Rest Decay Research Articles

Functional Role of TRPC1 Channels in Neonatal Cardiomyocytes

Transient receptor potential cation (TRPC) 1 channels are thought to play a role in store-operated and receptor-operated calcium entry in various mammalian cell types. However, our understanding of the functional role of these channels in cardiomyocytes remains to be ill-defined. Here, we test the hypothesis that the channels are involved in calcium leak from the sarcoplasmic reticulum (SR) and modulate the cytosolic calcium concentration in cardiomyocytes. Our studies were performed on neonatal rodent ventricular myocytes. After cell isolations, cells were plated on glass slides. Two days after isolation cells were infected with adenoviral vectors of TRPC1 tagged with eGFP. Measurements were performed 4-5 days after infection. Immunolabeling, three-dimensional scanning confocal microscopy and quantitative colocalization analysis revealed an abundant intracellular density of native TRPC1 and TRPC1 expressed via adenoviral vectors. TRPC1 was not associated with the sarcolemma, but the SR. We measured the rest decay and caffeine induced peak calcium release using rapid scanning confocal microscopy on infected cells loaded with the calcium sensitive dye Rhod-3. We found an increased SR calcium content in the presence of the TRPC channel blocker SKF-96365. SR calcium content exhibited a decreasing relationship with TRPC1 expression. In a computational model, activated SR TRPC1 channels increased the systolic and diastolic cytosolic calcium concentration with only minor effects on action potential and SR calcium content. Our studies indicate that TRPC1 channels are not involved in sarcolemmal electrophysiology of rodent ventricular myocytes, but localized in the SR. The studies support our hypothesis that the channels play a role as SR calcium leak channels. The findings could guide us to an understanding of TRPC channels as physiological modulators of intracellular calcium and contractility in cardiac myocytes.

A comparison of patient specific dosimetric calculations obtained by planar images and Monte Carlo simulation in 111In octreotide therapy

s / Physica Medica 30 (2014) e16ee44 e29 phase, as a function of the mean value of administered activity per patient treated each week was also acquired. Results: 675 patients were treated through 2011-2013. Mean administered activity 113,5±31.8mCi. Tank’s filling phase:42±4days.Rest decay duration:63±3days. I-131 activity,T, at the end of tank’s filling phase is given by the formula: T1⁄45.6*Aavg,[mCi] whereas Aavg[mCi] is the average amount of administered activity during tank’s filling time period. Tvalues:560-835 mCi and average monthly concentration 0.13mCi/lt. In accordance with Greek legislation, maximum limit of discharge is 110MBq /day. Liquid samples collected at releasing time documented radioactive releases of I-131, equal to those derived from estimations, much below recommended levels of once release. During the above time period a tank’s operation was interrupted twice as a consequence of clogging. Graphs that depict the measurements beyond are also provided. Conclusions: Above real practice data may be useful for further studies of environmental radiation protection from liquid waste of patients undergoing I-131 therapy for thyroid cancer. Physicists, taking into consideration ALARA principle, economic and social factors, must design in advance, emergency plan assuring that, any individual’s exposure is kept to the minimum practicable, in case of system’s mulfunction. A COMPARISON OF PATIENT SPECIFIC DOSIMETRIC CALCULATIONS OBTAINED BY PLANAR IMAGES AND MONTE CARLO SIMULATION IN 111IN OCTREOTIDE THERAPY Maria Argyrou , Maria Andreou , Nefeli Lagopati , Irini Baka , Ioannis Vamvakas , Maria Lyra . Radiation Physics Unit, A’ Radiology Department, Kapodistrian University of Athens, Aretaieion Hospital, 76 Vas. Sophias Ave, Athens, Greece; Medical Physics Department, Iaso Hospital,

EPAC Does Not Affect Diastolic Sarcoplasmic Reticulum Ca2+ Release in Rabbit Ventricular Myocytes

