Membrane polarization at the excitation threshold induced by external electric fields in cardiomyocytes of rats at different developmental stages.

External electric fields (E), used for cardiac pacing and defibrillation/cardioversion, induce a spatially variable change in cardiomyocyte transmembrane potential (ΔVm) that depends on cell geometry and E orientation. This study investigates E-induced ΔVm in cardiomyocytes from rats at different ages, which show marked size/geometry variation. Using a tridimensional numerical electromagnetic model recently proposed (NM3D), it was possible: (a) to evaluate the suitability of the simpler, prolate spheroid analytical model (PSAM) to calculate amplitude and location of ΔVm maximum (ΔVmax) for E = 1 V.cm-1; and (b) to estimate the ΔVmax required for excitation (ΔVT) from experimentally determined threshold E values (ET). Ventricular myocytes were isolated from neonatal, weaning, adult, and aging Wistar rats. NM3D was constructed as the extruded 2D microscopy cell image, while measured minor and major cell dimensions were used for PSAM. Acceptable ΔVm estimates can be obtained with PSAM from paralelepidal cells for small θ. ET, but not ΔVT, was higher for neonate cells. ΔVT was significantly greater in the cell from older animals, which indicate lower responsiveness to E associated with aging, rather than with altered cell geometry/dimensions. ΔVT might be used as a non-invasive indicator of cell excitability as it is little affected by cell geometry/size.

Developmental differences in myocardial transmembrane Na+ transport: implications for excitability and Na+ handling.

Little is currently known about possible developmental changes in myocardial Na+ handling, which may have impact on cell excitability and Ca2+ content. Resting intracellular Na+ concentration ([Na+ ]i ), measured in freshly isolated rat ventricular myocytes with CoroNa green, was not significantly different in neonates (3-5 days old) and adults, but electrical stimulation caused marked [Na+ ]i rise only in neonates. Inhibition of L-type Ca2+ current by CdCl2 abolished not only systolic Ca2+ transients, but also activity-dependent intracellular Na+ accumulation in immature cells. This indicates that the main Na+ influx pathway during activity is the Na+ /Ca2+ exchanger, rather than voltage-dependent Na+ current (INa ), which was not affected by CdCl2 . In immature myocytes, INa density was two-fold greater, inactivation was faster, and the current peak occurred at less negative transmembrane potential (Em ) than in adults. Na+ channel steady-state activation and inactivation curves in neonates showed a rightward shift, which should increase channel availability at diastolic Em , but also require greater depolarization for excitation, which was observed experimentally and reproduced in computer simulations. Ventricular mRNA levels of Nav 1.1, Nav 1.4 and Nav 1.5 pore-forming isoforms were greater in neonate ventricles, while a decrease was seen for the ß1 subunit. Both molecular and biophysical changes in the channel profile may contribute to the differences in INa density and voltage-dependence, and also to the less negative threshold Em , in neonates compared to adults. The apparently lower excitability in immature ventricle may confer protection against the development of spontaneous activity in this tissue. KEY POINTS: Previous studies showed that myocardial preparations from immature rats are less sensitive to electrical field stimulation than adult preparations. Freshly isolated ventricular myocytes from neonatal rats showed lower excitability than adult cells, e.g. less negative threshold membrane potential and greater membrane depolarization required for action potential triggering. In addition to differences in mRNA levels for Na+ channel isoforms and greater Na+ current (INa ) density, Na+ channel voltage-dependence was shifted to the right in immature myocytes, which seems to be sufficient to decrease excitability, according to computer simulations. Only in neonatal myocytes did cyclic activity promote marked cytosolic Na+ accumulation, which was prevented by abolition of systolic Ca2+ transients by blockade of Ca2+ currents. Developmental changes in INa may account for the difference in action potential initiation parameters, but not for cytosolic Na+ accumulation, which seems to be due mainly to Na+ /Ca2+ exchanger-mediated Na+ influx.

Accuracy of electromagnetic models to estimate cardiomyocyte membrane polarization.

