The interactive effects of temperature and pH on the isometric contraction of toad sartorius muscle

Summary The main purpose of this study was to examine the interactive effects of pH and temperature on the performance of isolated muscle in order to estimate the effect of temperature in vivo. The development of tetanic tension by the sartorius muscle isolated from toads was tested at either 5 or 25°C. The pH of the physiological solution bathing the muscle was changed from 9.0 to 6.0 stepwise by 0.5 pH units.In order to see if the temperature at which the toads were maintained modified the muscle activity, two groups of toads were used: those acclimated to low temperature (5°C) and those acclimated to high temperature (25°C). Acclimation temperature had no significant effect (Fig. 1, Table 1).Both the amount and rate of tension development increased when test temperature increased (Fig. 1A, B). Maximum tension and rate of tension development were significantly reduced at low pH. The latent period, time to half-maximum tension, and half-relaxation time were longer at low pH (Fig. 1C, D, E). The interactive effects of pH and test temperature were significant for all variables, except the latent period (Table 1). That is, the effects of pH were greater at 5 than at 25°C and the effects of test temperature depend on extracellular pH. We calculated that the in vivo decrease in tetanic tension when the body temperature of a toad decreases from 25 to 5°C is smaller than previously supposed. This is because in previous studies on the effect of test temperature on muscle function, unrealistic pH regimes generally have been employed.This work is supported by an NSERC operating grant to E.D. Stevens     

Is the change in intracellular pH during fatigue large enough to be the main cause of fatigue?

The intracellular pH of frog sartorius muscles exposed to an extracellular pH 8.0 (25 mM HCO3-, 1% CO2) was 6.9-7.1. Following a fatiguing stimulation period (one tetanic contraction per second for 3 min), the intracellular pH was 6.5-6.7. When similar experiments were repeated with frog sartorius muscles exposed to pH 6.4 (2mM HCO3-, 1% CO2), the intracellular pH was 6.8-6.9 at rest and 6.3-6.4 following fatigue. So, in both experiments the intracellular pH decreased by 0.4-0.5 pH unit during fatigue. When the CO2 concentration of the bathing solution was increased from 1 to 30%, the intracellular pH of resting muscles decreased from 7.0 to 6.2-6.3. Although the effect of CO2 on the intracellular pH was greater than the fatigue effect, the decrease in tetanic force with CO2 was less than 40%, while during fatigue the tetanic force decreased by at least 70%. Therefore in frog sartorius muscle the decrease in tetanic force during fatigue exceeds the decrease that is expected from just a change in intracellular pH.     

Tetanic fatigue and proximate post-tetanic recuperation in sartorius and flexor carpi radialis muscles of the male frog. Effects of iodoacetic acid (author's transl)]

The time-course of the isometric tension output, at 20 degrees C, during a long tetanus and after a short period of rest, was investigated in two isolated frog muscles : the sartorius and flexor carpi radialis muscles. To prevent aerobie and glycolytic recovery processes, some muscles were poisoned with 0,4 mM iodoacetic acid (IAA) and nitrogen, for 20 or 40 min. 1. For the unpoisoned sartorius muscle, tetanic tension declined quickly, but after a 0,8 sec period of rest, the muscle was able to develop high tension. Poisoning with IAA-N2 increased fatigue without suppressing the property of a proximate post-tetanic recuperation. 2. In the flexor carpi radialis muscle resistance to fatigue was very large before poisoning and diminished after poisoning. Proximate recuperation was very weak. 3. The results show that the recovery processes are not a primary factor of the development of the short-term fatigue ; they enhance the hypothesis that a failure of the electromechanical coupling can explain the rate of the tension fall in tetanized sartorius muscles.     

