antagonist was previously shown to enhance specifically the spontaneous acetylcholine (ACh) release at concentrations that blocked neuromuscular transmission. 2. In both rat diaphragm and frog sartorius muscles, phenthonium (Phen, 1-100 microM) depressed the muscle twitches elicited by nerve stimulation (IC50: 23 microM and 5 microM, respectively), and blocked the nerve-evoked muscle action potential. The neuromuscular blockade was not reversed after incubation with neostigmine. 3. Equal concentrations of Phen decreased the rate of rise and prolonged the falling phase of the directly elicited action potential in frog sartorius muscle fibres, indicating that the drug also affects the sodium and potassium conductance. 4. Phen (50 and 100 microM) protected the ACh receptor against alpha-bungarotoxin (BUTX) blockade in the mouse diaphragm allowing recording of endplate potentials and action potentials after 5 h wash with physiological salt solution. 5. Phen (10-100 microM) produced a concentration- and voltage-dependent decrease of the endplate current (e.p.c.), and induced nonlinearity of the current-voltage relationship. At high concentrations Phen also shortened the decay time constant of e.p.c (tau(e.p.c.)) and reduced its voltage sensitivity. 6. At the same range of concentrations, Phen also reduced the initial rate of [125I]-BUTX binding to junctional ACh receptors of the rat diaphragm (apparent dissociation constant = 24 microM), the relationship between the degree of inhibition and antagonist concentration being that expected for a competitive mechanism. 7. It is concluded that Phen affects the electrical excitability of the muscle fibre membrane, and blocks neuromuscular transmission through a mechanism that affects the agonist binding to its recognition site and ionic channel conductance of the nicotinic ACh receptor.

ATP and adenosine inhibit transmitter release at the frog neuromuscular junction through distinct presynaptic receptors.

1. The effects of exogenous ATP or adenosine on end-plate currents (e.p.cs; evoked by simultaneous action of a few hundred quanta of ACh) or on miniature e.p.cs (m.e.p.cs) were studied under voltage clamp conditions on frog sartorius muscle fibres. 2. ATP or adenosine (100 microM(-1) mM) reduced the e.p.c. amplitude but did not affect m.e.p.c. amplitude, decay time constant and voltage-dependence of m.e.p.c., suggesting that e.p.c. depression induced by these purines had presynaptic origin only. 3. The action of ATP, unlike that of adenosine, was prevented by the P2-purinoceptor antagonist suramin (100 microM). The stable ATP analogue alpha,beta-methylene ATP (100 microM), known to be desensitizing agent on P2X receptors, also abolished the depressant effect of ATP while sparing the action of adenosine. Concanavalin A, an inhibitor of ecto-5'-nucleotidase, did not affect the presynaptic action of exogenously applied ATP. 4. The presynaptic action of adenosine was prevented by theophylline (1 mM), a blocker of adenosine receptors, while the effect of ATP was not changed under these conditions. The selective blocker of A1 adenosine receptors, 8-cyclopentyl-1,3,dipropylxanthine (DPCPX; 0.1 microM), abolished the presynaptic action of adenosine but did not prevent the depressant effect of ATP. 5. The effects of ATP and adenosine (at nearly saturating concentration) were additive suggesting that these purines activated not only distinct receptors but also different intracellular signalling mechanisms. 6. In contrast to the hypothesis that at the neuromuscular junction ATP reduces transmitter release via enzymatic degradation to presynaptically active adenosine, our data suggest that ATP (through its own presynaptic receptors) directly inhibits ACh release. Thus, ATP and adenosine might be almost equipotent as endogenous prejunctional neuromodulators at the neuromuscular junction.

Reappearance of miniature endplate potentials in frog neuromuscular junctions “silenced” by lanthanum ions

Miniature endplate potentials and currents were recorded from frog sartorius muscle fibres. As expected from previous work, La 3+ caused a very large increase in miniature endplate potential frequency, followed by a progressive decrease, ending in the virtually complete disappearance of miniature endplate potentials. Soon after its application, La 3+ caused an increase in the amplitude of miniature endplate potentials due to a lengthening of the underlying miniature endplate currents. Analysis of membrane current fluctuations produced by acetylcholine showed that La 3+ caused an increase in the lifetime of the channels opened by acetylcholine. After miniature endplate potentials had vanished following La 3+ treatment, the muscles were placed in organ culture conditions to see if miniature endplate potentials would reappear. After several hours miniature endplate potential activity was resumed in about 25% of the endplates, and disappeared again a few hours later, presumably because of accelerated nerve terminal degeneration. Preliminary electron microscopic observations suggest that the recovery of miniature endplate potential activity was accompanied by re-formation of synaptic vesicles in the motor endings.

