Tobacco-specific N-nitrosamines NNN and NNK levels in cigarette brands between 2000 and 2014.

The evolution of the levels of tobacco-specific N-nitrosamines (TSNA), N-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in mainstream (MS) cigarette smoke is investigated based on smoke and tobacco chemistry data of cigarette brands sold by Philip Morris International (PMI) between 2000 and 2014. A total of 315 cigarette samples representing a wide range of product and design characteristics manufactured by PMI between 2008 and 2014 were analyzed and compared to a previously published dataset of PMI brands manufactured in 2000. The data indicate that there is a substantial reduction of NNN and NNK levels in tobacco fillers and MS cigarette smoke per mg of tar and per mg of nicotine using Health Canada Intense (HCI) machine-smoking regime. This observed reduction in NNN and NNK levels in MS cigarette smoke is also supported by the downward trend observed on NNN and NNK levels in USA flue-cured Virginia and Burley tobacco lots from 2000 to 2014 crops, reflecting effectiveness of measures taken on curing and agricultural practices designed to minimize TSNA formation in tobacco.

Cerebral glucose transporters expression and spatial learning in the K-ATP Kir6.2(-/-) knockout mice.

K-ATP channels formed of the Sur and Kir subunits are widely distributed in the brain. Sur1-Kir6.2 is the most common combination of K-ATP channel subunits in the brain and Kir6.2 plays an important role in glucose metabolism through pancreatic insulin secretion or hypothalamic glucose sensing. K-ATP channels have also been reported to play a role in memory processing. Therefore, the aim of the present experiment is to assess the gene and protein expression of GLUT1, GLUT3 and GLUT4 in various brain regions of Kir6.2(-/-) K-ATP knockout mice and to test their working memory performance. GLUT4 was measured using two antibodies, one recognizing an intracellular epitope and the other, an extracellular epitope. Relative to their corresponding wild type, semi-quantitative immunohistochemistry showed that GLUT4 protein expression as measured by a GLUT4 antibody recognizing an extracellular epitope was increased in the Kir6.2(-/-) K-ATP mice. However, there was only a small increase in GLUT4 labeling using the GLUT4 antibody recognizing the intracellular epitope. These results suggest a compensatory higher GLUT4 inclusion at the cellular neuronal membrane in the cerebral cortex, hippocampus and cerebellum of the Kir6.2(-/-) K-ATP knockout mice. However, there was no change in GLUT4 gene expression assessed by TaqMan PCR except for a decrease in the cerebellum of these mice. Working memory performance of the Kir6.2(-/-) K-ATP mice was disrupted at age of 12 weeks but not at 5 weeks. The mild glucose intolerance that is observed in the Kir6.2 knockout mice is unlikely to have created the memory deficits observed. Rather, in light of the effects of K-ATP channel modulators on memory, the memory deficits in the Kir6.2(-/-) K-ATP mice are more likely due to the absence of the Kir6.2 and possible disruption of the GLUT4 activity in the brain.

Treadmill running causes significant fiber damage in skeletal muscle of KATP channel-deficient mice.

Although it has been suggested that the ATP-sensitive K(+) (K(ATP)) channel protects muscle against function impairment, most studies have so far given little evidence for significant perturbation in the integrity and function of skeletal muscle fibers from inactive mice that lack K(ATP) channel activity in their cell membrane. The objective was, therefore, to test the hypothesis that K(ATP) channel-deficient skeletal muscle fibers become damaged when mice are subjected to stress. Wild-type and K(ATP) channel-deficient mice (Kir6.2(-/-) mice) were subjected to 4-5 wk of treadmill running at either 20 m/min with 0 degrees inclination or at 24 m/min with 20 degrees uphill inclination. Muscles of all wild-type mice and of nonexercised Kir6.2(-/-) mice had very few fibers with internal nuclei. After 4-5 wk of treadmill running, there was little evidence for connective tissues and mononucleated cells in Kir6.2(-/-) hindlimb muscles, whereas the number of fibers with internal nuclei, which appear when damaged fibers are regenerated by satellite cells, was significantly higher in Kir6.2(-/-) than wild-type mice. Between 5% and 25% of the total number of fibers in Kir6.2(-/-) extensor digitum longus, plantaris, and tibialis muscles had internal nuclei, and most of such fibers were type IIB fibers. Contrary to hindlimb muscles, diaphragms of Kir6.2(-/-) mice that had run at 24 m/min had few fibers with internal nuclei, but mild to severe fiber damage was observed. In conclusion, the study provides for the first time evidence 1) that the K(ATP) channels of skeletal muscle are essential to prevent fiber damage, and thus muscle dysfunction; and 2) that the extent of fiber damage is greater and the capacity of fiber regeneration is less in Kir6.2(-/-) diaphragm muscles compared with hindlimb muscles.