The effects of pH on the kinetics of fatigue and recovery in frog sartorius muscle

The effects of pH on the kinetics of fatigue and recovery in frog sartorius muscle were studied to establish whether the pH to which muscles are exposed (extracellular pH) has an effect on both the rate of fatigue development and recovery from fatigue. When frog sartorius muscles were stimulated with short tetanic stimuli at rates varying from 0.2 to 2.0 trains/s, a time- and frequency-dependent decrease in force development was observed, but extracellular pH had comparatively little effect. The recovery of tetanic force was dependent on the extracellular pH. This effect was characterized by a rapid recovery in force at pH 8.0 and an inhibition of recovery at pH 6.4 even when force decreased by only 25% during stimulation. Even when muscles were fatigued at pH 8.0 the rate of force recovery was still very small at pH 6.4. A model is proposed in which a step of the contraction cycle changes from a normal to a fatigued state. The rate of this transition is a function of the stimulation frequency and not pH. The reverse transition, from a fatigued to normal state is pH dependent; i.e., it is inhibited by H+. Measurements of resting and action potentials show that extracellular pH influences these parameters in the fatigue state, but there is no evidence that these changes are directly responsible for the pH-dependent step in the reversal of fatigue.     

Isometric contractile properties and instantaneous stiffness of amphibian skeletal muscle in the temperature range from 0 to 20 degrees C

The isometric contractile properties of frog (Rana pipiens) and toad (Bufo bufo) sartorii have been studied over the temperature range from 0 to 20 degrees C. The isometric twitch tension was found to vary considerably between these two species and between muscles in the same species. Between 0 and 4 degrees C there was very little change in maximum isometric twitch tension. Between 4 and 12 degrees C several muscles from frog or toad showed a potentiation of twitch tension whereas others showed a decline. Over this temperature range the toad sartorii consistently demonstrated a greater potentiation. By 12 degrees C a steady decline in twitch tension in both muscles was seen as the temperature range the toad sartorii consistently demonstrated a greater potentiation. By 12 degrees C a steady decline in twitch tension in both muscles was seen as the temperature approached 20 degrees C. The maximum isometric tetanic tension recorded between 18 and 20 degrees C increased fractionally to an average of 1.504 +/- 0.029 (n = 4) for frog sartorii and to 1.377 +/- 0.008 (n = 5) for toad sartorii. The time to peak twitch tension and the half-relaxation time decreased markedly with an increase in temperature. Moreover, the half-relaxation time was reduced by a greater proportion than the time to peak twitch tension. Measurements of instantaneous stiffness by controlled velocity releases from the plateau of isometric tetani revealed that the large increase in isometric tetanus tension as the muscle was warmed was not accompanied by a corresponding increase in the total number of active cross-bridges. The possibility that a decreased availability of intracellular Ca2+ ions at the contractile sites contributing to the fall of isometric twitch tension at elevated temperatures is discussed. The possibility exists that at elevated temperatures a change inthe intrinsic contractile ability of the muscle occurs which produces an increased tension per cross-bridge.     

Measurement of rate constants for the contractile cycle of intact mammalian muscle fibers.

Small, random length changes were applied to bundles of intact fibers from rat and mouse extensor digitorum longus (EDL) and soleus muscles, while they were being tetanically stimulated. With increasing frequency of length changes, EDL muscle stiffness (tension change per unit change in length) increased, then decreased and increased again. The decrease was not seen in the soleus muscles. The EDL frequency-response could be well fitted by three exponential components with apparent rate constants of approximately 25, 150, and 500 s-1 at 20 degrees C. All rate constants increased steadily with temperature and for each 10 degrees C increase in temperature, the rates in the mouse EDL increased by a factor (Q10) between 1.8 and 2.4. With tetanic stimulation, force increased nearly exponentially to a steady level with a rate constant of 24 s-1 at 20 degrees C in mouse EDL muscles, and a Q10 of 2.4. These values correspond closely to the lowest frequency rate constant measured with length perturbations, which suggests that this process may limit the rate of rise of force in intact muscle fibers. During fatigue the high frequency and intermediate frequency rate constants declined, but the low frequency rate constant remained unchanged. These results are discussed in relation to current biochemical models for cross-bridge cycling.     