Intracellular pH recovery from lactic acidosis of single skeletal muscle fibers.

Intracellular pH (pHi), measured with H+-selective microelectrodes, in quiescent frog sartorius muscle fibres was 7.29 +/- 0.09 (n = 13). Frog muscle fibres were superfused with a modified Ringer solution containing 30 mM HEPES buffer, at extracellular pH (pHo) 7.35. Intracellular pH decreased to 6.45 +/- 0.14 (n = 13) following replacement of 30 mM NaCl with sodium lactate (30 mM MES, pHo 6.20). Intracellular pH recovery, upon removal of external lactic acid, depended on the buffer concentration of the modified Ringer solution. The measured values of the pHi recovery rates was 0.06 +/- 0.01 delta pHi/min (n = 5) in 3 mM HEPES and was 0.18 +/- 0.06 delta pHi/min (n = 13) in 30 mM HEPES, pHo 7.35. The Na+-H+ exchange inhibitor amiloride (2 mM) slightly reduced pHi recovery rate. The results indicate that the net proton efflux from lactic acidotic frog skeletal muscle is mainly by lactic acid efflux and is limited by the transmembrane pH gradient which, in turn, depends on the extracellular buffer capacity in the diffusion limited space around the muscle fibres.

A microelectrode study of the mechanisms of L-lactate entry into and release from frog sartorius muscle.

1. Changes in intracellular pH and intracellular anion levels were monitored in frog sartorius muscle fibres during exposure to extracellular L-lactate, using ion-sensitive microelectrodes. 2. Resting intracellular pH (pHi) in 20 mmol l-1 HEPES buffer was 7.18 +/- 0.015 (S.E. of mean, n = 62). Exposure to an extracellular solution at pH 6.5 buffered with 20 mmol l-1 3-(N-morpholino)propanesulphonic acid (MOPS) resulted in a slow intracellular acidification. 3. A reversible decrease in pHi and an increase in intracellular anion levels was observed when L-lactate replaced chloride in equimolar amounts. The increase in intracellular anion level is consistent with intracellular accumulation of L-lactate ion. 4. The rate and steady-state change in pHi and anion level was a function of both extracellular pH and L-lactate concentration, providing evidence for the coupled movement of lactate and proton equivalents. 5. The initial rate of uptake of L-lactate, as measured by the change of pHi, was a non-linear function of the extracellular L-lactate concentration at extracellular pH 6.8 and 7.35. 6. No saturation was observed with concentrations of L-lactate between 5 and 60 mmol l-1 at pH 7.35 and 2.5 and 40 mmol l-1 at pH 6.8. 7. The non-linear relationship between the initial rate of change in pHi and extracellular L-lactate was well fitted by a curve defining uptake as the sum of a carrier process displaying Michaelis-Menten kinetics and a passive diffusion component. The apparent Km of the carrier was 10 mmol l-1 at pHo 7.35 and 4 mmol l-1 at pHo 6.8. 8. The initial rate of change of pHi in the presence of L-lactate was significantly inhibited 39.1 +/- 6.2% by 2-5 mmol l-1 alpha-cyano-4-hydroxycinnamate (n = 9; P less than 0.05, paired t test). 9. alpha-Cyano-4-hydroxycinnamate had no detectable effect on the initial rate of change of pHi induced by propionate exposure. 10. The initial rate of change of pHi induced by L-lactate was not affected by 20-100 mumol l-1 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS). 11. We conclude that L-lactate crosses the membrane of the frog sartorius muscle with proton equivalents via (1) a carrier-mediated process, and (2) passive diffusion of lactic acid. In the physiological range of L-lactate concentrations and pH the transport process dominates.

