Impact of RyR2 potentiation on myocardial function.

This perspective attempts to shed light on an old and not yet solved controversy in cardiac physiology, i.e., the impact of increasing ryanodine receptor (RyR)2 open probability on myocardial function. Based on an already proven myocyte model, it was shown that increasing RyR2 open probability results in a purely short-lived increase in Ca2+ transient amplitude, and, therefore, it does not increase cardiac contractility. However, potentiation of RyR2 activity permanently enhances fractional Ca2+ release, shifting the intracellular Ca2+ transient versus sarcoplasmic reticulum (SR) Ca2+ content curve to a new state of higher efficiency. This would allow the heart to maintain a given contractility despite a decrease in SR Ca2+ content, to enhance contractility if SR Ca2+ content is simultaneously preserved or to successfully counteract the effects of a negative inotropic intervention.NEW & NOTEWORTHY Increasing ryanodine receptor (RyR)2 open probability does not increase cardiac contractility. However, RyR2 potentiation shifts the intracellular Ca2+ transient-sarcoplasmic reticulum (SR) Ca2+ content relationship toward an enhanced efficiency state, which may contribute to a positive inotropic effect, preserve contractility despite decreased SR Ca2+ content, or successfully counteract the effects of a negative inotropic action.

Multiscale model of the human cardiovascular system: Description of heart failure and comparison of contractility indices.

A multiscale model of the cardiovascular system is presented. Hemodynamics is described by a lumped parameter model, while heart contraction is described at the cellular scale. An electrophysiological model and a mechanical model were coupled and adjusted so that the pressure and volume of both ventricles are linked to the force and length of a half-sarcomere. Particular attention was paid to the extreme values of the sarcomere length, which must keep physiological values. This model is able to reproduce healthy behavior, preload variations experiments, and ventricular failure. It also allows to compare the relevance of standard cardiac contractility indices. This study shows that the theoretical gold standard for assessing cardiac contractility, namely the end-systolic elastance, is actually load-dependent and therefore not a reliable index of cardiac contractility.

Phospholamban ablation rescues the enhanced propensity to arrhythmias of mice with CaMKII-constitutive phosphorylation of RyR2 at site S2814.

KEY POINTS: Mice with Ca(2+) -calmodulin-dependent protein kinase (CaMKII) constitutive pseudo-phosphorylation of the ryanodine receptor RyR2 at Ser2814 (S2814D(+/+) mice) exhibit a higher open probability of RyR2, higher sarcoplasmic reticulum (SR) Ca(2+) leak in diastole and increased propensity to arrhythmias under stress conditions. We generated phospholamban (PLN)-deficient S2814D(+/+) knock-in mice by crossing two colonies, S2814D(+/+) and PLNKO mice, to test the hypothesis that PLN ablation can prevent the propensity to arrhythmias of S2814D(+/+) mice. PLN ablation partially rescues the altered intracellular Ca(2+) dynamics of S2814D(+/+) hearts and myocytes, but enhances SR Ca(2+) sparks and leak on confocal microscopy. PLN ablation diminishes ventricular arrhythmias promoted by CaMKII phosphorylation of S2814 on RyR2. PLN ablation aborts the arrhythmogenic SR Ca(2+) waves of S2814D(+/+) and transforms them into non-propagating events. A mathematical human myocyte model replicates these results and predicts the increase in SR Ca(2+) uptake required to prevent the arrhythmias induced by a CaMKII-dependent leaky RyR2. ABSTRACT: Mice with constitutive pseudo-phosphorylation at Ser2814-RyR2 (S2814D(+/+) ) have increased propensity to arrhythmias under ß-adrenergic stress conditions. Although abnormal Ca(2+) release from the sarcoplasmic reticulum (SR) has been linked to arrhythmogenesis, the role played by SR Ca(2+) uptake remains controversial. We tested the hypothesis that an increase in SR Ca(2+) uptake is able to rescue the increased arrhythmia propensity of S2814D(+/+) mice. We generated phospholamban (PLN)-deficient/S2814D(+/+) knock-in mice by crossing two colonies, S2814D(+/+) and PLNKO mice (SD(+/+) /KO). SD(+/+) /KO myocytes exhibited both increased SR Ca(2+) uptake seen in PLN knock-out (PLNKO) myocytes and diminished SR Ca(2+) load (relative to PLNKO), a characteristic of S2814D(+/+) myocytes. Ventricular arrhythmias evoked by catecholaminergic challenge (caffeine/adrenaline) in S2814D(+/+) mice in vivo or programmed electric stimulation and high extracellular Ca(2+) in S2814D(+) /(-) hearts ex vivo were significantly diminished by PLN ablation. At the myocyte level, PLN ablation converted the arrhythmogenic Ca(2+) waves evoked by high extracellular Ca(2+) provocation in S2814D(+/+) mice into non-propagated Ca(2+) mini-waves on confocal microscopy. Myocyte Ca(2+) waves, typical of S2814D(+/+) mice, could be evoked in SD(+/+) /KO cells by partially inhibiting SERCA2a. A mathematical human myocyte model replicated these results and allowed for predicting the increase in SR Ca(2+) uptake required to prevent the arrhythmias induced by a Ca(2+) -calmodulin-dependent protein kinase (CaMKII)-dependent leaky RyR2. Our results demonstrate that increasing SR Ca(2+) uptake by PLN ablation can prevent the arrhythmic events triggered by SR Ca(2+) leak due to CaMKII-dependent phosphorylation of the RyR2-S2814 site and underscore the benefits of increasing SERCA2a activity on SR Ca(2+) -triggered arrhythmias.