Recent evidence gathered in ventricular myocytes from rodents points out that the EPAC pathway is a strong promoter of the release of Ca2+ from the sarcoplasmic reticulum (SR; Pereira et al., 2007, J. Physiol. 583:685-94). Encouraged by the above observations, we studied the effects of 2 μM 8-CPT (a specific EPAC activator) on the diastolic SR Ca2+ release in rabbit ventricular myocytes. Our initial studies used epifluorescence and focused on the relationship between the SR load and the diastolic SR Ca2+ release (i.e., the so called SR Ca2+ leak-load relationship). Contrary to the observations in rodents, our rabbit ventricular myocytes displayed no alterations of the leak-load relationship upon 8-CPT application. Since the leak-load relationship requires a steady state to be reached prior to the measurements, we also tested for non steady-state effects of EPAC stimulation using confocal microscopy. We studied the frequency and properties of Ca2+ sparks during the first 30 seconds of rest decay following 2 minutes of 8-CPT application and field stimulation at 1 Hz. Our results showed no effects of EPAC stimulation on the spark frequency or the spatio-temporal properties of the sparks. In summary, our results suggest that EPAC does not affect diastolic SR Ca2+ release in rabbit ventricular myocytes. Future studies will target the species dependence of the effect of EPAC and the effect of SR [Ca2+] upon the release properties under these conditions.

What can we learn from the decay of NX(1625) in the molecule picture?

Considering two assumptions on the molecular state, i.e. the S-wave Λ̄–K- and S-wave Σ̄0–K- molecular states, we study the possible decays of N̄X(1625) that include N̄X(1625)→K-Λ̄,π0p̄,ηp̄,π-n̄. Our results indicate that (1) if N̄X(1625) is the Λ̄–K- molecular state, K-Λ̄ is the main decay mode of N̄X(1625), and the branching ratios of the rest decay modes are tiny; (2) if N̄X(1625) is the Σ̄0–K- molecular state, the branching ratio of N̄X(1625)→K-Λ̄ is one or two orders smaller than that of N̄X(1625)→π0p̄,ηp̄,π-n̄. Thus the search for N̄X(1625)→π0p̄,ηp̄,π-n̄ will be helpful to shed light on the nature of N̄X(1625).

The Na +/Ca 2+ Exchanger/SR Ca 2+ ATPase Transport Capacity Regulates the Contractility of Normal and Hypertrophied Feline Ventricular Myocytes

Background Pressure overload leads to cardiac hypertrophy, which is often followed by heart failure. We tested the hypothesis that depressed contractility in this process results from an imbalance in Ca 2+ transport by the sarcoplasmic reticulum (SR) Ca 2+ ATPase (SERCA) and the sarcolemmal Na +/Ca 2+ exchanger (NCX). Methods and Results Left ventricular (LV) myocytes (n = 79) from 12 normal (N) and 5 hypertrophied (LVH, by aortic banding) feline hearts were studied. Adenoviral gene transfer was used to introduce green fluorescent protein (GFP), SERCA2, and NCX into N and LVH myocytes. Contraction (videomicroscopy) and Ca 2+ transients (Fluo-3) were measured in steady state and after rest periods of 2 to 120 seconds (rest decay and potentiation). LVH hearts were significantly larger than N (7.1 ± 1.4 versus 4.2 ± 0.2 g/kg). SERCA protein was significantly less abundant in LVH versus N. Steady state contractions and Ca 2+ transients of LVH-GFP myocytes decayed more slowly and rest decay of contractility was more pronounced compared with N-GFP. Infection of LVH (and N) myocytes with SERCA increased basal contractility and reduced rest decay. Infection of LVH myocytes with NCX almost abolished contraction and in N myocytes reduced contractility and increased rest decay. Conclusion These findings suggest that an imbalance of Ca 2+ transport by SERCA and the NCX produces the characteristic contractile abnormalities of hypertrophied cardiac myocytes.

