Action Potential-Evoked Calcium Release Is Impaired in Single Skeletal Muscle Fibers from Heart Failure Patients (original) (raw)
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The Journal of Physiology, 2004
The mdx mouse, a model of the human disease Duchenne muscular dystrophy, has skeletal muscle fibres which display incompletely understood impaired contractile function. We explored the possibility that action potential-evoked Ca(2+) release is altered in mdx fibres. Action potential-evoked Ca(2+)-dependent fluorescence transients were recorded, using both low and high affinity Ca(2+) indicators, from enzymatically isolated fibres obtained from extensor digitorum longus (EDL) and flexor digitorum brevis (FDB) muscles of normal and mdx mice. Fibres were immobilized using either intracellular EGTA or N-benzyl-p-toluene sulphonamide, an inhibitor of the myosin II ATPase. We found that the amplitude of the action potential-evoked Ca(2+) transients was significantly decreased in mdx mice with no measured difference in that of the surface action potential. In addition, Ca(2+) transients recorded from mdx fibres in the absence of EGTA also displayed a marked prolongation of the slow decay phase. Model simulations of the action potential-evoked transients in the presence of high EGTA concentrations suggest that the reduction in the evoked sarcoplasmic reticulum Ca(2+) release flux is responsible for the decrease in the peak of the Ca(2+) transient in mdx fibres. Since the myoplasmic Ca(2+) concentration is a critical regulator of muscle contraction, these results may help to explain the weakness observed in skeletal muscle fibres from mdx mice and, possibly, Duchenne muscular dystrophy patients.
Effects of Congestive Heart Failure on Ca2+ Handling in Skeletal Muscle During Fatigue
Circulation Research, 2006
Skeletal muscle weakness and decreased exercise capacity are major symptoms reported by patients with congestive heart failure (CHF). Intriguingly, these skeletal muscle symptoms do not correlate with the decreased heart function. This suggests that CHF leads to maladaptive changes in skeletal muscles, and as reported most markedly in slow-twitch muscles. We used rats at 6 weeks after infarction to measure expression of key proteins involved in SR Ca 2ϩ release and uptake in slow-twitch soleus muscles. We also measured force and myoplasmic free [Ca 2ϩ ] ([Ca 2ϩ ] i) in intact single fibers of soleus muscles. CHF rats showed clear signs of severe cardiac dysfunction with marked increases in heart weight and left ventricular end-diastolic pressure compared with sham operated rats (Sham). There were small, but significant, changes in the content of proteins involved in cellular Ca 2ϩ handling in CHF muscles: slight increases in SR Ca 2ϩ release channels (ie, the ryanodine receptors) and in SR Ca 2ϩ-ATPase. Tetanic force and [Ca 2ϩ ] i were not significantly different between CHF and Sham soleus fibers under resting conditions. However, during the stimulation period there was a decrease in tetanic force without changes in [Ca 2ϩ ] i in CHF fibers that was not observed in Sham fibers. The fatigue-induced changes recovered rapidly. We conclude that CHF soleus fibers fatigue more rapidly than Sham fibers because of a reversible fatigue-induced decrease in myofibrillar function.
Circulation Research, 2001
A decreased exercise tolerance is a common symptom in patients with congestive heart failure (CHF). This decrease has been suggested to be partly due to altered skeletal muscle function. Therefore, we have studied contractile function and cytoplasmic free Ca 2ϩ concentration ([Ca 2ϩ ] i , measured with the fluorescent dye indo 1) in isolated muscles from rats in which CHF was induced by ligation of the left coronary artery. The results show no major changes of the contractile function and [Ca 2ϩ ] i handling in unfatigued intact fast-twitch fibers isolated from flexor digitorum brevis muscles of CHF rats, but these fibers were markedly more susceptible to damage during microdissection. Furthermore, CHF fibers displayed a marked increase of baseline [Ca 2ϩ ] i during fatigue. Isolated slow-twitch soleus muscles of CHF rats displayed slower twitch contraction and tetanic relaxation than did muscles from sham-operated rats; the slowing of relaxation became more pronounced during fatigue in CHF muscles. Immunoblot analyses of sarcoplasmic reticulum proteins and sarcolemma Na ϩ ,K ϩ-ATPase showed no difference in flexor digitorum brevis muscles of sham-operated versus CHF rats. In conclusion, functional impairments can be observed in limb muscle isolated from rats with CHF. These impairments seem to mainly involve structures surrounding the muscle cells and sarcoplasmic reticulum Ca 2ϩ pumps, the dysfunction of which becomes obvious during fatigue.