Effect of preservation solution on graft viability in single-lung transplantation from heart-beating donors in pigs.

BACKGROUND: Low-potassium-dextran preservation solution Perfadex (PER) may provide better outcome of transplanted lungs than high-potassium Euro-Collins (EC) solution. However, there are no comparative studies of the recipient inflammatory response to the graft. PURPOSE: The purpose of this study was to compare EC versus PER as preservation solutions with respect to the functional performance and inflammatory response in single-lung transplantation from heart-beating donors in pigs. MATERIALS AND METHODS: The donor left lung flushed with the corresponding cold preservation solution was stored at 3 degrees C for 3 hours. We assessed hemodynamic values and pulmonary function in the recipient over a 2-hour reperfusion period calculated as percent of basal values, and expressed as mean of the reperfusion period. Interleukin-8 (IL-8) concentration in the donor was estimated in bronchoalveolar lavage fluid 2 hours after recipient reperfusion. Biopsies of the donor right lung and the transplanted lung were obtained to measure myeloperoxidase (MPO) activity. IL-8 and MPO values were expressed as percent of the donor value. We evaluated the wet/dry pulmonary weight ratio (W/D), polymorphonuclear neutrophil count (PMN), and a score of histological damage in the transplanted graft. RESULTS: Pulmonary function evaluated by % static: 66.6 +/- 6.8 (EC), 82.3 +/- 10.2 (PER), and dynamic: 74.0 +/- 7.3 (EC), 89.3 +/- 7.7 (PER) compliances, as well as % IL-8: 562.5 +/- 168.6 (EC), 232.3 +/- 148.7 (PER), % MPO: 485.9 +/- 194.9 (EC), 140.8 +/- 21.1 (PER), W/D: 9.9 +/- 3.1 (EC), 6.8 +/- 1.4 (PER), PMN 13.5 +/- 6.8 (EC), 5.5 +/- 3.3 (PER) and the histological damage score: 3.0 +/- 1.5 (EC), 0.7 +/- 0.4 (PER) showed significant differences between the EC and the PER (P < .01). CONCLUSIONS: PER affords good lung preservation with early graft function and modest evidences of inflammation, lung injury, and edema compared with the EC perfused lung.

Glibenclamide action on myocardial function and arrhythmia incidence in the healthy and diabetic heart.

Myocardial sarcolemmal ATP-dependent potassium (KATP) channels, which are normally closed by high ATP concentration, open during ischemia when ATP generation decreases favoring K(+) efflux. This reduces action potential duration (APD) decreasing the time of Ca(2+) influx and Ca(2+) overload. This behavior suggested that they might be involved in the protection against stunning and arrhythmias and in the mechanism of ischemic preconditioning. Sulfonylureas, used as hypoglycemic agents for the treatment of type 2 diabetes also block myocardial KATP channels prolonging APD during ischemia, which by allowing Ca(2+) entry for a longer period of time, is potentially harmful to the heart. Controversial findings have been reported regarding the protective effect of sulfonylureas. Due to their importance in the clinical setting, their action on the heart of large conscious animal models is relevant. The effect of glibenclamide, a representative sulfonylurea, has been studied in a conscious sheep model submitted to regional 12 min ischemia. Glibenclamide (0.4 mg/kg) completely blocked KATP channels, as assessed by monophasic APD, producing a deleterious effect on reperfusion-induced arrhythmias and myocardial recovery from stunning in normal animals. This adverse effect was more noticeable in alloxan-induced diabetic sheep, where a lower dose (0.1 mg/kg) inhibited KATP channel opening worsening mechanical recovery and arrhythmia incidence. However, glibenclamide did not abolish ischemic late preconditioning against stunning and arrhythmias in normal animals. Because diabetic sheep do not develop this cardioprotective phenomenon, probably due to KATP channel dysfunction, it was not possible to assess glibenclamide effect on preconditioning in this pathological condition. In conclusion, in large conscious animals, glibenclamide interferes with the beneficial action of KATP channel opening during acute ischemia-reperfusion events both in normal and diabetic animals. Therefore, despite some studies claiming no added cardiovascular risk due to glibenclamide treatment, this pharmacological agent should be further investigated to ensure its safe administration in patients with concurrent heart disease.