A Mathematical Treatment of Integrated Ca Dynamics within the Ventricular Myocyte

We have developed a detailed mathematical model for Ca 2+ handling and ionic currents in the rabbit ventricular myocyte. The objective was to develop a model that: 1), accurately reflects Ca-dependent Ca release; 2), uses realistic parameters, particularly those that concern Ca transport from the cytosol; 3), comes to steady state; 4), simulates basic excitation-contraction coupling phenomena; and 5), runs on a normal desktop computer. The model includes the following novel features: 1), the addition of a subsarcolemmal compartment to the other two commonly formulated cytosolic compartments (junctional and bulk) because ion channels in the membrane sense ion concentrations that differ from bulk; 2), the use of realistic cytosolic Ca buffering parameters; 3), a reversible sarcoplasmic reticulum (SR) Ca pump; 4), a scheme for Na-Ca exchange transport that is [Na] i dependent and allosterically regulated by [Ca] i; and 5), a practical model of SR Ca release including both inactivation/adaptation and SR Ca load dependence. The data describe normal electrical activity and Ca handling characteristics of the cardiac myocyte and the SR Ca load dependence of these processes. The model includes a realistic balance of Ca removal mechanisms (e.g., SR Ca pump versus Na-Ca exchange), and the phenomena of rest decay and frequency-dependent inotropy. A particular emphasis is placed upon reproducing the nonlinear dependence of gain and fractional SR Ca release upon SR Ca load. We conclude that this model is more robust than many previously existing models and reproduces many experimental results using parameters based largely on experimental measurements in myocytes.

Impaired contractile performance of cultured rabbit ventricular myocytes after adenoviral gene transfer of Na(+)-Ca(2+) exchanger.

Na(+)-Ca(2+) exchanger (NCX) gene expression is increased in the failing human heart. We investigated the hypothesis that upregulation of NCX can induce depressed contractile performance. Overexpression of NCX was achieved in isolated rabbit ventricular myocytes through adenoviral gene transfer (Ad-NCX). After 48 hours, immunoblots revealed a virus dose-dependent increase in NCX protein. Adenoviral beta-galactosidase transfection served as a control. The fractional shortening (FS) of electrically stimulated myocytes was analyzed. At 60 min(-1), FS was depressed by 15.6% in the Ad-NCX group (n=143) versus the control group (n=163, P:<0.05). Analysis of the shortening-frequency relationship showed a steady increase in FS in the control myocytes (n=26) from 0.027+/-0.002 at 30 min(-1) to 0. 037+/-0.002 at 120 min(-1) (P:<0.05 versus 30 min(-1)) and to 0. 040+/-0.002 at 180 min(-1) (P:<0.05 versus 30 min(-1)). Frequency potentiation of shortening was blunted in NCX-transfected myocytes (n=27). The FS was 0.024+/-0.002 at 30 min(-1), 0.029+/-0.002 at 120 min(-1) (P:<0.05 versus 30 min(-1), P:<0.05 versus control), and 0. 026+/-0.002 at 180 min(-1) (NS versus 30 min(-1), P:<0.05 versus control). Caffeine contractures, which indicate sarcoplasmic reticulum Ca(2+) load, were significantly reduced at 120 min(-1) in NCX-transfected cells. An analysis of postrest behavior showed a decay of FS with longer rest intervals in control cells. Rest decay was significantly higher in the Ad-NCX group; after 120 seconds of rest, FS was 78+/-4% in control and 65+/-3% in the Ad-NCX group (P:<0.05) relative to steady-state FS before rest (100%). In conclusion, the overexpression of NCX in rabbit cardiomyocytes results in the depression of contractile function. This supports the hypothesis that upregulation of NCX can result in systolic myocardial failure.

Monensin-induced reversal of positive force-frequency relationship in cardiac muscle: role of intracellular sodium in rest-dependent potentiation of contraction.