External electric fields (E) induce a spatially heterogeneous variation in the membrane potential (ΔVm) of cardiomyocytes that, if sufficiently large, results in an action potential and contraction. Insights into the phenomenon of ΔVm induction by E have been classically gained with electromagnetic models due to the lack of adequate experimental approaches. However, it is not clear yet how reliable these models are. To assess the accuracy of commonly used models, a reference 3D numerical model for cardiomyocytes (NMReal) was developed, consisting of the cell membrane shell reconstructed from rendered confocal microscopy images of freshly isolated ventricular myocytes. NMReal was used to estimate the E-induced maximum ΔVm values (ΔVmax), which were compared with estimates from seven other electromagnetic models. Accurate ΔVmax estimates (average error < 2%) were obtained with a less complex 3D model (NM3D) based on the extruded 2D image of the cell longitudinal section. Acceptable ΔVmax estimates (average error < 5%) were obtained with the prolate spheroid analytical model (PSAM) when the angle of E incidence and the cell major axis was < 30°. In this case, PSAM, a much simpler model requiring only the measurement of the longitudinal and transversal cell dimensions, can be a suitable alternative for ΔVmax calculation. Graphical abstract (A) Confocal images of the cell were used to reconstruct the realistic geometry of cardiomyocytes (NMReal). (B) NMReal was used to estimate the maximum variation in the transmembrane potential (ΔVmax) induced by an external electric field (E) applied at different angles with respect to the cell major axis. Plus (anode) and minus (cathode) signs indicate electrode position (E direction is from minus to plus). (C) Relative error (vs. NMReal) of ΔVmax estimation with simplified electromagnetic models, presented in descending order of accuracy (left-to-right, top-to-bottom). NM2D: 2D numerical model based on the longitudinal cell image; NM3D: numerical model based on the z extrusion of NM2D; EAM, PSAM, and CAM: ellipsoidal, prolate spheroidal, and cylindrical analytical models, respectively; PNM and CNM: parallelepipedal and cylindrical numerical models, respectively.

Sources of Ca2+ for contraction of the heart tube of Tenebrio molitor (Coleoptera: Tenebrionidae).

Insect and vertebrate hearts share the ability to generate spontaneously their rhythmic electrical activity, which triggers the fluid-propelling mechanical activity. Although insects have been used as models in studies on the impact of genetic alterations on cardiac function, there is surprisingly little information on the generation of the inotropic activity in their hearts. The main goal of this study was to investigate the sources of Ca2+ for contraction in Tenebrio molitor hearts perfused in situ, in which inotropic activity was assessed by the systolic variation of the cardiac luminal diameter. Increasing the pacing rate from 1.0 to 2.5 Hz depressed contraction amplitude and accelerated relaxation. To avoid inotropic interference of variations in spontaneous rate, which have been shown to occur in insect heart during maneuvers that affect Ca2+ cycling, experiments were performed under electrical pacing at near-physiological rates. Raising the extracellular Ca2+ concentration from 0.5 to 8 mM increased contraction amplitude in a manner sensitive to L-type Ca2+ channel blockade by D600. Inotropic depression was observed after treatment with caffeine or thapsigargin, which impair Ca2+ accumulation by the sarcoplasmic reticulum (SR). D600, but not inhibition of the sarcolemmal Na+/Ca2+ exchanger by KB-R7943, further depressed inotropic activity in thapsigargin-treated hearts. From these results, it is possible to conclude that in T. molitor heart, as in vertebrates: (a) inotropic and lusitropic activities are modulated by the heart rate; and (b) Ca2+ availability for contraction depends on both Ca2+ influx via L-type channels and Ca2+ release from the SR.

Minimally invasive measurement of vesical pressure for diagnosis of infravesical obstruction.