Entrance in mitosis of adult cardiomyocytes in ischemic pig hearts after plasmid-mediated rhVEGF165 gene transfer.

Replacement of the cell loss occurring after acute myocardial infarction has been proposed as a potential treatment to prevent heart remodeling and failure. On account that cardiomyocytes express VEGF receptors and that VEGF triggers mitogen-activated protein kinases, we investigated if VEGF gene transfer may induce cardiomyocyte replication. In a pig model of chronic myocardial ischemia achieved by Ameroid occlusion of the left circumflex coronary artery, we observed that direct intramyocardial injection of a plasmid encoding human VEGF(165) induced a several-fold increase in cardiomyocyte mitotic index and in the number of cardiomyocyte nuclei per unit volume as compared with pigs receiving plasmid devoid of gene. Despite images of conventional cytokinesis were not observed, the fact that caryokinesis is an obligatory step for cell division suggests that our finding may contribute to the issue of heart regeneration and may potentially widen the therapeutic spectrum of VEGF gene transfer.

Glibenclamide effects on reperfusion-induced malignant arrhythmias and left ventricular mechanical recovery from stunning in conscious sheep.

INTRODUCTION: Sulfonylureas have been associated with a high incidence of cardiovascular death in diabetic patients treated with these drugs. Although the evidence on the cardiovascular effects of sulfonylureas is contradictory and scarce, many experiments have shown that the second-generation compound glibenclamide has a protective effect on mechanical function and against generation of malignant arrhythmias. OBJECTIVE: The purpose of this study was to assess whether glibenclamide elicits protection on postischemic myocardial functional recovery (stunning) and against reperfusion-induced arrhythmias in a conscious sheep model. METHODS: Sheep were divided into three groups: control, glibenclamide (0.4 mg/kg) and vehicle. After a 12-min ischemic period, the heart was reperfused and recordings for index calculation were acquired during 2 h of reperfusion. Percent systolic wall thickening fraction (%WTH), radial diastolic compliance (CR), arrhythmia incidence and Bernauer's arrhythmia severity index (ASI) were calculated for each group. RESULTS: Glibenclamide infusion had a high proarrhythmic action (ASI: glibenclamide 143, control 54 and vehicle 23; ANOVA P < 0.001 drug vs. control and vehicle) and a detrimental effect on regional systolic (%WTH: glibenclamide 26.9 +/- 6.7, control 65.7 +/- 3.5 and vehicle 68.6 +/- 5.6, ANOVA P < 0.01 drug vs. control and vehicle) and diastolic function (CR: glibenclamide 76.2 +/- 7.8, control 104.7 +/- 4.2 and vehicle 106 +/- 4.9, ANOVA P < 0.05 drug vs. control and vehicle) during reperfusion. CONCLUSIONS: Glibenclamide infusion resulted in adverse cardiovascular effects. The combined deleterious effects on reperfusion-induced arrhythmias and on myocardial recovery from stunning could be the cause of the unexplained high mortality in diabetic patients treated with sulfonylurea derivatives. The mechanism involved seems to be the blockade of the cardiac ATP sensitive potassium (K-ATP) channel.

Concentration and elongation of attached cross-bridges as pressure determinants in a ventricular model.

