Rest Decay Research Articles

Rest decay of calcium transients and contractility in feline ventricular myocytes

To define the respective contributions of sarcoplasmic reticulum Ca2+ release and action potential-dependent Ca2+ influx to the activator Ca2+, rest decay of the cytosolic free Ca2+ (Ca2+i) transient and contractility was measured in feline ventricular myocytes loaded with the Ca2(+)-sensitive fluorescent dye indo-1. Myocytes were stimulated to a steady state and then rested for periods ranging from 2 to 120 s. Rest decay was assayed with both resumption of stimulation and rapid extracellular applications of caffeine. The protocol was then repeated after exposure to ryanodine. Ryanodine changed the Ca2+i transients during steady-state stimulation from large, rapidly rising transients starting near the resting Ca2+i level to small, slowly rising Ca2+i transients superimposed on an elevated diastolic Ca2+i. Under control conditions the stimulation- and caffeine-induced Ca2+i transients exhibited slow monotonic rest decay with similar time constants of decay (180 and 202 s, respectively, from a representative cell). After ryanodine, the rest decay of stimulation-induced Ca2+i transients was seemingly no different. However, no Ca2+i transient could be elicited by caffeine after 15 s of rest. These results suggest that 1) under normal conditions the primary source of activator Ca2+ is the sarcoplasmic reticulum, and 2) the rest decay of contractility seen in feline ventricular myocytes results from time-dependent loss of Ca2+ from this organelle.

Functional interconversion of rest decay and ryanodine effects in rabbit and rat ventricle depends on Na/Ca exchange

Rapid cooling contractures were used to assess changes in sarcoplasmic reticulum (SR) Ca content in isolated rabbit and rat ventricular muscle during rest, with altered transsarcolemmal [Na] and [Ca] gradients and in the presence and absence of 100 nM ryanodine. In rabbit there is normally a rest-duration dependent decline in SR Ca content (rest decay), whereas in rat there is a short-term increase in SR Ca content (rest potentiation) and little evidence of rest decay. Ryanodine greatly accelerates the rate of rest decay in rabbit, depleting the SR of Ca in approximately 1 s, whereas in rat, ryanodine does not appear to drain the SR even after a 10 min rest. Elevation of intracellular Na activity in rabbit (by Na-pump inhibition) to a level similar to that measured in control rat during rest (Shattock and Bers, Am. F. Physiol., 256: C813-C822, 1989) makes rest-dependent changes of SR Ca content in these two tissues similar. The rest decay in rabbit in the presence of ryanodine is also markedly slowed after Na-pump inhibition. In rat, reduction of [Ca]0 allows rest decay to occur (+/- ryanodine), but this rest decay can be largely prevented by simultaneous reduction of [Na]o (to maintain [Na]3/[ Ca] constant) which serves to keep the thermodynamic driving force on a 3:1 Na/Ca exchange constant. We conclude that the process of rest decay and rest potentiation in both rabbit and rat ventricle depends on the sarcolemmal Na/Ca exchange. Furthermore, these species can be functionally interconverted by manipulation of the [Na] and [Ca] gradients. The ability of ryanodine to deplete the SR of Ca also depends critically on other transport systems (particularly Na/Ca exchange) to remove Ca from the cytoplasm.

Rapid cooling contractures as an index of sarcoplasmic reticulum calcium content in rabbit ventricular myocytes

Rapid cooling contractures (RCCs) were used to assess changes in sarcoplasmic reticulum (SR) Ca content in both isolated rabbit ventricular myocytes and multicellular preparations. The main difference observed between these preparations was the magnitude of RCCs relative to twitches, apparently due to differences in measured parameters, i.e., unloaded shortening vs. isometric tension. When multicellular preparations were unloaded, RCC shortening was similar to that observed in myocytes. RCC magnitude decreased as the time between the last electrical stimulation and the RCC was increased (rest decay). RCC rest decay closely paralleled that of postrest twitches, suggesting that SR Ca loss is responsible for this process. Paired RCC experiments were used to investigate RCC relaxation and rest decay. When a second RCC (RCC2) was induced immediately after the first (RCC1), a large contracture was still observed (RCC2/RCC1 x 100 = 77.8 +/- 7.3%, mean +/- SD), indicating that the SR resequestered the majority of Ca on rewarming. This fraction was increased (to 92.9 +/- 5.5%) if Na and Ca-free solution was used during RCCs and rewarming, indicating that Na-Ca exchange also contributes to RCC relaxation. Increasing the interval between paired RCCs led to a decrease in RCC2, analogous to rest decay. This rest decay was abolished by inhibiting Na-Ca exchange, indicating that SR Ca loss during rest is mediated primarily by this process. RCCs were abolished by 10 mM caffeine. Ryanodine (1 microM) greatly accelerated RCC rest decay but had less effect on RCCs generated immediately after a train of stimulation. This accelerated rest decay was also dependent on Na-Ca exchange.(ABSTRACT TRUNCATED AT 250 WORDS)

Intracellular Ca2+ transients during rapid cooling contractures in guinea-pig ventricular myocytes.