Qualitative measurements of the entry of L-lactate into single surface fibres of frog skeletal muscle using a lactate-sensitive microelectrode

The present results demonstrate the sensitivity of the Corning chloride liquid ion exchanger 477913 to L-lactate. Microelectrodes filled with this exchanger showed responses to changes in L-lactate concentration in chloride-free solutions. In these experiments L-lactate replaced gluconate in equimolar amounts. Microelectrodes filled with this exchanger were used to qualitatively detect changes in intracellular anion in chloride-depleted frog sartorius muscle fibres during exposure to extracellular concentrations of L-lactate. The increase in intracellular anion concentration is consistent with the movement of L-lactate into the cell. This microelectrode enables one to qualitatively monitor changes in intracellular L-lactate in chloride-free experiments without incorporating selectivity coefficients.

TMB-8 can block twitches without blocking high K + or caffeine induced contractures in frog's skeletal muscle

TMB-8 [8-(diethylamino)-octyl-3,4, 5trimethoxybenzoate] is known to inhibit calcium ion dependent processes in several tissues by stabilizing some intracellular stores of membrane-bound calcium. TMB-8 was used to study the excitation-contraction (E-C) coupling process in frog's skeletal muscle. TMB-8 (5 × 10 −5 − 10 −4 M) blocked electrically evoked twitches but not high K + (123 mM)- or caffeine (2.36 mM)-induced contractures in isolated, curarized toe muscles. TMB-8 (10 −4 M) produced a small decrease (16%) in the action potential of frog's sartorius muscle fibres. However, reducing extracellular Na + to 44.7 mM produced a similar reduction (17%) in action potential amplitude but did not suppress the twitch; i.e. it produced only a small increase (about 10%) in twitch amplitude. It is known that potassium contractures are produced by extracellular Ca ++ ions which enter through calcium channels in the t-tubules and that caffeine produces contractures by sensitizing the sarcoplasmic reticulum to Ca ++-induced Ca ++ release. The present results suggest that TMB-8 blocks twitches by preventing the release of Ca ++ ions bound to the intracellular surface of the t-tubular membrane which is often called the store of ‘trigger-calcium’ ions.

The interactive effects of fatigue and pH on the ionic conductance of frog sartorius muscle fibers

The effects of fatigue on the membrane conductance of frog sartorius muscle at the resting potential and during an action potential were studied. When muscles were exposed to an extracellular pH of 8.0 the membrane conductance at the resting potential increased during fatigue by about 20% and returned to prefatigue level in about 20 min. The membrane conductance of muscle fibers exposed to pH 6.4 was about three times less than that of pH 8.0 and decreased further during fatigue. Furthermore, the recovery of a normal membrane conductance was slow at pH 6.4. Both the inward, depolarizing and the outward, repolarizing currents during the action potential are reduced in fatigue. In each case the effect is greater at pH 6.4 than at 8.0 and recovery towards normal values is slower at pH 6.4. It is concluded that the ionic conductance of the sarcolemmal membrane at the resting potential and during an action potential are modified by fatigue and that these changes are modulated by pHo.

Verapamil alters the amplitude and time course of miniature endplate current

The effect of verapamil on neuromuscular transmission was examined by recording miniature endplate currents (mepcs) in voltage-clamped frog sartorius muscle fibres. In the presence of 100 μM verapamil, the amplitude and time constant of decay of the mepcs (τ D) were reduced to 68% and 55% of control respectively, and the normal voltage dependency of τ D was decreased. Part of the decrease in amplitude of the mepc was independent of changes in τ D because, on washout, recovery of τ D was nore rapid than that of amplitude, and in some cells smaller concentrations of verapamil (1 and 10 μM) decreased amplitude without affecting τ D. Evidence of open and closed channel blockade by verapamil (5–20 μM) was obtained from ionophoretic end-plate current trains and it is proposed that these effects are mediated via an allosteric mechanism.