Carbonic anhydrase III inhibition in normocapnic and hypercapnic contracting mouse soleus

The physiological role of carbonic anhydrase III in slow-twitch skeletal muscle was investigated using isolated mouse soleus (N = 30) contracting once every 1.7 min for 75 min in Krebs-Henseleit solution gassed with either 95% oxygen - 5% carbon dioxide (normocapnia) or 90% oxygen - 10% carbon dioxide (hypercapnia). Each contraction was 500 ms in duration at 50 Hz. When muscles contracted in normocapnic solution (pH 7.42), the developed tension decreased an average of 6.1 +/- 0.8% over 25 min. For the next 50 min, 15 muscles remained normocapnic, while the remainder contracted in hypercapnic solution (pH 7.20). Tension decreased significantly more with hypercapnia. For the last 25 min, both normocapnic and hypercapnic muscles were divided into three treatment groups (N = 5). One group continued in the same environment, while acetazolamide (final concentration of 10(-5) M) was added to the bath of the second and sodium cyanate (final concentration of 10(-5) M) was added to the bath of the third group. Acetazolamide had no effect on tension in either carbon dioxide environment. Sodium cyanate significantly decreased tension from the hypercapnic control but had no effect in normocapnia. Thus carbonic anhydrase III inhibition with sodium cyanate increased the effect of hypercapnia implying that carbonic anhydrase III assists in the regulation of free hydrogen ion concentration in slow-twitch skeletal muscle.     

Time-resolved X-ray diffraction by skinned skeletal muscle fibers during activation and shortening

Force, sarcomere length, and equatorial x-ray reflections (using synchrotron radiation) were studied in chemically skinned bundles of fibers from Rana temporaria sartorius muscle, activated by UV flash photolysis of a new photolabile calcium chelator, NP-EGTA. Experiments were performed with or without compression by 3% dextran at 4 degrees C. Isometric tension developed at a similar rate (t(1/2) = 40 +/- 5 ms) to the development of tetanic tension measured in other studies (Cecchi et al., 1991). Changes in intensity of equatorial reflections (I(11) t(1/2), 15-19 ms; I(10) t(1/2), 24-26 ms) led isometric tension development and were faster than for tetanus. During shortening at 0.14P(o), I(10) and I(11) changes were partially reversed (18% and 30%, respectively, compressed lattice), in agreement with intact cell data. In zero dextran, activation caused a compression of A-band lattice spacing by 0.7 nm. In 3% dextran, activation caused an expansion of 1.4 nm, consistent with an equilibrium spacing of 45 nm. But, in both cases, discharge of isometric tension by shortening caused a rapid lattice expansion of 1.0-1.1 nm, suggesting discharge of a compressive cross-bridge force, with or without compression by dextran, and the development of an additional expansive force during activation. In contrast to I(10) and I(11) data, these findings for lattice spacing did not resemble intact fiber data.     

Studies of the diffuse x-ray scattering from contracting frog skeletal muscles.

Using x-rays from synchrotron radiation, we studied diffuse scattering, sometimes together with the myosin layer lines. With an area detector, sartorius muscles and a time resolution of 150 ms, earlier results from semitendinosus muscles contracting isometrically at 6 degrees C (Lowy, J., and F. R. Poulsen. 1987. J. Mol. Biol. 194:595-600) were confirmed and extended. Evidence from intensity changes both in the diffuse scattering and in the myosin layer lines showed that the majority of the heads become disordered at peak tetanic tension. With a linear detector and a time resolution of 5 ms, it was found that during tension rise the intensity increase of the diffuse scattering (which amounted maximally to 12% recorded near the meridian) runs approximately 20 ms ahead of the mechanical change, comparing half-completion times. This suggests that an appreciable number of heads change orientation before peak tension is reached. In quick release experiments the diffuse scattering intensity showed very little change. Recorded near the meridian during rapid shortening, however, it decreased progressively with a half-time of approximately 40 ms. This change amounted to approximately 35% of that observed during the initial tension rise. We interpret this to indicate that during rapid shortening a certain number of heads assume an orientation characteristic of the relaxed state. Viewed in the context of the behavior of the first myosin layer line and the (1, 1) equatorial reflection in similar experiments (Huxley, H. E., M. Kress, A. R. Faruqi, and R. M. Simmons. 1988.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of chronic hypobaria on isolated tracheal preparation.