AIMS: This study was focused on the clinical test of an improved and portable version of a previously described urethral connector (UC), designed for minimally invasive measurement of vesical pressure and diagnosis of infravesical obstruction in men. METHODS: The conventional pressure-flow study (PFS) and the test with the new version of UC were applied to individuals reporting lower urinary tract symptoms (LUTS), who were then classified as obstructed or non-obstructed/equivocal based on the bladder outlet obstruction index (BOOI) from PFS data. Two-way analysis of variance was used to compare the values of urine flow rate and vesical pressure between methods and diagnoses. RESULTS: Vesical pressure and urine flow values were not significantly different between methods (P > 0.05), while the former was greater in the group classified as obstructed. CONCLUSION: The present results showed that the UC test can support the diagnosis of infravesical obstruction in a comparable way as that of the conventional urodynamic method, however with the advantages of simplicity and minimal invasiveness, having thus the potential to be an alternative method for long term follow-up of individuals reporting LUTS.

Atrial chronotropic reactivity to catecholamines in neonatal rats: Contribution of ß-adrenoceptor subtypes.

Although increase in heart rate is a crucial determinant for enhancement of cardiac output in the neonate, information on the chronotropic reactivity to catecholamines during postnatal development is scarce. The present study was aimed at investigating the role of ß-adrenoceptor subtypes and catecholamine removal mechanisms in the adrenergic chronotropic response during the early post-natal period. Right atria isolated from immature (0-21 day old) and adult (4-6 month old) rats were used for determination of the responsiveness to agonists and quantitation of the transcripts of proteins involved in ß-adrenergic signaling. The main results were: (a) the maximum response (Rmax) to norepinephrine increased with age, whereas sensitivity decreased; (b) age-dependent differences in sensitivity to norepinephrine were abolished by inhibition of the neuronal norepinephrine transporter; (c) Rmax to isoproterenol was similar in immature and adult atria, and depressed only in the former by ß2-adrenoceptor blockade with ICI118,551; (d) neonatal atria showed greater ß2-adrenoceptor mRNA levels, and more prominent positive chronotropic response to the ß2- and ß3-adrenoceptor agonists zinterol and YM178, respectively (nanomolar range); (e) in atria of immature rats, transcript levels of the extraneuronal monoamine transporter were lower, and its inhibition did not affect sensitivity to isoproterenol; and (f) reactivity to forskolin and 3-isobutyl-1-methylxanthine was not affected by age. The increased ß2- and ß3-adrenoceptor participation in the adrenergic chronotropic response, in addition to weaker catecholamine removal, may compensate for the immature cardiac innervation and the apparently reduced efficiency of ß1-adrenoceptor signaling in the neonate, increasing the responsiveness to endogenous and exogenous ß2-adrenoceptor agonists.

Electrocardiogram, heart movement and heart rate in the awake gecko (Hemidactylus mabouia).

The electrocardiogram (ECG) is the simplest and most effective non-invasive method to assess the electrical activity of the heart and to obtain information on the heart rate (HR) and rhythm. Because information on the HR of very small reptiles (body mass <10 g) is still scarce in the literature, in the present work we describe a procedure for recording the ECG in non-anesthetized geckos (Hemidactylus mabouia, Moreau de Jonnès, 1818) under different conditions, namely manual restraint (MR), spontaneous tonic immobility (TI), and in the non-restrained condition (NR). In the gecko ECG, the P, QRS and T waves were clearly distinguishable. The HR was 2.83 ± 0.02 Hz under MR, which was significantly greater (p < 0.001) than the HR under the TI (1.65 ± 0.09 Hz) and NR (1.60 ± 0.10 Hz) conditions. Spontaneously beating isolated gecko hearts contracted at 0.84 ± 0.03 Hz. The in vitro beating rate was affected in a concentration-dependent fashion by adrenoceptor stimulation with noradrenaline, as well as by the muscarinic cholinergic agonist carbachol, which produced significant positive and negative chronotropic effects, respectively (p < 0.001). To our knowledge, this is the first report on the ECG morphology and HR values in geckos, particularly under TI. The methodology and instrumentation developed here are useful for non-invasive in vivo physiological and pharmacological studies in small reptiles without the need of physical restraint or anesthesia.