Acta physiologica (Oxford, England), 2007
Aim: The purpose was to evaluate the effects of fatiguing eccentric contractions (EC) on calcium (Ca2+) handling properties in mammalian type I muscles. We hypothesized that EC reduces both endogenous sarcoplasmic reticulum (SR) content of releasable Ca2+ (eSRCa2+) and myofibrillar Ca2+ sensitivity.Methods: Isolated rat soleus muscles performed 30 EC bouts. Single fibres were isolated from the muscle and after mechanical removal of sarcolemma used to measure eSRCa2+, rate of SR Ca2+ loading and myofibrillar Ca2+ sensitivity.Results: Following EC maximal force in whole muscle was reduced by 30% and 16/100 Hz force ratio by 33%. The eSRCa2+ in fibres from non-stimulated muscles was 45 ± 5% of the maximal loading capacity. After EC, eSRCa2+ per fibre CSA decreased by 38% (P = 0.05), and the maximal capacity of SR Ca2+ loading was depressed by 32%. There were no effects of EC on either myofibrillar Ca2+ sensitivity, maximal Ca2+ activated force per cross-sectional area and rate of SR Ca2+ loading, or in SR vesicle Ca2+ uptake and release.Conclusions: We conclude that EC reduces endogenous SR content of releasable Ca2+ but that myofibrillar Ca2+ sensitivity and SR vesicle Ca2+ kinetics remain unchanged. The present data suggest that the long-lasting fatigue induced by EC, which was more pronounced at low frequencies (low frequency fatigue), is caused by reduced Ca2+ release occurring secondary to reduced SR content of releasable Ca2+.
Ca-dependent slow action potentials in human skeletal muscle
Journal of Cellular Physiology, 1988
Slow Ca-action potentials (CaAP) were studied in normal human skeletal muscle fibers obtained during surgery (fibers with both ends cut). Control studies also were carried out with intact as well as cut rat skeletal muscle fibers. Experiments were performed in hypertonic CI-free saline with 10 or 84 m M Ca and K-channel blockers; muscles were preincubated in a saline containing Cs and tetraethylammonium. A current-clamp technique with two intracellular microelectrodes was used. In human muscle, 14.5% of the fibers showed fully developed CaAPs, 21 % displayed nonregenerative Ca responses, and 64.5% showed only passive responses; CaAPs were never observed in 10 m M Ca. In rat muscle, nearly 90% of the fibers showed CaAPs, which were not affected by the cut-end condition. Human and rat muscle fibers had similar membrane potential and conductance in the resting state. In human muscle (22-32" C, 84 m M Ca), the threshold and peak potential during a CaAP were + 26 k 6 mV and + 70 ? 3 mV, respectively, and the duration measured at threshold level was 1.7 2 0.5 sec. In rat muscle, the duration was four times longer. During a CaAP, membrane conductance was assumed to be a leak conductance in parallel with a Ca and a K conductance. In human muscle (22-32" C, 84 m M Ca, 40 p m fiber diameter), values were 0.4 2 0.1 pS, 1.1 f 0.7 p,S, and 0.9 * 0.4 p,S, respectively. Rat muscle (22-24" C, 84 m M Ca) showed leak and K conductances similar to those found in human fibers. Ca-conductance in rat muscle was double the values obtained in human muscle fibers. Ca-dependent action potentials have been observed in mammalian muscle, when C1 was replaced by a
Effects of external calcium deprivation on single muscle fibers
1967