Isolated tracheal preparation (ITP) has been studied from rabbits adapted to 24 days simulated hypobaria (PB = 395 mmHg), corresponding to an elevation of 17,000ft (5,000m). During this daily hypobaric exposure, the growth of these animals was continuous and final hematocrit reached 70%. After 24 days of hypobaric exposure, the sensitivity of ITP was tested to acute hypercapnia and acute anoxia. Acute hypercapnia was induced by aerating a muscle chamber with a gas mixture of 85% oxygen and 15% carbon dioxide. During that time, the PCO2 of the experimental solution increased from 24 to 66 mmHg with a corresponding change of pH from 7.39 to 7.10. ITP from rabbits exposed to chronic hypoxia reveals a greater sensitivity to acute hypercapnia as measured by active tension (AT max) and rate of tension development (dT/dt). Acute anoxia was induced by aerating a muscle chamber with a gas mixture of 95% nitrogen and 5% carbon dioxide. During 20 min of these conditions, the Po2 decreased from 520 to 25 mmHg. Although the contractility diminished at approximately equal rates in both groups in response to acute anoxia, preparation from animals exposed to hypobaria showed statistically a significantly greater ability to recover from the anoxic state.     

Application of high-field 1H-NMR spectroscopy for the study of perifused amphibian and excised mammalian muscles

Frog sartorius and gastrocnemius muscles were perifused at 20 degrees C, the intracellular pH (pHi) and the concentration of phosphocreatine were determined in the resting muscle by 1H-NMR spectroscopy at 470 MHz; values of pHi = 7.31 +/- 0.05 (n = 7) and concentration of phosphocreatine = 20.4 +/- 1.1 mumol/g wet wt. (n = 6) were found. The hydrolysis of phosphocreatine and the simultaneous increase in lactate upon perifusion with 10 mM caffeine (in Ringer's solution) was followed with a time resolution of 1 min. Lactate increased at a rate of 1.0 mumol/g per min, but no pHi change was recorded during the time monitored. The lower limit for the buffering capacity of the muscle cytosol was estimated to be 16.7 mumol/g muscle per pH unit from the uncertainty in pHi determination (+/- 0.03 pH units) and from the amount of lactate produced and phosphocreatine hydrolyzed. Changes in pHi, lactate concentration and fatty acyl chain intensity were monitored by 1H-NMR spectroscopy at 361 MHz in ischemic rat skeletal muscle, excised and stored at 20 degrees C. The resonances in the 1H-NMR spectrum of a human skeletal muscle perchloric acid extract are reported and tentatively assigned.     

The variation of characteristics of twitch and tetanic contractions with sarcomere length in isolated muscle fibres of the frog.