An ATP-sensitive potassium channel blocker decreases diaphragmatic circulation in anesthetized dogs.

The goal of this study was to determine whether in the dog ATP-sensitive K+ channels blocked with glibenclamide affect diaphragmatic blood flow [phrenic arterial blood flow (Qpa)] during both spontaneous breathing at rest and increased diaphragmatic activity. A control group (no glibenclamide; n = 4) and an experimental group (50 mg/kg of glibenclamide; n = 5) were studied. During spontaneous breathing at rest, Qpa was 15.0 ml.min-1 x 100 g-1 and decreased by 5% in the presence of glibenclamide. Diaphragmatic pacing (30 min-1) generated by phrenic nerve pacing produced an initial diaphragmatic tension-time index of 0.25 in both groups. A 50% decay in transdiaphragmatic pressure was reached at 165 s in the experimental group compared with 421 s in the control group. Diaphragmatic pacing increased Qpa by 46% in the experimental group and 65% in the control group, yielding a 63% greater vascular resistance in the experimental group. Phrenic vein K+ content at rest was unchanged by the presence of glibenclamide, being 3.6 +/- 0.16 mmol/l compared with 3.5 +/- 0.19 mmol/l in the control group. Phrenic nerve pacing in the control group produced a 13% increase in phrenic vein K+ content, whereas in the experimental group a 16% decrease was observed. We suggest that ATP-sensitive K+ channels play an important role in the modulation of Qpa.

Tolbutamide, but not glyburide, affects the excitability and contractility of unfatigued frog sartorius muscle.

The goal of this study was to characterize the effects of tolbutamide and glyburide, two known KATP channel blockers, on intact, unfatigued sartorius muscle fibres of the frog, Rana pipiens. Tetanic contractions were elicited by field stimulation with 200 ms long train of pulses (0.5 ms, 6 V, 140 Hz). Resting and action potentials were measured using conventional microelectrodes. At pHo 7.2 (extracellular pH), the tetanic force was unaffected by 0.5 mM and 1.0 mM tolbutamide, but at 2.0 mM it decreased by 15.5 +/- 1.0%. The effect of tolbutamide on the tetanic force was significantly greater at pHo 6.4: all three tolbutamide concentrations caused a significant decrease in tetanic force, being 62.3 +/- 9.4% at 2 mM. In the presence of tolbutamide a large number of fibres became unexcitable at pHo 6.4, but not at pHo 7.2. Glyburide at 10 microM, on the other hand, caused a 5-7% decrease in tetanic force at both pHo 6.4 and 7.2, but no further decreases in tetanic force were observed when the glyburide concentration was increased up to 100 microM. Unlike tolbutamide, glyburide did not affect the excitability of muscle fibres, but significantly prolonged the repolarization phase of action potentials, especially at pHo 6.4. We suggest that several of the tolbutamide effects reported in this study cannot be accounted for by a direct effect on KATP channels, and that the large decrease in membrane excitability and muscle contractility in the presence of tolbutamide must seriously be taken into consideration when this channel blocker is used to study the physiological role of KATP channels in intact muscle fibres.

Force-length properties and functional demands of cat gastrocnemius, soleus and plantaris muscles.

The purpose of this study was to measure isometric force-length properties of cat soleus, gastrocnemius and plantaris muscle-tendon units, and to relate these properties to the functional demands of these muscles during everyday locomotor activities. Isometric force-length properties were determined using an in situ preparation, where forces were measured using buckle-type tendon transducers, and muscle-tendon unit lengths were quantified through ankle and knee joint configurations. Functional demands of the muscles were assessed using direct muscle force measurements in freely moving animals. Force-length properties and functional demands were determined for soleus, gastrocnemius and plantaris muscles simultaneously in each animal. The results suggest that isometric force-length properties of cat soleus, gastrocnemius and plantaris muscles, as well as the region of the force-length relation that is used during everyday locomotor tasks, match the functional demands.