Deprivation of external calcium causes sudden potentiation of the twitch response of single muscle fibers. The potentiation was 64 4-8 %. Potentiation is simultaneous with membrane depolarization occurring after Ca +÷ removal. This depolarization amounted to 9 4-2 my. Ca ++ removal also alters the action potential. 3 rain after calcium withdrawal, action potential amplitude fell by 36 4-3 my; maximum rates of rise and fall of the spike decreased by 55 4-5 and 63 4-5 % respectively. Changes in shape of the A. P. differ from those seen with other potentiators of the twitch response, such as Zn ++. After short exposure to calcium-free media, potassium-induced contractures show potentiation of peak tension. The S-shaped curve relating potassium contracture tension to log [K]o shifts to the left after such treatment. Calcium deprivation also increased the rate of relaxation of the contractures. This effect depends on the duration of calcium deprivation, and is probably related to the effect of calcium lack on the membrane. The change in relaxation occurred immediately after calcium deprivation, and was reversed by sudden readmission of calcium. Relaxation of twitch and tetanus responses also were affected by Ca lack, but not as rapidly as potassium contractures. The results suggest that external calcium is not directly involved in the process responsible for tension development, supporting the view that this process is mediated by translocation of intracellular calcium. The relaxation process, however, appears to be rapidly affected by deprivation of external calcium. Normally the contractile response of muscle fibers is triggered by the lowering of the fiber m e m b r a n e potential to a threshold value at which the excitationcontraction coupling (ECC) process begins to be effective (1-4). T h e hypothesis which postulated t h a t calcium ions entering the fibers from the extracellular space upon depolarization of the m e m b r a n e activated the contractile material (5, 6) appears to be inadequate on the basis of theoretically calculated diffusion delays (4, 7, 8). However, the presence in the interior of the fiber of the system of transverse tubules which open out directly into the external space could reduce such delays (9, 10). a~77
ATP Released by Electrical Stimuli Elicits Calcium Transients and Gene Expression in Skeletal Muscle
Journal of Biological Chemistry, 2009
ATP released from cells is known to activate plasma membrane P2X (ionotropic) or P2Y (metabotropic) receptors. In skeletal muscle cells, depolarizing stimuli induce both a fast calcium signal associated with contraction and a slow signal that regulates gene expression. Here we show that nucleotides released to the extracellular medium by electrical stimulation are partly involved in the fast component and are largely responsible for the slow signals. In rat skeletal myotubes, a tetanic stimulus (45 Hz, 400 1-ms pulses) rapidly increased extracellular levels of ATP, ADP, and AMP after 15 s to 3 min. Exogenous ATP induced an increase in intracellular free Ca 2؉ concentration, with an EC 50 value of 7.8 ؎ 3.1 M. Exogenous ADP, UTP, and UDP also promoted calcium transients. Both fast and slow calcium signals evoked by tetanic stimulation were inhibited by either 100 M suramin or 2 units/ml apyrase. Apyrase also reduced fast and slow calcium signals evoked by tetanus (45 Hz, 400 0.3-ms pulses) in isolated mouse adult skeletal fibers. A likely candidate for the ATP release pathway is the pannexin-1 hemichannel; its blockers inhibited both calcium transients and ATP release. The dihydropyridine receptor co-precipitated with both the P2Y 2 receptor and pannexin-1. As reported previously for electrical stimulation, 500 M ATP significantly increased mRNA expression for both c-fos and interleukin 6. Our results suggest that nucleotides released during skeletal muscle activity through pannexin-1 hemichannels act through P2X and P2Y receptors to modulate both Ca 2؉ homeostasis and muscle physiology.