We have investigated the role of a rest-dependent inotropic factor in determining species-related differences in cardiac force-frequency relationships (FFR). Isolated rat, rabbit or guinea-pig papillary muscles, as well as guinea-pig ventricular myocytes were superfused with 1.8 mM Ca2+ Tyrode. In rat muscles, isometric force amplitude decreased, while in rabbit or guinea-pig muscles force increased with frequency (0.02-1 Hz). Paired-pulse pacing potentiated contraction markedly at all frequencies in rabbit muscles, but not at low frequencies in rat muscles. We tested the hypothesis that high intracellular Na+ levels (Nai) are responsible for negative FFR. The ionophore monensin increased Nai, reversed the FFR of rabbit and guinea-pig muscles from positive to negative, by increasing force mostly at low frequencies, and decreased the paired-pulse potentiation of contraction at low frequencies. Monensin added during rest also reversed rest-induced decay. In isolated myocytes, monensin had qualitatively similar effects on cell shortening as well as on Cai transients. Monensin also decreased the action potential duration (APD) but did not change the pattern of its variation with frequency. Cells intracellularly dialyzed with 20 mM Na+ via a patch pipette also showed rest potentiation of the Cai transients, in contrast to cells dialyzed with 10 mM Na+, which showed rest decay of the transients. APD was also shorter in myocytes dialyzed with 20 mM Na+ than in those dialyzed with lower Na+. The results indicate that in the presence of high Nai, sarcoplasmic reticular Ca2+ load is increased during diastole, possibly via reverse-mode Na+/Ca2+ exchange, and therefore that Nai is an important factor determining the FFR. In addition, the data suggest that short APDs in preparations showing negative FFR may be partly a consequence of increased Nai.

Effects of FK-506 on contraction and Ca2+ transients in rat cardiac myocytes.

FK-506 binding protein (FKBP) has been reported to be closely associated with the ryanodine receptor in skeletal and cardiac muscle and to modulate sarcoplasmic reticulum (SR) Ca2+ release channel gating in isolated channels. FK-506 can inhibit the activity of FKBP, thereby reversing its effects on SR Ca2+ release. We investigated the function of FKBP during normal contractions and Ca2+ transients in intact rat ventricular myocytes loaded with fluorescent Ca2+ indicators. FK-506 significantly increased steady state twitch Ca2+ transients and contraction amplitudes even under conditions in which the SR Ca2+ load and Ca2+ current were unaltered, suggesting that FK-506 increases the fraction of SR Ca2+ released during excitation-contraction (E-C) coupling. Action potentials were somewhat prolonged, consistent with the larger Ca2+ transients causing greater inward Na(+)-Ca2+ exchange current. FK-506 did not affect SR Ca2+ uptake but modestly decreased Ca2+ extrusion via Na(+)-Ca2+ exchange in intact cells (although no effect on Na(+)-Ca2+ exchange was seen in sarcolemmal vesicles). In most cells, FK-506 caused an increase in SR Ca2+ content during steady state stimulation, as assessed by caffeine-induced contractures. This was probably due to the inhibition of Ca2+ efflux via Na(+)-Ca2+ exchange. FK-506 also accelerated the rest decay of SR Ca2+ content and increased the frequency of resting Ca2+ sparks about fourfold. The increase in frequency of these basic Ca2+ release events was not associated with changes in the amplitude or duration of the Ca2+ sparks. We conclude that FK-506 increases the fraction of SR Ca2+ released during normal twitches and enhances the rate of SR Ca2+ release during rest. FK-506 also inhibits Na(+)-Ca2+ exchange, although this effect may be indirect. These effects are consistent with an important SR-stabilizing effect of FKBP in intact rat ventricular myocytes.

Diminished post-rest potentiation of contractile force in human dilated cardiomyopathy. Functional evidence for alterations in intracellular Ca2+ handling.