The effect of lactate on intracellular pH and force recovery of fatigued sartorius muscles of the frog, Rana pipiens.

1. The effects of pHo (extracellular pH) and lactic acid on pHi (intracellular pH) and tetanic force were examined in frog sartorius muscle. Ion-selective microelectrodes were used to measure pHi. Tetanic force was elicited by field stimulation. Experiments were performed in HEPES-buffered solution equilibrated with 100% O2. 2. Mean pHi values (+/- S.E.M.) of unfatigued frog sartorius muscles were 7.14 +/- 0.02 and 7.05 +/- 0.09 at pHo 7.2 and 6.4, respectively. 3. A stimulation at a rate of one 100 ms tetanic contraction per second for 3 min reduced pHi to 6.21 +/- 0.09 and 6.20 +/- 0.04 at pHo 7.2 and 6.4, respectively. Meanwhile at pHo 7.2, the tetanic force (defined as the maximum force developed during a tetanus) decreased by 82.9 +/- 2.6%, the maximum rate of relaxation decreased by 92.9 +/- 0.9%, and the rate constant of the relaxation decreased by 88.5 +/- 1.6%. At pHo 6.4, the decrease in tetanic force, maximum rate of relaxation and rate constant were 90.6 +/- 1.8%, 93.8 +/- 0.5 and 87.5 +/- 2.7%, respectively. 4. The maximum rates of recovery of pHi following fatigue were 0.068 +/- 0.05 and 0.025 +/- 0.05 pH units min-1 at pHo 7.2 and 6.4, respectively. Recovery of normal tetanic force and relaxation rate was also slower at acidic pHo than at neutral pHo. 5. In the presence of 40 mmol l-1 L-lactic acid at pHo 7.2, the maximum rate of pHi recovery following fatigue was only 0.027 +/- 0.03 pH units min-1 at pHo 7.2. The presence of lactic acid also reduced the recovery of the relaxation phase, but not the recovery of tetanic force. 6. It is suggested that pHi recovery is not a limiting factor for tetanic force recovery and that the extracellular H+ inhibits tetanic force recovery by acting at a site located on the outer surface of the sarcolemma. The recovery of the relaxation phase is believed to be pHi dependent.

Modulation of force development by Na+, K+, Na+ K+ pump and KATP channel during muscular activity.

Extracellular K+ concentration increases during exercise and especially during fatigue development. It has been proposed that K+ is an important factor in the etiology of skeletal muscle fatigue because it suppresses membrane excitability and eventually force development. Based on the effect of K+, it has then been proposed the Na+ K+ pump reduces increases in extracellular K+ concentration while the ATP-sensitive K+ channel (KATP channel) allows for rapid increases in extracellular K+ to suppress force development when ATP levels start to fall or when the levels of metabolic end-products become high. However, recent studies have now demonstrated that an increase in extracellular K+ concentration can be advantageous to muscle during exercise because it not only stimulates vasodilation and the exercise pressor reflex, but it also potentiates force development when the Na+ concentration gradient is maintained. A new hypothesis is therefore proposed in which the Na+ K+ pump is important in maintaining the Na+ concentration gradient (and not the K+ concentration gradient as previously suggested), while the activation of KATP channels is important to increase the K+ efflux and extracellular concentration. This situation then optimizes the development of force during exercise. Another hypothesis is proposed in which more KATP channels are activated while the activity of the Na+ K+ pump is reduced when ATP levels start to decrease to allow for an accumulation of intracellular Na+ and further increases in extracellular K+ concentration. These concentration changes then reduce membrane excitability and force development (i.e., fatigue) to protect muscle against large ATP depletion and function impairment.

The effect of K+ on the recovery of the twitch and tetanic force following fatigue in the sartorius muscle of the frog, Rana pipiens.