Muscarinic stimulation and pinacidil produce similar facilitation of tachyarrhythmia induction in rat isolated atria.

Atrial tachyarrhythmias, the most common type of cardiac arrhythmias, are associated with greater stroke risk. Muscarinic cholinergic agonists have been shown to facilitate atrial tachyarrhythmia maintenance in the absence of cardiac disease. This has been attributed to action potential shortening, which enhances myocardial electrical anisotropy, and thus creates a substrate for reentrant excitation. In this study, we describe a similar effect of the ATP-sensitive K(+) channel (KATP) opener pinacidil on tachyarrhythmia induction in isolated rat atria. Pinacidil, which activates a weakly inwardly-rectifying current in isolated atrial myocytes, enhanced arrhythmia induction in the right and left atria. This effect was abolished by the KATP blocker glibenclamide, but not by atropine, which rules out a possible indirect effect due to stimulation of acetylcholine release. However, pinacidil attenuated carbachol-induced tachyarrhythmia facilitation, which may indicate that the action of these agonists converges to a common cellular mechanism. Both agonists caused marked action potential shortening in isolated atrial myocytes. Moreover, during arrhythmia in the presence of pinacidil and carbachol, the atrial vectorelectrographic patterns were similar and consistent with reentrant propagation of the electrical activity. From these results, we conclude that the KATP channel opening is pro-arrhythmic in atrial tissue, which may pose as an additional risk in the scenario of myocardial hypoxia. Moreover, the similarity of the electrophysiological effects of pinacidil and carbachol is suggestive that the sole increase in background K(+) conductance is sufficient for atrial tachyarrhythmia facilitation.

Greater cardiac cell excitation efficiency with rapidly switching multidirectional electrical stimulation.

Electric field (E) stimulation is widely used in experiments with myocardial preparations and in the clinical setting (e.g., defibrillation). As a rule, stimuli are applied in a single direction, which limits excitatory cell recruitment because myocytes are disposed in different directions and their sensitivity to E depends on the stimulus orientation with respect to the cell major axis. Here, we propose a stimulatory approach, namely rapidly switching multidirectional stimulation (RSMS), in which stimuli are delivered in three directions within the electric refractory period. In populations of randomly oriented isolated rat cardiomyocytes, RSMS doubled the percentage of cells excited by near-threshold E (P < 0.001), which was more than the increase in recruitment in a single direction achieved by doubling E intensity. This effect was similar for monophasic and biphasic pulses, but for the latter, a given percent recruitment was obtained with 20-30% lower E intensity ( P < 0.01), so that RSMS with biphasic pulses allowed at least 60% reduction of E intensity for recruitment of >70% of the cells. RSMS can be applied to improve stimulation efficiency in experiments with isolated cardiac myocytes, and may be a promising alternative for decreasing shock intensity requirements for cardioversion and defibrillation.

Estimation of the fractional sarcoplasmic reticulum Ca2+ release in intact cardiomyocytes using integrated Ca2+ fluxes.

The sarcoplasmic reticulum (SR) is the main source of contraction-activating Ca2+ in the adult mammalian myocardium. The fraction of the SR Ca2+ content released at a twitch (fractional SR Ca2+ release, FR) is an important parameter for assessing the efficiency of excitation-contraction coupling under physiological and pathophysiological conditions, as well as for identification of modulators of this process. We here describe and propose an approach for FR quantitation based on the estimation of integrated Ca2+ fluxes mediated by different transporters that remove the ion from the cytosol. These fluxes may be calculated solely from the measurement of cytosolic free Ca2+ concentration ([Ca2+]i) during Ca2+ transients evoked under selective inhibition of the transporters, and from the cell Ca2+ buffering parameters available in the literature. The FR values obtained with this approach in intact rat ventricular myocytes (0.63 ± 0.04; n = 12) were comparable to those estimated in the same cell type with an already established method, based on electrophysiological measurements with the patch-clamp technique, in addition to [Ca2+]i measurement (0.69 ± 0.05; n = 6; p > 0.40). We conclude that the proposed method might be a suitable and technically simpler alternative to the electrophysiological method for FR estimation.