Post-rest contractile behavior of isolated myocardium indicates the capacity of the sarcoplasmic reticulum (SR) to store and release Ca2+. We investigated post-rest behavior in isolated muscle strips from nonfailing (NF) and endstage failing (dilated cardiomyopathy [DCM]) human hearts. At a basal stimulation frequency of 1 Hz, contractile parameters of the first twitch after increasing rest intervals (2-240 s) were evaluated. In NF (n = 9), steady state twitch tension was 13.7 +/- 1.8 mN/mm2. With increasing rest intervals, post-rest twitch tension continuously increased to maximally 29.9 +/- 4.1 mN/mm2 after 120s (P < 0.05) and to 26.7 +/- 4.5 mN after 240 s rest. In DCM (n = 22), basal twitch tension was 10.0 +/- 1.5 mN/mm2 and increased to maximally 13.6 +/- 2.2 mN/mm2 after 20 s rest (P < 0.05). With longer rest intervals, however, post-rest twitch tension continuously declined (rest decay) to 4.7 +/- 1.0 mN/mm2 at 240 s (P < 0.05). The rest-dependent changes in twitch tension were associated with parallel changes in intracellular Ca2- transients in NF and DCM (aequorin method). The relation between rest-induced changes in twitch tension and aequorin light emission was similar in NF and DCM, indicating preserved Ca(2-)-responsiveness of the myofilaments. Ryanodine (1 microM) completely abolished post-rest potentiation. Increasing basal stimulation frequency (2 Hz) augmented post-rest potentiation, but did not prevent rest decay after longer rest intervals in DCM. The altered post-rest behavior in failing human myocardium indicates disturbed intracellular Ca2- handling involving altered function of the SR.

Regional differences in rest decay and recoveries of contraction and the calcium transient in rabbit ventricular muscle

The rates of rest decay (for rest periods of between 0.5 min and 10 min) and recovery from the rested state (following 10 min of rest) of cell shortening and the amplitude of the intracellular calcium transient were compared in epicardial and endocardial ventricular myocytes isolated from rabbit hearts. The object of these experiments was to determine whether reported transmural differences in action potential duration, myosin type expression and metabolic enzyme content are able to influence the control of contraction. Cells isolated from these two regions of the ventricular wall displayed almost identical twitch shortening and calcium transient characteristics during steady-state electrical stimulation at 0.5 Hz. Despite this, rest decay of cell shortening was faster and recovery from the rested state slower in endocardial cells than in epicardial cells. Neither of these differences could be explained in terms of changes of calcium transient amplitude or time course. We tried to mimic the effect of prolonged rest by application of caffeine to empty the sarcoplasmic reticulum of calcium. The regional differences in recovery of contraction from the rested state were not reproduced in the recovery of contraction after caffeine application, suggesting that the effect is produced by something other than refilling of the sarcoplasmic reticulum. It is suggested that changes in factors that affect myofilament calcium sensitivity produce the regional differences in rest decay and post-rest recovery of contraction.

Sarcoplasmic reticulum-related changes in cytosolic calcium in pressure-overload-induced feline LV hypertrophy.

Alterations in Ca2+ homeostasis that involve the sarcoplasmic reticulum (SR) were studied in feline left ventricular (LV) myocytes isolated from hearts with LV hypertrophy induced by slow, progressive pressure overload. At death, severe hypertrophy was evidenced by increased heart weight-to-body weight ratio (8.4 +/- 0.6 vs. 4.2 +/- 0.2 g/kg in controls). Steady-state Ca2+ transients (measured as. indo 1 fluorescence at 410 nm/480 nm; I410/I480) in LV hypertrophy (LVH) myocytes had diminished peak amplitudes (I410/I480 2.28 +/- 0.07 vs. 2.53 +/- 0.07 in controls) and prolonged durations (0.75 +/- 0.03 vs. 0.59 +/- 0.02 s in controls). The magnitude of shortening was reduced and the contractile duration was prolonged in LVH myocytes. The idea that changes in SR function are responsible for these alterations in the Ca2+ transient was tested by studying two aspects of SR-related Ca2+ homeostasis. Restitution of releasable SR Ca2+ was studied by measuring indo 1 transients and contractions during premature beats. The time course of restitution of both the indo 1 transient and contraction of hypertrophy myocytes was significantly slower than in controls. These data suggest that restitution of releasable SR Ca2+ is slowed in hypertrophy myocytes. The reduction of the indo 1 transient and contraction in beats following long rest periods (rest decay) was measured to determine the rate of Ca2+ loss from the SR. Rest decay was significantly (P < 0.05) more pronounced in hypertrophy myocytes, suggesting that Ca2+ loss from the SR is accelerated in these myocytes. (ABSTRACT TRUNCATED AT 250 WORDS)