The relation between sarcomere length, tension and time course of tension development in twitch and tetanic contractions at 20 degrees C was determined for isolated fibres from the semitendinosus muscle of the frog (Rana esculenta). In twenty fibres at about 2.15 micron sarcomere length, the peak twitch tension, the maximum tetanic tension and the twitch/tetanus ratio ranged, respectively, from 0.22 to 1.6 kg/cm2, from 2.13 o 3.96 kg/cm2 an from 0.07 to 0.53. The peak twitch tension was found to be: i) directly correlated with the twitch/tetanus ratio and the time to the peak of the first derivative of the twitch tension, ii) inversely correlated with the time to the peak of the first derivative of tetanic tension. No significant correlation was found between the maximal tetanic tension and the peak twitch tension or the twitch/tetanus ratio. Peak twitch tension and twitch/tetanus ratio were not correlated with the fibre cross-sectional area which ranged from 1.052 to 6,283 micron2. Sarcomere length-tension curves for twitch and tetanic isometric contractions at 20 degrees C were determined in twelve fibres. Increases in sarcomere length from about 2.15 to 2.85 micron produced, depending on the peak twitch tension or the twitch/tetanus ratio at about 2.15 micron, either decrease and no change or increase in peak twitch tension, but constantly enhanced the twitch/tetanus ratio and the degree of this potentiation was inversely correlated with the twitch/tetanus ratio at 2.15 micron. Increase in sarcomere length above 2.15 micron did not alter the course of the early development of twitch and tetanic tensions, reduced considerably the variation in peak twitch tension and twitch/tetanus ratio, without altering that of tetanic tension and swamped the correlation between the peak twitch tension and the time to peak of the differentiated twitch tension. However, the peak twitch tension at about 2.85 micron resulted to be directly correlated with the peak twitch tension at about 2.15 micron and in addition the relative length-dependent change in the time of the peak of the first derivative of the twitch tension resulted to be directly correlated with the relative length-dependent change in the peak twitch tension. It is concluded that both the duration of the active state and the rate factors of activation contribute to the determining of the large variation in peak twitch tension at about 2.15 micron, whereas the length-dependent increase in twitch/tetanus ratio appears to be mainly determined by prolongation of the active state duration.     

A strong constant magnetic field affects muscle tension development in bullfrog neuromuscular preparations

Effects of a constant magnetic field (CMF) of 0.65 T on muscle tension over 9 h were studied in the neuromuscular preparation of the bullfrog sartorius muscle. Tension was developed every 30 min by stimulation of the sciatic nerve (nerve stimulation) or of the sartorius muscle itself (muscle stimulation). In sciatic nerve stimulation, tension decreased rapidly for the first 3-4 h at a similar rate in both test (exposed to CMF) and control muscles. However, the rate of decrease became smaller and almost leveled off after 3-4 h in the test muscles, whereas tension continued to decrease monotonically in control muscles. The slope of the decrease for these later periods was significantly different between the test and the control conditions. Accordingly, tension was larger in test than in control muscles. In muscle stimulation, tension decreased monotonically from the start of experiments in control muscles, while tension in test muscles maintained their initial values for almost 3 h. Thereafter they started to decrease with a similar rate to the control. Hence, tension was always larger in test than in control muscles. A similar pattern of temporal change was observed for the rate of rise of the maximum tension for nerve or muscle stimulation. However, a significant difference was detected only in the case of muscle stimulation. The present results showed that a strong CMF of 0.65 T had biological effects on tension development of the bullfrog sartorius muscle by stimulation of the sciatic nerve as well as by stimulation of muscle itself. The presence of a small AC magnetic field component leaves open the possibility of an AC, rather than a CMF effect.     

Thermal stress and Ca-independent contractile activation in mammalian skeletal muscle fibers at high temperatures.

Temperature dependence of the isometric tension was examined in chemically skinned, glycerinated, rabbit Psoas, muscle fibers immersed in relaxing solution (pH approximately 7.1 at 20 degrees C, pCa approximately 8, ionic strength 200 mM); the average rate of heating/cooling was 0.5-1 degree C/s. The resting tension increased reversibly with temperature (5-42 degrees C); the tension increase was slight in warming to approximately 25 degrees C (a linear thermal contraction, -alpha, of approximately 0.1%/degree C) but became more pronounced above approximately 30 degrees C (similar behavior was seen in intact rat muscle fibers). The extra tension rise at the high temperatures was depressed in acidic pH and in the presence of 10 mM inorganic phosphate; it was absent in rigor fibers in which the tension decreased with heating (a linear thermal expansion, alpha, of approximately 4 x 10(-5)/degree C). Below approximately 20 degrees C, the tension response after a approximately 1% length increase (complete < 0.5 ms) consisted of a fast decay (approximately 150.s-1 at 20 degrees C) and a slow decay (approximately 10.s-1) of tension. The rate of fast decay increased with temperature (Q10 approximately 2.4); at 35-40 degrees C, it was approximately 800.s-1, and it was followed by a delayed tension rise (stretch-activation) at 30-40.s-1. The linear rise of passive tension in warming to approximately 25 degrees C may be due to increase of thermal stress in titin (connectin)-myosin composite filament, whereas the extra tension above approximately 30 degrees C may arise from cycling cross-bridges; based on previous findings from regulated actomyosin in solution (Fuchs, 1975), it is suggested that heating reversibly inactivates the troponin-tropomyosin control mechanism and leads to Ca-independent thin filament activation at high temperatures. Additionally, we propose that the heating-induced increase of endo-sarcomeric stress within titin-myosin composite filament makes the cross-bridge mechanism stretch-sensitive at high temperatures.     