Medical equipment classification: method and decision-making support based on paraconsistent annotated logic.

As technology evolves, the role of medical equipment in the healthcare system, as well as technology management, becomes more important. Although the existence of large databases containing management information is currently common, extracting useful information from them is still difficult. A useful tool for identification of frequently failing equipment, which increases maintenance cost and downtime, would be the classification according to the corrective maintenance data. Nevertheless, establishment of classes may create inconsistencies, since an item may be close to two classes by the same extent. Paraconsistent logic might help solve this problem, as it allows the existence of inconsistent (contradictory) information without trivialization. In this paper, a methodology for medical equipment classification based on the ABC analysis of corrective maintenance data is presented, and complemented with a paraconsistent annotated logic analysis, which may enable the decision maker to take into consideration alerts created by the identification of inconsistencies and indeterminacies in the classification.

Pacemaker activity in the insect (T. molitor) heart: role of the sarcoplasmic reticulum.

The electrophysiological properties of the myogenic cardiac cells of insects have been analyzed, but the mechanisms that regulate the pacemaker activity have not been elucidated yet. In mammalian pacemaker cells, different types of membrane ion channels seem to be sequentially activated, perhaps in a cooperative fashion with the current generated by Ca(2+) extrusion mediated by the electrogenic Na(+)/Ca(2+) exchanger, which is sustained by the diastolic sarcoplasmic reticulum (SR) Ca(2+) release. The objective of the present work was to investigate the role of the SR function on the basal beating rate (BR), and BR modulation by extracellular Ca(2+) concentration ([Ca(2+)](o)) and neurotransmitters in the in situ dorsal vessel (heart) of the mealworm beetle Tenebrio molitor. The main observations were as follows: 1) basal BR was reduced by 50% by inhibition of SR function, but not affected by perfusion with CsCl or ZD7288; 2) spontaneous activity was abolished by Cd(2+); 3) a robust positive chronotropic response could be elicited to serotonin (5-HT), but not to norepinephrine or carbamylcholine; 4) SR inhibition abolished the sustained chronotropic stimulation by [Ca(2+)](o) elevation and by 5-HT, while the latter was unaffected by CsCl. It is concluded that, in T. molitor heart, BR is markedly, but not exclusively, dependent on the SR function, and that BR control and modulation by both [Ca(2+)](o) and 5-HT requires a functional SR.

Determination of the vectorelectrogram in isolated rat atria: application to the study of arrhythmias.

Atrial tachyarrhythmias, the most frequent type of cardiac arrhythmia, are associated with increased stroke risk. Reentry and focal activity are considered as the main mechanisms underlying this dysfunction. In this study, we describe determination of the vectorelectrogram in isolated rat atria as a means to distinguish different patterns of electrical propagation. In all studied right atria beating at sinus rhythm, the mean electric vector (MEV) trajectory was clockwise, and each cycle was preceded by electric diastole (null MEV), either in the absence or presence of muscarinic cholinergic or beta-adrenergic receptor stimulation. During cholinergic tachyarrhythmia (induced by high-rate electric stimulation in both atria, plus exposure to carbachol in left atria), vector loops were ellipsoidal and stable, with variable direction, and did not cross the origin, which is consistent with reentrant activation and with findings obtained in vivo by other authors. In contrast, during spontaneous activity induced by rapid pacing in isoproterenol-exposed left atria, vector loops were similar to those in right atria at sinus rhythm, thus suggestive of focal activity. It is concluded that the vectorelectrogram approach allows discrimination of different patterns of propagation during arrhythmia in isolated atria and may be useful for high-output tests of pro- and anti-arrhythmic compounds.

The role of extracellular potassium dynamics in the different stages of ictal bursting and spreading depression: a computational study.