Effects of thapsigargin and cyclopiazonic acid on twitch force and sarcoplasmic reticulum Ca2+ content of rabbit ventricular muscle.

Thapsigargin (TG) and cyclopiazonic acid (CPA) are reported to be specific high-affinity inhibitors of the sarcoplasmic reticulum (SR) Ca2+ pump in isolated membranes and cells, with TG causing complete pump inhibition at nanomolar concentrations. To evaluate the effectiveness of TG and CPA in small multicellular cardiac preparations, we used rapid cooling contractures (RCCs) to assess the SR Ca2+ load. In contrast to observations in single myocytes, TG caused remarkably slow and incomplete SR Ca2+ depletion in multicellular preparations. A 45-minute exposure to 500 microM TG at 30 degrees C and 0.5-Hz stimulation only decreased RCCs by 76 +/- 5% (and 100 microM CPA reduced RCCs by 59 +/- 10% [mean +/- SEM]). In contrast, 10 minutes with 20 mM caffeine completely abolished RCCs. This confirms that there was still a caffeine-sensitive pool of Ca2+ in the TG-treated muscle. The time constant of rest decay of RCCs was accelerated by both TG (from 83 +/- 18 to 26 +/- 6 seconds) and CPA (from 68 +/- 11 to 10 +/- 5 seconds). This might be expected since Ca2+ leaking from the SR during rest cannot be taken back up as efficiently, favoring Ca2+ extrusion by the sarcolemmal Na(+)-Ca2+ exchanger. TG and CPA decreased twitch force (by 44 +/- 7% and 40 +/- 11%, respectively) and increased twitch duration, presumably because of the SR effects. We conclude that complete blockade of SR Ca2+ uptake by TG or CPA in multicellular preparations cannot be assumed, even at high [TG] or [CPA], unless evaluated (eg, by RCC).

Paradoxical Twitch Potentiation After Rest in Cardiac Muscle: Increased Fractional Release of SR Calcium

Rest interval dependent changes in contractile force (rest decay and rest potentiation) were studied in rabbit, rat and ferret ventricular muscle and myocytes. The SR Ca content was assessed by rapid cooling contractures or caffeine induced contractures. Intracellular Ca transients, action potentials and Ca current were also recorded. Rest decay of twitches ventricle are roughly paralleled by a decline in SR Ca content. Rat ventricle exhibits primarily rest potentiation, which is not necessarily paralleled by an increased SR Ca content. Ferret ventricle exhibits both rest potentiation and rest decay. However, the SR Ca content in ferret appears to decline monotonically throughout the rest. It is demonstrated that the rest potentiation is not due to an increase in Ca current or in action potential duration. We conclude that there is increase in the fraction of SR Ca content which is released the time that rest potentiation develops. The differences in post-rest contractile function among different cardiac preparations can be described by a simple unifying mechanistic model. In this model there is an exponential time dependent recovery of the ability of the SR to release Ca (e.g. recovery from inactivation) which can be considered to increase the fraction os SR Ca release in response to activation. This fractional SR Ca release is multiplied by the SR Ca content (which may decline exponentially) to provide a measure of the Ca available for activation of contractile force.