The goal of this study was to investigate how an increase in the extracellular K+ (K0+) concentration immediately after fatigue affects the recovery of the resting potential, the twitch and tetanic contraction of frog sartorius muscle to further understand the role of K+ in the mechanism of fatigue. Resting potentials were measured with conventional microelectrodes. Twitch and tetanic contractions were elicited by field stimulation. All muscles were fatigued with tetanic contractions at a rate of one contraction per second for 3 min while being exposed to 3 mmole l-1 K0+. During fatigue development the resting potential decreased by 16 mV (control group and pH0 7.2, extracellular pH), while the decrease in the twitch force was 32.8%, compared to 79.3% for the tetanic force, and 84.6% for the maximum rate of force development of the tetanus. Fatigued muscles were also unable to maintain a plateau phase during a tetanus: force declined by 14.8% during this phase. During the recovery period under control conditions (3 mmole l-1 K0+), all four parameters returned to their pre-fatigue values, the recovery of the plateau phase was the fastest (10 min), while that of the twitch force was the slowest (80 min). When K0+ was increased to 7.5 or 9.5 mmole l-1 immediately after fatigue, the recovery rate of the tetanic force and plateau phase was reduced. The maximum rate of force development of the tetanus, however, recovered at a faster rate than control muscles. The recovery of the twitch force was also increased above that of control when K0+ was increased to 9.0 mmole l-1 (a concentration which maximally potentiates the twitch force of unfatigued muscle). Frog sartorius muscles were also tested at pH0 6.4, a pH0 which inhibits force recovery. At that pH0 the effects of K0+ were similar to those observed at pH0 7.2. It is concluded that the role of K+ in muscle fatigue is more complex and may not involve just a contribution to the decrease in force during fatigue development, but may also contribute to an increase in force development under some conditions.

Effects of K+ on the twitch and tetanic contraction in the sartorius muscle of the frog, Rana pipiens. Implication for fatigue in vivo.

The effects of increasing the extracellular K+ concentration on the capacity to generate action potentials and to contract were tested on unfatigued muscle fibers isolated from frog sartorius muscle. The goal of this study was to investigate further the role of K+ in muscle fatigue by testing whether an increased extracellular K+ concentration in unfatigued muscle fibers causes a decrease in force similar to the decrease observed during fatigue. Resting and action potentials were measured with conventional microelectrodes. Twitch and tetanic force was elicited by field stimulation. At pHo (extracellular pH) 7.8 and 3 mmol K+.L-1 (control), the mean resting potential was -86.6 +/- 1.7 mV (mean +/- SEM) and the mean overshoot of the action potential was 5.6 +/- 2.5 mV. An increased K+ concentration from 3 to 8.0 mmol.L-1 depolarized the sarcolemma to -72.2 +/- 1.4 mV, abolished the overshoot as the peak potential during an action potential was -12.0 +/- 3.9 mV, potentiated the twitch force by 48.0 +/- 5.7%, but did not affect the tetanic force (maximum force) and the ability to maintain a constant force during the plateau phase of a tetanus. An increase to 10 mmol K+.L-1 depolarized the sarcolemma to -70.1 +/- 1.7 mV and caused large decreases in twitch (31.6 +/- 26.1%) and tetanic (74.6 +/- 12.1%) force. Between 3 and 9 mmol K+.L-1, the effects of K+ at pHo 7.2 (a pHo mimicking the change in interstitial pH during fatigue) and 6.4 (a pHo known to inhibit force recovery following fatigue) on resting and action potentials as well as on the twitch and tetanic force were similar to those at pHo 7.8. Above 9 mmol K+.L-1 significant differences were found in the effect of K+ between pHo 7.8 and 7.2 or 6.4. In general, the decrease in peak action potential and twitch and tetanic force occurred at higher K+ concentrations as the pHo was more acidic. The results obtained in this study do not support the hypothesis that an accumulation of K+ at the surface of the sarcolemma is sufficiently large to suppress force development during fatigue. The possibility that the K+ concentration in the T tubules reaches the critical K+ concentration necessary to cause a failure of the excitation-contraction coupling mechanism is discussed.

Blocking ATP-sensitive K+ channel during metabolic inhibition impairs muscle contractility.