1. We measured intracellular Ca2+ transients during rapid cooling contractures (RCCs) in guinea-pig ventricular myocytes using the fluorescent Ca2+ indicator, Indo-1. 2. Rapid cooling of myocytes from 22 to 0-1 degrees C induced a rapid increase in [Ca2+]i which preceded the peak of the contraction and was sometimes large enough to saturate Indo-1. This indicates that [Ca2+]i may reach greater than 10 microM during an RCC. 3. The [Ca2+]i during the RCC slowly declined from its peak value and most of this decline in [Ca2+]i can be attributed to slow reaccumulation of Ca2+ by the sarcoplasmic reticulum (SR) in the cold. RCCs induced in the absence of Cao2+, were not different from control, supporting previous conclusions that RCCs depend exclusively on intracellular Ca2+ stores. 4. RCCs are depressed by long rest periods (rest decay) or by exposure to ryanodine or caffeine, which supports conclusions that RCCs are due to Ca2+ release from the SR. The rest decay of RCCs can be almost completely prevented by applying Nao(+)-free solution during the rest period. This implies that the loss of SR Ca2+ during rest depends on the sarcolemmal Na(+)-Ca2+ exchange (and not the sarcolemmal Ca2(+)-ATPase pump). 5. Rapid rewarming during an RCC normally leads to an additional transient contraction (or rewarming spike), without any increase in [Ca2+]i. Thus, the rewarming spike might be attributable to an increase in myofilament Ca2+ sensitivity induced by rewarming. 6. A second RCC is used to assess the fraction of Ca2+ which is re-sequestered by the SR during relaxation from the first RCC. In control solution progressive RCCs decline in amplitude, but in Na(+)-free, Ca2(+)-free solution they are of constant amplitude. We conclude that the SR Ca2+ pump and Na(+)-Ca2+ exchange are responsible for relaxation and that the latter may account for 20-50% of relaxation. 7. These results support the use of RCCs as a useful means of assessing SR Ca2+ content in intact cardiac muscle cells.

SR Ca loading in cardiac muscle preparations based on rapid-cooling contractures

The influence of rest periods on twitches and rapid-cooling contractures (RCCs) was examined in trabeculae from rabbit, rat, guinea pig, and frog ventricle and rabbit atrium. RCCs were used as a relative index of sarcoplasmic reticulum (SR) Ca content. After increasing rest duration, rabbit and guinea pig ventricles exhibit a decline of both twitch force and RCC force (rest decay). When stimulation is resumed, both twitches and RCCs recover to steady-state levels. The SR (and cells) in these tissues may lose Ca during quiescence and become reloaded with progressive stimulation. Rat ventricle and rabbit atrium exhibited an increase in both twitch and RCC tension as a function of rest duration (rest potentiation). Resumption of stimulation resulted in parallel declines of both twitch and RCC tension approaching steady state. Thus stimulation in rat ventricle and rabbit atrium may lead to a net Ca loss from the SR (and the cell) and quiescence may lead to replenishment of cellular Ca. This major difference in Ca metabolism in mammalian cardiac muscles might be due to a fundamental difference in SR properties or, alternatively, different sarcolemmal transport properties (e.g., action potential configuration, Na-pump). After long rest intervals in rabbit and guinea pig ventricle, RCCs return toward their steady-state value in considerably fewer beats than does twitch tension. This implies that something other than SR refilling is responsible for the slow phase of twitch recovery after rest. In rabbit ventricle increasing frequency or extracellular Ca concentration ([Ca]o) generally increases both twitch and RCC tension. However, decreasing [Ca]o (to 0.2 mM) does not decrease RCCs much despite a dramatic decline in twitch tension (suggesting low twitch tension despite a loaded SR). Rapid rewarming during an RCC usually results in a transient rise in tension (or rewarming "spike"), which is due to a warming-induced increase in myofilament Ca sensitivity. Differences in rewarming spikes among the tissues studied suggest differences in temperature effects on myofilament Ca sensitivity.

Spontaneous sarcoplasmic reticulum calcium release in rat and rabbit cardiac muscle: relation to transient and rested-state twitch tension.