Rested-state contractions and rest potentiation in spontaneously hypertensive rats.

To gain further insight into the excitation-contraction coupling mechanisms in hypertrophy, we studied rested-state contractions, rest decay curves, and rest potentiation under different experimental conditions using papillary muscles of spontaneously hypertensive rats (SHR) and age-matched normotensive Wistar and Wistar-Kyoto (WKY) rats. Under constant stimulation at 1.1 Hz, contractility and relaxation were not significantly different in hypertensive when compared with normotensive animals. Rested-state contraction (the first beat after a rest interval of 15 minutes) increased to 159.2 +/- 23% and 123.5 +/- 7.5% of prerest values in Wistar and WKY rats, respectively, whereas in SHR it did not differ from prerest values (92.8 +/- 9.8%). Ryanodine, used to preferentially inhibit sarcoplasmic reticulum function, eliminated the differences in rested-state contractions observed between hypertensive and normotensive rats. Maximal rest potentiation (the first beat after a rest interval of 1 minute) was also significantly higher in Wistar and WKY rats than in SHR. These differences persisted at low extracellular Na+, when Ca2+ efflux via the Na(+)-Ca2+ exchanger was inhibited. Rest decay curves (the decay in contractility from maximal rest potentiation to rested-state contraction) showed a similar pattern in the three rat strains. The results suggest that the altered inotropic responses of the SHR arise from an alteration in calcium handling by the sarcoplasmic reticulum. Experiments on saponin-skinned trabeculae indicated that fractional calcium release induced by caffeine was significantly reduced in the SHR. We conclude that the altered inotropic response observed in SHR may reflect a diminished release of calcium from the sarcoplasmic reticulum.

Cellular mechanisms of endotoxin-induced myocardial depression in rabbits.

We investigated the mechanisms by which endotoxic shock induces intrinsic myocardial depression by studying cardiac myocytes isolated from 10 anesthetized instrumented rabbits given 172 +/- 42 (mean +/- SD) micrograms/kg IV endotoxin. Left ventricular (LV) depression developed 4 +/- 1 hours after endotoxin administration, with a 15 +/- 4% increase in LV internal end-systolic diameter, measured with sonomicrometers at a matched LV end-systolic pressure of 65 +/- 10 mm Hg. Normal LV pressure, arterial PO2, and pH were maintained to minimize confounding effects of ischemia, hypoxia, and acidosis. Cardiac myocytes from endotoxin-exposed rabbits had less unloaded cell shortening and lower peak rates of cell shortening (-dL/dt) and lengthening (+dL/dt) at [Ca2+] levels ranging from 0.5 to 16 mM when compared with myocytes isolated from normal rabbits or rabbits undergoing an identical protocol but without exposure to endotoxin. At 2 mM [Ca2+], cell shortening was depressed by approximately 25% because of a decrease in action potential duration (207 +/- 70 versus 375 +/- 64 milliseconds). In contrast, there was only mild impairment of sarcoplasmic reticulum (SR) function. When myocytes were restimulated after rest periods of 4 to 480 seconds, the decrement in cell shortening (rest decay), peak -dL/dt and peak +dL/dt, and the recovery from rest decay were similar in myocytes from endotoxin-treated and normal rabbits. There was a greater decrement in cell shortening in the second beat of postrest recovery in myocytes from endotoxin-treated rabbits than in normal myocytes. This was partly due to a 12% decrement in action potential duration with rest decay, which did not occur in normal myocytes. The SR Ca2+ content assessed by contractures in 10 mM caffeine was similar in the two groups. We conclude that endotoxic shock produces a LV depression in vivo that persists in isolated myocytes studied in vitro. This intrinsic myocardial depression is largely related to endotoxin-mediated sarcolemmal alterations, which shorten action potential duration, and is not due to alterations in SR function.