The objectives of this study were to determine the metabolic conditions in which ATP-sensitive K+ channels (K+(ATP) channels) contribute to a decrease in force. Sartorius muscles of the frog Rana pipiens were subjected to a 60-min metabolic inhibition by exposing them to cyanide (2 mM) and iodoacetate (1 mM). Muscles were exposed to glibenclamide (100 microM) to block K+ATP channels either 60 min before or 8 or 18 min into metabolic inhibition. Resting potentials, action potentials, and membrane conductance were measured using intracellular microelectrodes. Tetanic and resting tension were measured with a force transducer. ATP, ADP, and phosphocreatine (PCr) were measured by high-pressure liquid chromatography. Glibenclamide completely blocked the shortening of action potential but only partially blocked the increase in membrane conductance. When glibenclamide was added 60 min before metabolic inhibition, the decrease in tetanic force was faster than in control muscle (no glibenclamide). This faster decrease in tetanic force was associated with significant membrane depolarizations, greater increases in resting tension, greater depletions of ATP and PCr contents, and greater increases in ADP content. Addition of glibenclamide 8 min into metabolic inhibition caused an increase in tetanic force followed by a faster decrease compared with control. Addition of glibenclamide 18 min into metabolic inhibition had no effect on the tetanic force compared with control muscles. The data indicate that K+ATP channels 1) were activated during metabolic inhibition and 2) contributed to the decrease in tetanic force but also 3) had a myoprotective effect protecting skeletal muscle against muscle function impairment.

Effect of acclimation temperature and pH on contraction of frog sartorius muscle.

The effect of acclimation temperature and pH on the isometric twitch and tetanus of sartorius muscle from frog, Rana pipiens, was studied at different experimental temperatures. Seven variables were measured, namely: tension, latent period, time to maximum tension, half-relaxation time, mean rate, maximum rate, and maximum acceleration of tension development. The effect of experimental temperature was similar to that reported in the literature. The effects of acclimation temperature were small and were not compensatory. Different pH's were obtained by varying CO2 in the gas phase, while the HCO3- concentration was kept constant. The main effects of a decrease in pH on the isometric twitch and tetanus were a reduction in tension and rate of tension development and an increase in latent period. A decrease in pH had no effect on the time to maximum tension or the half-relaxation time. Analysis of variance showed that the test temperature had the greatest effect of all three treatments on each variable, the effects of test and acclimation temperature were dependent on neither the test nor the acclimation temperatures. The in vivo relationships between these three treatments are discussed.

The effects of isotonic contractions on the rate of fatigue development and the resting membrane potential in the sartorius muscle of the frog, Rana pipiens.

The goal of this study was to characterize how isotonic contractions affect the rate of fatigue development. Muscle bundles dissected from frog sartorius muscles were stimulated with 100-ms long train of pulses (0.5 ms, 6 V, 140 Hz). To measure the effect of the isotonic contractions, isometric tetanus were elicited at regular time intervals during the stimulation to fatigue. In general, isotonic contractions caused a faster decrease in tetanic force than isometric contractions. The difference in tetanic force between an isotonic and isometric fatigue increased gradually over a 20-min period to 7.9 and 13.5% at 0.04 and 0.1 trains/s (TPS), respectively. At 0.2, 0.5, and 1.0 TPS, the decrease in tetanic force was also faster during an isotonic fatigue, which resulted in an initial difference in tetanic force between the two types of fatigue. The difference did not exceed 18.5% and did not persist throughout the stimulation period; i.e., the difference disappeared before the end of the fatigue stimulation. The half-relaxation time was prolonged during fatigue development, and the prolongation was greater during an isotonic fatigue, except at 0.04 TPS. The increases in the half-relaxation time at 0.2, 0.5, and 1.0 TPS were followed by a decrease, and the decreases were especially pronounced during an isotonic fatigue at 0.5 and 1.0 TPS. The results showed for the first time that isotonic contractions cause a faster rate of fatigue development in frog sartorius muscles, and this effect depends on the frequency of stimulation.

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.

The effect of acid-base balance on fatigue of skeletal muscle.