Scattered light intensity fluctuations (SLIF) (which monitor spontaneous Ca2+ release from the sarcoplasmic reticulum [SR]) and resting and twitch tension were measured during intervals after stimulation in rat and rabbit papillary muscles. For 1 to 5 seconds after stimulation of rat muscle bathed in 1.5 mM [Ca2+] (Cao), twitch and resting tension are depressed and SLIF are transiently abolished. SLIF and resting tension then recover simultaneously and monotonically but lag behind the restitution of twitch tension. In the absence of further stimulation, SLIF persist and the rested state twitch amplitude remains potentiated. When Cao is increased above 2.5 mM, the restitution of all parameters following stimulation is accelerated and becomes oscillatory; the lag of SLIF and resting tension restitution behind that of the twitch increases such that twitch amplitude increases, overshoots, and decreases to a nadir as SLIF and resting tension reach their initial maximum. In a given muscle, the maximum twitch amplitude occurs at approximately the same level of SLIF; when this level is exceeded, either transiently during monotonic or oscillatory recovery after stimulation in a given Cao or in the steady rested state by an increase in Cao, twitch tension decreases. Ryanodine (1 microM), caffeine (10 mM), or replacement of Cao with strontium abolishes SLIF and causes twitch amplitude to decay with rest. In contrast to rat, twitch amplitude in rabbit muscle bathed in physiological Cao decays with rest and SLIF are nonmeasurable at any interval following stimulation. When Cao is increased to 20 mM during rest, SLIF occur and the rest decay of the twitch is abolished. We interpret the parallel behavior of SLIF and rest potentiation to indicate that in the presence of SLIF, the average SR Ca2+ load within the tissue is high. Depolarization of a tissue exhibiting SLIF causes a large twitch but also a transient depletion of the average SR Ca2+ load. That the restitution of SLIF lags behind recovery of twitch amplitude suggests that the onset of spontaneous SR Ca2+ release requires a delay following SR Ca2+ replenishment. The simultaneous occurrence of spontaneous Ca2+ release in a sufficient number of cells places an upper limit on twitch amplitude either during recovery following stimulation or at rest.

Effect of acetylstrophanthidin on twitches, microscopic tension fluctuations and cooling contractures in rabbit ventricle.

1. We have measured the effect of the aglycone acetylstrophanthidin (ACS) on twitches, cooling contractures and microscopic tension fluctuations in rabbit ventricular muscle. 2. Both developed twitches and cooling contractures are strengthened by applications of ACS in the range 1-4 microM. This positive inotropy averages 150-160% of control (zero ACS) in both twitches and cooling contractures. Cooling contracture magnitude is assumed to reflect the availability of sarcoplasmic reticulum (SR) Ca2+ for contraction (Bridge, 1986). We infer that ACS increases the availability of SR Ca2+ by enlarging SR Ca2+ stores and this may contribute to the positive inotropy. 3. However, twitches appear to increase at lower concentrations of ACS than those required to increase cooling contractures. This observation suggests that the initial ACS inotropy may be achieved without an increase in SR Ca2+. Furthermore, low doses of ACS produce positive inotropy in the presence of 10.0 mM-caffeine where cooling contractures are abolished. This also suggests that positive inotropy occurs in the absence of SR Ca2+ accumulation. 4. Rest decay of both cooling contractures and twitches is significantly slowed in 4 and 8 microM-ACS. We infer that ACS slows the rate of decline of SR Ca2+ available for contraction by slowing the rate at which Ca2+ is lost from the cell during rest. This suggests that ACS produces a net slowing of Ca2+ efflux during activity which in the absence of altered Ca2+ influx will result in net Ca2+ gain and presumably enlarged SR Ca2+ stores. 5. Increasing the concentration of ACS (6-10 microM) results in a decline in developed twitch tension, total tension and an increase in rest tension. Measurement of microscopic tension fluctuations indicates that as developed twitches decline, the root mean square (r.m.s.) of the tension fluctuations increases in a reciprocal manner. This supports the suggestion of others that the decline in developed twitch tension and the appearance of tension fluctuations are causally related. 6. Although ACS (6-10 microM) causes a decline in twitch tension, rapid cooling contractures remain elevated. We suggest that in the presence of Ca2+ oscillations the magnitude of cooling contractures reflects the sum of cytosolic Ca2+ and Ca2+ that is available for release. If microscopic tension fluctuations do represent Ca2+ moving between the SR and cytosol the sum of SR and cytosolic Ca2+ and hence cooling contracture might not decline.(ABSTRACT TRUNCATED AT 400 WORDS)

Ryanodine accelerates rest decay of caffeine-induced but not stimulation-induced calcium transients in feline ventricular myocytes : W.H. duBell and S.R. Houser. Department of Physiology Temple University School of Medicine, Philadelphia, PA, USA

Ryanodine accelerates rest decay of caffeine-induced but not stimulation-induced calcium transients in feline ventricular myocytes : W.H. duBell and S.R. Houser. Department of Physiology Temple University School of Medicine, Philadelphia, PA, USA

Effects of rest duration and ryanodine on changes of extracellular [Ca] in cardiac muscle from rabbits.