Effect of axoplasmic transport blockade on end-plate currents in frog muscle fibers

Evoked and spontaneous end-plate currents (EPC) were studied in normal voltage-clamped frog sartorius muscle fibers and 2 weeks after application of colchicine to the nerve innervating the muscle to block axoplasmic transport in its fibers. Application of colchicine was found to reduce the rate of rise and to prolong decay of EPC without affecting the amplitude of the EPC and miniature EPC, the quantum composition of EPC, and the frequency of miniature EPC. The histogram of distribution of the time constant (τ) of EPC decay under normal conditions follows the normal law, but after application of colchicine to the nerve it is shifted to the right, with separation of two modes (τ1 and τ2). Three types of synapses can be distinguished from the character of EPC decay: monoexponential decay with τ1 (44%), biexponential decay with τ1 and τ2 (39%), and monoexponential decay with τ2 (19%). An increase in τ of EPC decay is accompanied by strengthening of the dependence of this process on the clamping voltage. The current-voltage characteristic and reversal potential of EPC are unchanged. It is suggested that the change in character of EPC decay after application of colchicine to the motor nerve is due to the appearance of acetylcholine-activated ionic channels in the muscle membrane with a longer duration of the open state and with potential-dependence of the open state similar to that taking place after muscle denervation.

The influence of the permeant ions thallous and potassium on inward rectification in frog skeletal muscle.

A three-electrode voltage-clamp method was used to investigate the inactivation of Tl currents through the inward rectifier of frog sartorius muscle fibres, and the interaction between the permeant ions Tl+ and K+. In 80 mM-Tl Ringer inward currents inactivated on hyperpolarization along an exponential time course, with time constants that initially increased and then fell with increasing hyperpolarization. Because of the inactivation process steady-state conductances were smaller than instantaneous conductances at all potentials in Tl Ringer. The steady-state conductance increased to a maximum value at around - 100 mV in 80 mM-Tl Ringer, and then fell with increasing hyperpolarization. In K Ringer the steady-state conductance was greater at all potentials than the instantaneous conductance because K currents activate (rather than inactivate) on hyperpolarization. Time constants of Tl inactivation were the same when measured from the decay of current during a single pulse, or from the rate of recovery from inactivation using either a two- or a three-pulse method, indicating that inactivation obeys first-order kinetics. In 80 mM-Tl Ringer steady-state inactivation increased with increasing hyperpolarization, e-fold every 48 mV. This would be consistent with the site at which inactivation occurs experiencing 0.5 of the membrane voltage field. Tl+ was more permeant than K+ through the inward rectifier, the permeability ratio PTl+/PK+ being 1.66. In solutions containing both Tl+ and K+ the membrane showed an anomalous mole-fraction dependence of conductance, the resting potential being more negative, and both instantaneous and steady-state conductances smaller than those recorded in solutions containing only Tl+ or only K+. The reduction in the amplitude of the instantaneous conductance in Tl-K mixtures was voltage-dependent, the block being initially increased and then falling with increasing hyperpolarization. Inward currents also inactivated on hyperpolarization in Tl-K mixtures. The time constants of inactivation, and the extent of inactivation which occurred, became less dependent on membrane potential in these solutions. When K+ is the major permeant ion in solution, Tl+ has a blocking effect on the currents carried by K+, and the degree of block is voltage-dependent. Increasing [Tl]o increased the block at all potentials. The results of our experiments in solutions containing both Tl+ and K+ are discussed in terms of an interaction between these ions within the channel.

Aequorin-calcium transients in frog twitch muscle fibres.

Intracellular Ca2+ transients, evoked either by action potentials or depolarizing clamp pulses, were studied in frog sartorius muscle fibres injected with aequorin. The time course of the Ca2+ transients became shorter as the temperature was increased. The half rise time and decay time constants showed straight lines between 3 and 30 degrees C in Arrhenius plots, with a Q10 of 2.5 and 2.3 respectively. The potential dependence of the Ca2+ transient was examined under voltage clamp. The peak light amplitude reached a plateau at around +50 mV, suggesting that Ca2+ release continues beyond the potential level at which contraction was saturated. During a prolonged depolarization, the Ca2+ transient gradually declined. The time course of decline became faster when long depolarizing pulses were repeated, or when the temperature was increased. The Q10 for half duration of the Ca2+ transient evoked by prolonged depolarization was 2.2. A Ca2+ transient could be evoked in Ca2+-free Ringer solution containing EGTA. Formamide, which is known to abolish excitation-contraction coupling, also abolished the Ca2+ transient. During maintained depolarization, the time integral of the Ca2+ transient was larger for larger depolarizations, suggesting that the total amount of Ca2+ released was greater for the more intense depolarization. The decline of the Ca2+ transient during maintained depolarization is probably due to inactivation of excitation-contraction coupling rather than the depletion of intracellular Ca2+ stores. These findings support the view that in frog skeletal muscle fibres the increase in intracellular Ca2+, caused by membrane depolarization, is produced by the release of Ca2+ from intracellular stores and that any influx of Ca2+ from the external medium does not contribute appreciably to the aequorin-Ca2+ transient.