H+ ions are generated rapidly when muscles are maximally activated. This results in an intracellular proton load. Typical proton loads in active muscles reach a level of 20-25 mumol X g-1, resulting in a fall in intracellular pH of 0.3-0.5 units in mammalian muscle and 0.6-0.8 units in frog muscle. In isolated frog muscles stimulated to fatigue a proton load of this magnitude is developed, and at the same time maximum isometric force is suppressed by 70-80%. Proton loss is slowed when external pH is kept low. This is paralleled by a slow recovery of contractile tension and seems to support the idea that suppression results from intracellular acidosis. Nonfatigued muscles subjected to similar intracellular proton loads by high CO2 levels show a suppression of maximal tension by only about 30%. This indicates that only a part of the suppression during fatigue is normally due to the direct effect of intracellular acidosis. Further evidence for a component of fatigue that is not due to intracellular acidosis is provided by the fact that some muscle preparations (rat diaphragm) can be fatigued with very little lactate accumulation and very low proton loads. Even under these conditions, a low external pH (6.2) can slow recovery of tension development 10-fold compared with normal pH (7.4). We must conclude that there are at least two components to fatigue. One, due to a direct effect of intracellular acidosis, acting directly on the myofibrils, accounts for a part of the suppression of contractile force. A second, which in many cases may be the major component, is not dependent on intracellular acidosis. This component seems to be due to a change of state in one or more of the steps of the excitation-contraction coupling process. Reversal of this state is sensitive to external pH which suggests that this component is accessible from the outside of the cell.

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.

Use of dP/dt and rise time to estimate speed of shortening in muscle.

We examined the relationship between the maximum speed of shortening at zero load (Vmax) and two variables occasionally used to estimate Vmax: maximal rate of change of force during an isometric tetanic contraction [(dP/dt)max] and reciprocal of one-half rise time (RHRT), the time to achieve one-half the maximal force during an isometric tetanic contraction. The relationship was examined in two experiments on isolated toad sartorius muscle: the effect of temperature and the effect of changing pHe (extracellular pH) to test the hypotheses that (dP/dt)max or RHRT can be used to estimate the magnitude of the effect of experimental variables on Vmax. In the temperature experiment both (dP/dt)max and RHRT could be used to estimate changes in Vmax. The effect of pH on Vmax was markedly overestimated by (dP/dt)max, but there was no significant difference between the magnitude of the changes in Vmax and RHRT. Our results, taken with the results of others, suggest that it is inappropriate to assume that the magnitude of the effect of a particular experimental protocol on Vmax can necessarily be predicted by measuring (dP/dt)max or RHRT.

Effects of step changes in pH on isometric tetanic tension of toad sartorius muscle.

The effect of a rapid change in pHe (pH of bathing solution) on the isometric tetanic tension developed by sartorius muscles of toads acclimated to 5 and 25 degrees C was measured at 5 and 25 degrees C. The pH was altered by changing the carbon dioxide concentration of a bicarbonate buffered physiological solution. Acclimation temperature did not modify the response to a rapid change in pH, but test temperature did. Following a pH decrease from 9.0 to 6.0, tetanic tension decreased at a faster rate at 5 degrees C than at 25 degrees C. A new steady state was reached in 15 min at 5 degrees C but in 40 min at 25 degrees C. Following a pH increase from 6.0 to 8.5, tetanic tension increased at a faster rate at 25 degrees C than at 5 degrees C. A new steady state was reached in 60 min at 5 degrees C but in 10 min at 25 degrees C. We conclude that the rate of carbon dioxide diffusion through the sartorius muscle is only one factor that determines how rapidly tetanic tension changes following the step change in pH, and that muscle resists pH change more effectively at higher temperatures.

The effects of pH on force and stiffness development in mouse muscles.

Changes in force and stiffness during contractions of mouse extensor digitorum longus and soleus muscles were measured over a range of extracellular pH from 6.4 to 7.4. Muscle stiffness was measured using small amplitude (less than 0.1% of muscle length), high frequency (1.5 kHz) oscillations in length. Twitch force was not significantly affected by changes in pH, but the peak force during repetitive stimulation (2, 3, and 20 pulses) was decreased significantly as the pH was reduced. Changes in muscle stiffness with pH were in the same direction, but smaller in extent. If the number of attached cross-bridges in the muscle can be determined from the measurement of small amplitude, high frequency muscle stiffness, then these findings suggest that (a) the number of cross-bridges between thick and thin filaments declines in low pH and (b) the average force per cross-bridge also declines in low pH. The decline in force per cross-bridge could arise from a reduction in the ability of cross-bridges to generate force during their state of active force production and (or) in an increased percentage of bonds in a low force, "rigor" state.