Experimental evidences point out the participation of nonsynaptic mechanisms (e.g., fluctuations in extracellular ions) in epileptiform bursting and spreading depression (SD). During these abnormal oscillatory patterns, it is observed an increase of extracellular potassium concentration [K(+)](o) and a decrease of extracellular calcium concentration [Ca(2+)](o) which raises the neuronal excitability. However, whether the high [K(+)](o) triggers and propagates these abnormal neuronal activities or plays a secondary role into this process is unclear. To better understand the influence of extracellular potassium dynamics in these oscillatory patterns, the experimental conditions of high [K(+)](o) and zero [Ca(2+)](o) were replicated in an extended Golomb model where we added important regulatory mechanisms of ion concentration as Na(+)-K(+) pump, ion diffusion and glial buffering. Within these conditions, simulations of the cell model exhibit seizure-like discharges (ictal bursting). The SD was elicited by the interruption of the Na(+)-K(+) pump activity, mimicking the effect of cellular hypoxia (an experimental protocol to elicit SD, the hypoxia-induced SD). We used the bifurcation theory and the fast-slow method to analyze the interference of K(+) dynamics in the cellular excitability. This analysis indicates that the system loses its stability at a high [K(+)](o), transiting to an elevated state of neuronal excitability. Effects of high [K(+)](o) are observed in different stages of ictal bursting and SD. In the initial stage, the increase of [K(+)](o) creates favorable conditions to trigger both oscillatory patterns. During the neuronal activity, a continuous growth of [K(+)](o) by outward K(+) flow depresses K(+) currents in a positive feedback way. At the last stage, due to the depression of K(+) currents, the Na(+)-K(+) pump is the main mechanism in the end of neuronal activity. Thus, this work suggests that [K(+)](o) dynamics may play a fundamental role in these abnormal oscillatory patterns.

A novel intraurethral device diagnostic index to classify bladder outlet obstruction in men with lower urinary tract symptoms.

Objectives. Using a urethral device at the fossa navicularis, bladder pressure during voiding can be estimated by a minimal invasive technique. This study purposes a new diagnostic index for patients with lower urinary tract symptoms (LUTSs). Methods. Fifty one patients presenting with LUTSs were submitted to a conventional urodynamic and a minimal invasive study. The results obtained through the urethral device and invasive classic urodynamics were compared. The existing bladder outlet obstruction index (BOOI) equation that classifies men with LUTSs was modified to allow minimal invasive measurement of isovolumetric bladder pressure in place of detrusor pressure at maximum urine flow. Accuracy of the new equation for classifying obstruction was then tested in this group of men. Results. The modified equation identified men with obstruction with a positive predictive value of 68% and a negative predictive value of 70%, with an overall accuracy of 70%. Conclusions. The proposed equation can accurately classify over 70% of men without resorting to invasive pressure flow studies. We must now evaluate the usefulness of this classification for the surgical treatment of men with LUTSs.

Osmolality- and Na+ -dependent effects of hyperosmotic NaCl solution on contractile activity and Ca2+ cycling in rat ventricular myocytes.

Hypertonic NaCl solutions have been used for small-volume resuscitation from hypovolemic shock. We sought to identify osmolality- and Na(+)-dependent components of the effects of the hyperosmotic NaCl solution (85 mOsm/kg increment) on contraction and cytosolic Ca(2+) concentration ([Ca(2+)](i)) in isolated rat ventricular myocytes. The biphasic change in contraction and Ca(2+) transient amplitude (decrease followed by recovery) was accompanied by qualitatively similar changes in sarcoplasmic reticulum (SR) Ca(2+) content and fractional release and was mimicked by isosmotic, equimolar increase in extracellular [Na(+)] ([Na(+)](o)). Raising osmolality with sucrose, however, augmented systolic [Ca(2+)](i) monotonically without change in SR parameters and markedly decreased contraction amplitude and diastolic cell length. Functional SR inhibition with thapsigargin abolished hyperosmolality effects on [Ca(2+)](i). After 15-min perfusion, both hyperosmotic solutions slowed mechanical relaxation during twitches and [Ca(2+)](i) decline during caffeine-evoked transients, raised diastolic and systolic [Ca(2+)](i), and depressed systolic contractile activity. These effects were greater with sucrose solution, and were not observed after isosmotic [Na(+)](o) increase. We conclude that under the present experimental conditions, transmembrane Na(+) redistribution apparently plays an important role in determining changes in SR Ca(2+) mobilization, which markedly affect contractile response to hyperosmotic NaCl solutions and attenuate the osmotically induced depression of contractile activity.