Electrophysiological studies of the action of ketamine in frog skeletal muscle

The effects of the dissociative analgesic anaesthetic, ketamine, on action potential production in frog sartorius muscle fibres, were studied with bipolar extracellular electrodes and intracellular microelectrode techniques. All concentrations of ketamine depressed the extracellularly recorded compound action potential. Low concentrations of ketamine (less than 1 × 10 −4 m) increased excitability by reducing the current needed to initiate an action potential. Higher concentrations of ketamine decreased excitability by inhibiting the maximum rate of rise, the overshoot, and the maximum amplitude of the action potential. The resting membrane potential was not significantly affected by any of the ketamine concentrations employed. The depressant effects of ketamine on the action potential were antagonized in an apparently competitive manner by sodium ions. Ketamine caused a shift in the relationship between rate of fall and amplitude to higher action potential amplitudes, indicating that the drug inhibited the increase in potassium conductance (gK) associated with the falling phase of the action potential. It can be concluded that ketamine decreases excitability in frog sartorius muscle by inhibiting the specific increase in sodium conductance, which normally follows a threshold stimulus and is responsible for the rising phase of the action potential.

Rubidium block and rubidium permeability of the inward rectifier of frog skeletal muscle fibres.

1. A three-electrode voltage clamp method was used to investigate the Rb block of inward rectification in frog sartorius muscle fibres. 2. In a solution containing 80 mM-K+, the potassium conductance increased with increasing hyperpolarization to 3.18 +/- 0.11 mS. cm-2 (n = 17) when (V - VK) was -150 mV. In the presence of Rb+, the conductance increased, fell and increased again with increasing hyperpolarization, i.e. the Rb block was first increased and then reduced by increasing hyperpolarization. Increasing [Rb]o increased the block at all voltages. 3. In a solution containing 80 mM-Rb+ (zero K+) inward currents were recorded when the membrane was hyperpolarized beyond about -60 mV. These currents, which were < 10% the amplitude of those in 80 mM-K solution, were blocked by tetraethylammonium ions. 4. Experiments were carried out in solutions either where both [K]o and [K]i were increased, or where [K]o only was increased. The form of the relation between K conductance and membrane potential appeared to depend on [K]o. The magnitude of the conductance appeared to depend on [K]o and on [K]i. 5. So far as the block by Rb+ is concerned, increasing [K]o appeared to enhance the release of Rb block under large hyperpolarizations. Increasing [K]o and [K]i reduced the Rb block at all membrane potentials. 6. The results of experiments in the presence of Rb+ and Cs+ suggest that these two ions do not compete with each other for a site at which they block inward rectification. Rather, over a range of membrane potentials from -25 to -65 mV, the presence of Cs+ enhances the Rb block and vice versa. 7. Single dissected muscle fibres (from semitendinosus) were used to measure sarcoplasmic resistivity in 80 mM-K solution and 160 mM-K (hyperosmotic) solution. The measured values were 163.2 +/- 11.7 omega x cm and 136.1 +/- 16.0 omega x cm, respectively (n = 7). 8. A semi-empirical model is presented, supposing that Rb interacts with a site in the membrane to produce its blocking effect, but is able to move on through into the sarcoplasm. Internal K+ is supposed to reduce the affinity of the site for Rb+; external K+ is able to enhance the moving on of Rb+ into the sarcoplasm. 9. The implications of our experiments for the nature of the permeability mechanism inward rectification are discussed.

Potassium depletion and sodium block of potassium currents under hyperpolarization in frog sartorius muscle.