Shape, Size, and Distribution of Ca2+ Release Units and Couplons in Skeletal and Cardiac Muscles
Biophysical Journal, 1999
Excitation contraction (e-c) coupling in skeletal and cardiac muscles involves an interaction between specialized junctional domains of the sarcoplasmic reticulum (SR) and of exterior membranes (either surface membrane or transverse (T) tubules). This interaction occurs at special structures named calcium release units (CRUs). CRUs contain two proteins essential to e-c coupling: dihydropyridine receptors (DHPRs), L-type Ca 2ϩ channels of exterior membranes; and ryanodine receptors (RyRs), the Ca 2ϩ release channels of the SR. Special CRUs in cardiac muscle are constituted by SR domains bearing RyRs that are not associated with exterior membranes (the corbular and extended junctional SR or EjSR). Functional groupings of RyRs and DHPRs within calcium release units have been named couplons, and the term is also loosely applied to the EjSR of cardiac muscle. Knowledge of the structure, geometry, and disposition of couplons is essential to understand the mechanism of Ca 2ϩ release during muscle activation. This paper presents a compilation of quantitative data on couplons in a variety of skeletal and cardiac muscles, which is useful in modeling calcium release events, both macroscopic and microscopic ("sparks").
PLoS ONE, 2009
Background: Abnormal Ca 2+ transients are often observed in heart muscles under a variety of pathophysiological conditions including ventricular tachycardia. To clarify whether these abnormal Ca 2+ transients can be attributed to abnormal action potential generation or abnormal Ca 2+ handling/excitation-contraction (EC) coupling, we developed a procedure to determine Ca 2+ and action potential signals at the cellular level in isolated heart tissues. Methodology/Principal Findings: After loading ventricular papillary muscle with rhod-2 and di-4-ANEPPS, monowavelength fluorescence images from rhod-2 and ratiometric images of two wavelengths of emission from di-4-ANEPPS were sequentially obtained. To mimic the ventricular tachycardia, the ventricular muscles were field-stimulated in nonflowing Krebs solution which elicited abnormal Ca 2+ transients. For the failed and alternating Ca 2+ transient generation, there were two types of causes, i.e., failed or abnormal action potential generation and abnormal EC coupling. In cells showing delayed initiation of Ca 2+ transients with field stimulation, action potential onset was delayed and the rate of rise was slower than in healthy cells. Similar delayed onset was also observed in the presence of heptanol, an inhibitor of gap junction channels but having a non-specific channel blocking effect. A Na + channel blocker, on the other hand, reduced the rate of rise of the action potentials but did not result in desynchronization of the action potentials. The delayed onset of action potentials can be explained primarily by impaired gap junctions and partly by Na + channel inactivation. Conclusions/Significance: Our results indicate that there are multiple patterns for the causes of abnormal Ca 2+ signals and that our methods are useful for investigating the physiology and pathophysiology of heart muscle.
Proceedings of the National Academy of Sciences, 1997
Applying a brief repolarizing pre-pulse to a depolarized frog skeletal muscle fiber restores a small fraction of the transverse tubule membrane voltage sensors from the inactivated state. During a subsequent depolarizing test pulse we detected brief, highly localized elevations of myoplasmic Ca 2؉ concentration (Ca 2؉ "sparks") initiated by restored voltage sensors in individual triads at all test pulse voltages. The latency histogram of these events gives the gating pattern of the sarcoplasmic reticulum (SR) calcium release channels controlled by the restored voltage sensors. Both event frequency and clustering of events near the start of the test pulse increase with test pulse depolarization. The macroscopic SR calcium release waveform, obtained from the spark latency histogram and the estimated open time of the channel or channels underlying a spark, exhibits an early peak and rapid marked decline during large depolarizations. For smaller depolarizations, the release waveform exhibits a smaller peak and a slower decline. However, the mean use time and mean amplitude of the individual sparks are quite similar at all test depolarizations and at all times during a given depolarization, indicating that the channel open times and conductances underlying sparks are essentially independent of voltage. Thus, the voltage dependence of SR Ca 2؉ release is due to changes in the frequency and pattern of occurrence of individual, voltage-independent, discrete release events.