Tension relaxation after stretch in resting mammalian muscle fibers: stretch activation at physiological temperatures.

Tension responses to ramp stretches of 1-3% Lo (fiber length) in amplitude were examined in resting muscle fibers of the rat at temperatures ranging from 10 degrees C to 36 degrees C. Experiments were done using bundles of approximately 10 intact fibers isolated from the extensor digitorum longus (a fast muscle) and the soleus (a slow muscle). At low temperatures (below approximately 20 degrees C), the tension response consisted of an initial rise to a peak during the ramp followed by a complex tension decay to a plateau level; the tension decay occurred at approximately constant sarcomere length. The tension decay after a standard stretch at approximately 3-4.Lo/s contained a fast, an intermediate, and a (small amplitude) slow component, which at 10 degrees C (sarcomere length approximately 2.5 microns) were approximately 2000.s-1, approximately 150.s-1, and approximately 25.s-1 for fast fibers and approximately 2000.s-1, approximately 70.s-1 and approximately 8.s-1 for slow fibers, respectively. The fast component may represent the decay of interfilamentary viscous resistance, and the intermediate component may be due to viscoelasticity in the gap (titin, connectin) filament. The two- to threefold fast-slow muscle difference in the rate of passive tension relaxation (in the intermediate and the slow components) compares with previously reported differences in the speed of their active contractions; this suggests that "passive viscoelasticity" is appropriately matched to contraction speed in different muscle fiber types. At approximately 35 degrees C, the fast and intermediate components of tension relaxation were followed by a delayed tension rise at approximately 10.s-1 (fast fibers) and 2.5.s-1 (slow fibers); the delayed tension rise was accompanied by sarcomere shortening. BDM (5-10 mM) reduced the active twitch and tetanic tension responses and the delayed tension rise at 35 degrees C; the results indicate stretch sensitive activation in mammalian sarcomeres at physiological temperatures.     

Positive inotropism in mammalian skeletal muscle in vitro during and after fatigue

We tested the hypothesis that positive inotropic factors decrease fatigue and improve recovery from fatigue in mammalian skeletal muscle in vitro. To induce fatigue, we stimulated mouse soleus and extensor digitorum longus (EDL) to perform isometric tetanic contractions (50 impulses x s(-1) for 0.5 s) at 6 contractions x min(-1) for 60 min in soleus and 3 contractions x min(-1) for 20 min in EDL. Muscles were submerged in Krebs-Henseleit bicarbonate solution (Krebs) at 27 degrees C gassed with 95% nitrogen - 5% carbon dioxide (anoxia). Before and for 67 min after the fatigue period, muscles contracted at 0.6 contractions x min(-1) in 95% oxygen - 5% carbon dioxide (hyperoxia). We added a permeable cAMP analog (N6, 2'-O-dibutyryladenosine 3':5'-cyclic monophosphate at 10(-3) mol x L(-1) (dcAMP)), caffeine (2 x 10(-3) mol x L(-1), or Krebs as vehicle control at 25 min before, during, or at the end of the fatigue period. In soleus and EDL, both challenges added before fatigue significantly increased developed force but only caffeine increased developed force when added during the fatigue period. At the end of fatigue, the decrease in force in challenged muscles was equal to or greater than in controls so that the force remaining was the same or less than in controls. EDL challenged with dcAMP or caffeine at any time recovered more force than controls. In soleus, caffeine improved recovery except when added before fatigue. With dcAMP added to soleus, recovery was better after challenges at 10 min and the end of the fatigue period. Thus, increased intracellular concentrations of cAMP and (or) Ca2+ did not decrease fatigue in either muscle but improved recovery from fatigue in EDL and, in some conditions, in soleus.     