Cumulative depletions of extracellular Ca were measured using double-barreled Ca-sensitive microelectrodes in the extracellular space of rabbit ventricular muscle. Depletions were produced by 1-Hz stimulation after rest intervals of 10 s to 10 min. With longer rest intervals, depletion size increased while the first postrest contraction decreased in a reciprocal manner. The depletions may represent refilling of sarcoplasmic reticulum (SR) Ca stores that have become partially depleted of Ca during the rest. Within this interpretive framework, the longer the rest interval the lower the SR Ca content, so the SR is then capable of taking up larger amounts of Ca. This may be related to the rest decay of tension of the first postrest beat, since this is thought to be SR dependent. Ryanodine (1 microM) increased the size of the depletions after short rest intervals (less than 2 min) but not after longer (greater than or equal to 2 min) intervals. Ryanodine also increased the rate of Ca loss from the cell on cessation of stimulation. This increased rate of Ca loss with ryanodine may deplete the SR of Ca such that more Ca can be taken up during subsequent stimulation than in untreated muscles. Thus cumulative depletions after short rest intervals are enhanced by ryanodine. When a Ca load was produced during 1) quiescence [by removal of extracellular Na (Nao)] or 2) continuous stimulation (in the presence of 3 microM acetylstrophanthidin), addition of ryanodine (5-10 microM) did not produce any apparent Ca loss. Caffeine (10 mM), added after ryanodine, induced contractures accompanied by Ca efflux, implying there was Ca in the SR after ryanodine exposure. The results of other investigators have suggested that ryanodine may inhibit cardiac SR Ca release. The present study suggests that ryanodine also enhances the loss of cellular (and probably SR) Ca on cessation of stimulation but not when applied during continuous stimulation or quiescence.

Relationships between the sarcoplasmic reticulum and sarcolemmal calcium transport revealed by rapidly cooling rabbit ventricular muscle.

Rabbit right ventricular papillary muscles were cooled from 30 to approximately 1 degree C immediately after discontinuing electrical stimulation (0.5 Hz). This produced a contracture that was 30-50% of the preceding twitch magnitude and required 20-30 s to develop. The contractures were identical in cooling solutions with normal (144 mM) or low (2.0 mM) Na. They were therefore not Na-withdrawal contractures. Contracture activation was considerably slower than muscle cooling (approximately 2.5 s to cool below 2 degrees C). Cooling contractures were suppressed by caffeine treatment (10.0 mM). Rapid cooling did not cause sufficient membrane depolarization (16.5 +/- 1.2 mV after 30 s of cooling) to produce either a voltage-dependent activation of contracture or a gated entry of Ca from the extracellular space. Contractures induced by treating resting muscles with 5 X 10(-5) M strophanthidin at 30 degrees C exhibited pronounced tension noise. The Fourier spectrum of this noise revealed a periodic component (2-3 Hz) that disappeared when the muscle was cooled. Cooling contractures decayed with rest (t1/2 = 71.0 +/- 9.3 s). This decay accelerated in the presence of 10.0 mM caffeine and was prevented and to some extent reversed when extracellular Na was reduced to 2.0 mM. 20 min of rest resulted in a net decline in intracellular Ca content of 1.29 +/- 0.38 mmol/kg dry wt. I infer that cooling contractures are principally activated by Ca from the sarcoplasmic reticulum (SR). The properties of these contractures suggest that they may provide a convenient relative index of the availability of SR Ca for contraction. The rest decay of cooling contractures (and hence the decay in the availability of activating Ca) is consistent with the measured loss in analytic Ca during rest. The results suggest that contraction in heart muscle can be regulated by an interaction between sarcolemmal and SR Ca transport.

Mean life-time of the neutral pion

The mean life-time of the π0- is measured by the method of Harris, Orear and Taylor. The decay distance of π0-produced by Kπ2-decay at rest (K+→π++π0) and subsequent decay by the Dalitz mode (π0→e++e−+γ) is measured. Using Ilford G-5 emulsions, exposed in the 300 MeV/c separated K+ beam from the Bevatron at Berkeley, 88 of the above events were found. The observed mean π0 life-time is (2.8±0.9)·10−16) s.