Combining stimulus direction and waveform for optimization of threshold stimulation of isolated ventricular myocytes.

Electric field stimulation is widely used for heart pacing and arrhythmia reversion. In this study, we analysed the influence of waveform and direction of external stimulating electric field on the excitation threshold of isolated ventricular myocytes. The threshold field (E(T)) was lower when the field was applied longitudinally (E(T,L)) rather than transversally (E(T,T)) to the cell major axis. Rheobase was greater for transversal stimulation, but chronaxie and estimated membrane polarization were similar for both directions. The calculated maximal variation in membrane potential at the threshold (DeltaV(T) approximately 15 mV) was insensitive to field direction. As DeltaV(T) values were similar, we assumed that the E(T,T)/E(T,L) ratio might be described solely as the ratio of the major and minor cell semi-axes. Accordingly, the ratio thus estimated was comparable to that determined experimentally. Stimulus waveform significantly affected both E(T) and DeltaV(T), which were greater for monophasic versus biphasic stimuli. Direction and waveform effects were independent. We conclude that (a) direction affects E(T) by its influence on the ability of a given field intensity to cause threshold membrane polarization and (b) threshold-lowering effects of longitudinal stimulation and biphasic waveforms apparently depend on different mechanisms, are additive and thus may be combined to decrease the energy requirement for myocardial stimulation.

Enhanced calcium mobilization in rat ventricular myocytes during the onset of pressure overload-induced hypertrophy.

Early cardiovascular changes evoked by pressure overload (PO) may reveal adaptive strategies that allow immediate survival to the increased hemodynamic load. In this study, systolic and diastolic Ca(2+) cycling was analyzed in left ventricular rat myocytes before (day 2, PO-2d group) and after (day 7, PO-7d group) development of hypertrophy subsequent to aortic constriction, as well as in myocytes from time-matched sham-operated rats (sham group). Ca(2+) transient amplitude was significantly augmented in the PO-2d group. In the PO-7d group, intracellular Ca(2+) concentration ([Ca(2+)](i)) was reduced during diastole, and mechanical twitch relaxation (but not [Ca(2+)](i) decline) was slowed. In PO groups, fractional sarcoplasmic reticulum (SR) Ca(2+) release at a twitch, SR Ca(2+) content, SR Ca(2+) loss during diastole, and SR-dependent integrated Ca(2+) flux during twitch relaxation were significantly greater than in sham-operated groups, whereas the relaxation-associated Ca(2+) flux carried by the Na(+)/Ca(2+) exchanger was not significantly changed. In the PO-7d group, mRNA levels of cardiac isoforms of SR Ca(2+)-ATPase (SERCA2a), phospholamban, calsequestrin, ryanodine receptor, and NCX were not significantly altered, but the SERCA2a-to-phospholamban ratio was increased 2.5-fold. Moreover, greater sensitivity to the inotropic effects of the beta-adrenoceptor agonist isoproterenol was observed in the PO-7d group. The results indicate enhanced Ca(2+) cycling between SR and cytosol early after PO imposition, even before hypertrophy development. Increase in SR Ca(2+) uptake may contribute to enhancement of excitation-contraction coupling (augmented SR Ca(2+) content and release) and protection against arrhythmogenesis due to buildup of [Ca(2+)](i) during diastole.