The Effect of Thapsigargin on Sarcoplasmic Reticulum Ca 2+ Content and Contractions in Single Myocytes of Guinea-pig Heart

Contractures initiated by 1 s superfusion of single myocytes of guinea-pig heart with 10 mM caffeine were used as a relative index of the SR Ca 2+ content. Thapsigargin (Tg) in concentration 2 × 10 -7M completely blocked Ca 2+ uptake during electrical stimulation by the SR from which Ca 2+ has been previously depleted by caffeine. Tg did not affect the SR Ca 2+ content in the resting myocytes and did not block release of the SR Ca 2+ during electrically stimulated contractions (ESCs). It is concluded that in guinea-pig myocytes Tg affects SR Ca 2+ by selective blocking the SR Ca 2+ uptake. The amplitude of steady state ESCs dropped to 68±5.4% (S.D., n=20) of that of the pre-Tg control. Time to peak contraction increased from 454±82.4 ms to 820±157.4 ms and time of relaxation increased from 368±90.8 ms to 474±87 ms after the SR Ca 2+ has been depleted by Tg. Rest decay of contractions, post-extrasystolic potentiation and post-rest potentiation were inhibited.

New monochromatic muon beam channel using two body decay K + → μ+v at rest in a production target

A new type of monochromatic μ + beam channel of high resolution was designed and constructed at the KEK 12 GeV proton synchrotron (KEK-PS). The principle of the channel is to transport 236 MeV/ c μ + emitted in the two body at rest decay of positive kaons, K + → μ + v, in the production target as a beam. In order to eliminate background μ + from in-flight π + decay, the channel has the following features. Firstly, the μ + beam is extracted at a backward angle of 147° from the direction of the 12 GeV primary proton beam, where the π + yield is relatively small. Secondly, the distance from the production target to the first bending magnet is designed to be as short as possible. Thirdly, the high momentum resolution of the beam channel also enables us to observe a very sharp peak in the μ + momentum spectrum, so that the background is efficiently subtracted. The typical intensity of the 236 MeV/ c monochromatic μ + is 3500 per 10 12 incident protons on a thin Pt target. The signal-to-background ratio finally achieved at the peak momentum was 60. This pure Kμ 2 muon beam was successfully utilized for an experiment designed to search for V + A currents in the weak interaction from a precise determination of the μ + longitudinal polarization.

Does cell size affect the rest decay of contractility in guinea-pig myocytes?

Does cell size affect the rest decay of contractility in guinea-pig myocytes?

Contraction and sarcoplasmic reticulum Ca2+ content in single myocytes of guinea pig heart: effect of ryanodine

The relationship between the ability of sarcoplasmic reticulum (SR) to accumulate and retain Ca2+ and the electrically stimulated contractions (ESCs) of isolated cells from guinea pig ventricular myocardium was investigated. Caffeine contractures or rapid cooling contractures were used as a relative measure of the SR Ca2+ content. Depletion of SR Ca2+ by short exposure to caffeine (15 mM) or by prolonged rest resulted in a reduction of the amplitude of the ESCs by 83 +/- 14 and 65 +/- 11% (means +/- SD), respectively. This result points to SR as a major source of the Ca2+ that activates contraction. However, depriving the SR of the ability to retain Ca2+ by means of prolonged (up to 75 min) exposure to 0.1 microM ryanodine (as shown by the absence of contractile response to caffeine or cooling) did not prevent an ESC of nearly normal amplitude (81 +/- 24% control), albeit with a reduced contraction velocity and a time to peak contraction prolonged by 51 +/- 11%. Additionally, while rest decay of ESCs was present after ryanodine treatment, the time for the ESCs to recover their steady-state amplitude was prolonged at least twofold. Thus, in contrast with the normal guinea pig cells, ESCs of the myocytes exposed to ryanodine are controlled by sarcolemmal processes. This change in the state of excitation-contraction coupling results mainly in modification of the time course of the ESCs and of the time course of the response of the cells to the change in the rate of stimulation.