1. A three-electrode voltage clamp method was used to investigate the mechanism of the fall in resting potassium permeability which occurs under extreme hyperpolarization in frog sartorius muscle fibres. 2. Experiments were performed which show that this permeability change is due to a potential dependent block by Na+ ions present in the external solution. 3. Inward K-currents recorded on hyperpolarization turned off exponentially with time. In the presence of Na the steady-state current-voltage relation had a region of negative slope beyond -140 mV. This negative-slope region was removed when Na was replaced by TMA, Tris or Li. Increasing [Na] to 140 mM shifted the negative-slope region to less negative membrane potentials; reducing [Na] to 14 mM shifted the region to more negative potentials. 4. The time constant for the turn-off of the currents (tau) was the same in Na and TMA-containing solutions at membrane potentials positive to -140 mV. At more negative membrane potentials the tau s in Na became progressively shorter than those in TMA. Increasing [Na] to 140 mM (from 70 mM) gave smaller tau s at all potentials. 5. If fibres were hyperpolarized to -240 mV and then repolarized to -160 mV in 70 mM-Na the current recorded during the second pulse turned on with time, often reaching a value greater than that at the end of the first pulse. This behaviour was removed when Na was replaced by TMA or Tris. 6. An estimate of the steady-state relationship between the degree of block and membrane potential was obtained, and could be fitted by an expression for a potential-dependent ionic block with a very low affinity binding site for Na+ in the membrane. 7. The recovery after hyperpolarization of K-currents at the holding potential was examined in two-pulse experiments. In 70 mM-TMA recovery occurred at the same rate whether the initial hyperpolarization was to -120 or to -210 mV. In 70 mM-Na recovery after an initial pulse to -120 mV occurred at the same rate as in TMA, but recovery after a pulse to -210 mV occurred about 9 times faster. These results are consistent with depletion of K from the lumen of the T-system dominating the turn-off of K currents in TMA and in Na for the hyperpolarization to -120 mV, but a different mechanism being involved for the -120 mV pulse in Na. 8. A three-compartment model is presented which attempts to describe the depletion of K from the T-system. The model accurately predicts the time-course for the decline of inward K-currents, both in 10 and 80 mM-K solutions.

Cooperative effect of calcium ions in mediator release from neuromuscular synapses of large and small frog sartorius muscle fibers

Cooperative effect of calcium ions in mediator release from neuromuscular synapses of large and small frog sartorius muscle fibers

A dual effect of formaldehyde on the inwardly rectifying potassium conductance in skeletal muscle.

1. The inwardly rectifying potassium conductance of the membrane of frog sartorius muscle fibres is greatly reduced by treatment of muscles for 30 min with a solution containing formaldehyde (10 mM). 2. A transient increase in the conductance of the inward rectifier is observed early during formaldehyde action. 3. Analysis of the biphasic time course of the conductance changes, as determined under controlled voltage conditions, suggests that treatment with formaldehyde alters simultaneously, but in opposite ways, two factors that determine the conductance of the inward rectifier. 4. The linear component of the current-voltage relation, which dominates the relation at strongly positive potentials, is not affected while the above changes occur. But on prolonged exposure to formaldehyde the leak conductance increases. 5. The effects of formaldehyde on the inward rectifier are reversible on prolonged superfusion with normal Ringer solution. 6. The slight inward rectification remaining after most of the extracellular K is replaced by Rb, is similarly reduced by treatment with formaldehyde. 7. The results are interpreted in terms of the chemical properties of formaldehyde and present views of the mechanisms of inward rectification.

Electrophysiological, morphometric, and histochemical characteristics of muscle fibers of the frog sartorius muscle

Investigation of succinate dehydrogenase (SD) activity in the fibers of the frog sartorius muscle revealed three groups of muscle fibers (dark, pale, and intermediate). The SD activity in the muscle fibers was inversely proportional to their diameter. The auter surface of the sartorius muscle consists mainly of dark muscle fibers, the inner surface mainly of pale fibers. A microelectrode study showed that fibers of the outer surface have lower values of action potentials, a longer negative after-potential, a lower quantum composition of the end-plate potentials, and higher amplitude and lower frequency of miniature end-plate potentials than fibers of the inner surface. Analysis of the results reveals definite relations between the histochemical profile of frog sartorius muscle fibers and their electrophysiological characteristics.