Carbon dioxide induces endotrophic germ tube formation in Candida albicans

Candida albicans formed germ tubes when exposed to air containing 5 to 15% carbon dioxide (CO2). The CO2-mediated germ tube formation occurred optimally at 37 degrees C in a pH range of 5.5 to 6.5. No germ tubes were produced at 25 degrees C, even when the optimal concentration of CO2 (10%) was present in the environment. The requirement of CO2 for germ tube formation could be partially substituted by sodium bicarbonate but not by N2. Carbon dioxide was required to be present throughout the entire course of germ tube emergence suggesting that its role is not limited to an initial triggering of morphogenic change. We suggest that carbon dioxide may be a common effector responsible for the germ tube promoting activity of certain chemical inducers for C. albicans.     

The effect of acid--base changes on skeletal muscle twitch tension.

The effects of acid--base alterations produced by changing bicarbonate (metabolic type), carbon dioxide tension (respiratory type), or both bicarbonate and carbon dioxide tension (compensated type) on skeletal muscle twitch tension, intracellular pH, and intracellular potassium were studied in vitro. Hemidiaphragm muscles from normal rats and rats fed a potassium-deficient diet were used. Decreasing the extracellular pH by decreasing bicarbonate or increasing CO2 in the bathing fluid produced a decrease in intracellular pH, intracellular K+, and muscle twitch tension. However, at a constant extracellular pH, an increase in CO2 (compensated by an increase in bicarbonate) produced an increase in intracellular K+ and twitch tension in spite of a decrease in intracellular pH. The effect on twitch tension of the hemidiaphragms showed a rapid onset, was reversible, persisted until the buffer composition was changed, and was independent of synaptic transmission. It is concluded that the twitch tension of the skeletal muscle decrease with a decrease in intracellular K+. The muscle tension also decreases with an increase in the ratio of intracellular and extracellular H+ concentration. However, there is no consistent relationship between muscle tension and extracellular or intracellular pH. The muscle tension of the diaphragms taken from K+-deficient rats is more sensitive to variations in CO2, PH, and bicarbonate concentration of the medium than that of the control rat diaphragms.     

Effect of fatigue on the compliance series of isolated muscles]

Frog sartorius muscles are stretched at rest and during tetanic contractions, and series-compliance of normal muscle is compared to that of a fatigued one. The series-compliance is calculated from the parallel compliance and the active one. The compliance decreases when the tension increases and becomes quite constant for the highest levels of tension. These results are discussed on the basis of the sliding filaments theory.     

Reactions of ethyleneimine with guanosine and deoxyguanosine

Ethyleneimine was reacted with guanosine in aqueous medium and the products were purified by Sephadex G-10 and high-performance liquid chromatography (HPLC). Two products were identified by UV-spectroscopy: imidazole-ring opened 7-alkylguanosine and 1-alkylguanosine, accounting for 80% and 14% of all adduct radioactivity, respectively. When the incubation with ethyleneimine and guanosine or deoxyguanosine was carried out at pH 6.0 for 1 h intact 7-alkylation products were detected. The half-life of imidazole ring opening of 7-alkylguanosine was 11, 5 and 2.8 min at pH-values 7.0, 7.7 and 8.0, respectively, as measured fluorometrically at 25 degrees C. The respective half-life of alkylated deoxyguanosine was 21 min at pH 7.7. The half-life of depurination of alkylated deoxyguanosine was 42 min at pH 6.0 and 25 degrees C.