A ventricular model based on a muscle model relating sarcomere dynamics to Ca(2+)kinetics was used to establish the relative contribution to pressure development of the two components of cross-bridge dynamics: attached cross-bridge concentration and elongation of its elastic structure. The model was tested by reproduction of experiments reflecting myofibrillar behavior at the ventricular level as well as chamber mechanical properties. It was then used to study cross-bridge behavior independently of Ca(2+)activation, by simulation of flow-clamp experiments at constant Ca(2+)concentration. During the volume ramp, both reduced cross-bridge elongation and decreased concentration by cross-bridge detachment caused a fall of pressure; at end-ejection there was a fast partial increase of pressure by recovery of cross-bridge elongation, and during post-ejection there was a slower pressure change towards the value corresponding to end-ejection volume by cross-bridge reattachment according to the rate of constants of Ca(2+)kinetics. Likewise, during a physiological normal ejection, results showed that a maximal decrease in cross-bridge elongation (Deltah) produced a maximal reduction of ejecting pressure with respect to that at constant cross-bridge elongation (DeltaP), both in simulated beats (DeltaP=20%, Deltah=17%), and in experimentally fitted pressure-volume data from open-chest dogs (DeltaP=43.7+/-3.8%, Deltah=30.7+/-8.3%), Deltah being dependent of peak flow (Deltah=0. 1471 peak flow+6.0788, r=0.72). We conclude that normal ejecting pressure depends not only on cross-bridge concentration, but also on the elongation of its elastic structure, which reduces pressure according to flow.

Left ventricular regional systolic and diastolic function in conscious sheep undergoing ischemic preconditioning.

OBJECTIVE: Late preconditioning diastolic protection and cardiac function optimization by the combined effects of late and early preconditioning have not been studied in conscious animals. This study assessed in fully conscious sheep: (1) whether 24 h after a reversible ischemia, a new ischemic episode results in lesser systo-diastolic dysfunction (late preconditioning protection) and (2) whether the addition of early preconditioning (brief episodes of ischemia-reperfusion before the subsequent sustained ischemia) on the second day of late preconditioning optimized the second window of protection. METHODS: Three protocols were assessed: (a) late preconditioning, 9 min ischemia and 2 h reperfusion was done on two consecutive days; (b) early plus late preconditioning, as in protocol (a) except that on day 2 the heart underwent three periods of 3 min ischemia-6 min reperfusion prior to the sustained 9 min ischemia; (c) early preconditioning, the same protocol as in (b) except that day 2 was separated 1 week from day 1. RESULTS: Late preconditioning decreased regional radial diastolic stiffness from 147 +/- 26% (day 1) to 96 +/- 14% (day 2), at 2 h of reperfusion (mean +/- SEM, p < 0.05), but did not protect against systolic stunning (thickening fraction and regional stroke work). Early plus late preconditioning did not improve late preconditioning findings. Early preconditioning alone did not protect either systolic or diastolic functions. CONCLUSION: In conscious sheep, there is diastolic but not systolic mechanical protection with late preconditioning. Diastolic protection is not enhanced by the addition of early preconditioning.

A cardiac muscle model relating sarcomere dynamics to calcium kinetics.

A muscle model establishing the link between cross-bridge dynamics and intracellular Ca2+ kinetics was assessed by simulation of experiments performed in isolated cardiac muscle. The model is composed by the series arrangement of muscle units formed by inextensible thick and thin filaments in parallel with an elastic element. Attached cross-bridges act as independent force generators whose force is linearly related to the elongation of their elastic structure. Ca2+ kinetics is described by a four-state system of sites on the thin filament associated with troponin C: sites with free troponin C (T), sites with Ca2+ bound to troponin C (TCa); sites with Ca2+ bound to troponin C and attached cross-bridges (TCa*); and sites with troponin C not associated with Ca2+ and attached cross-bridges (T*). The intracellular Ca2+ concentration ([Ca2+]) is controlled solely by the sarcoplasmic reticulum through an inflow function and a saturated outflow pump function. All the simulations were performed using the same set of parameters. The model was able to reproduce the following experiments in cardiac muscle: (a) time course of isometric force (peak force: 46.5 mN/mm2), intracellular [Ca2+] (peak [Ca2+]: 1.5 microM); (b) force-length-[Ca2+] relations; (c) transient response of force to step changes in length; (d) force-velocity relation (maximum velocity: 3 microns/s); (e) the force response to length pulses to estimate the time course of [TCa]; (f) force response to quick releases showing the superactivating and deactivating effects of shortening; (g) stiffness response to sinusoidal length changes; and (h) time course of active state. The good accordance of the simulations with experimental results indicates that the model is an adequate representation of the link between cross-bridge dynamic behaviour and Ca2+ kinetics.

The contractile mechanism as an approach to building left ventricular pump models.

OBJECTIVE: The aim was to construct a model linking a simplified interpretation of the contractile process at the myofilament level to the mechanical behaviour of the left ventricle to improve the ability of elastic-resistive models to represent the pumping response of the left ventricle. The mechanical model, consisting of an elastic component connected in series with a contractile component and an elastic component parallel to both the series elastic and contractile components, is able to develop pressure by the binding of a structural substance T to an excitatory substance C, the behaviour of which is a simplification of miofibrillar Ca2+ kinetics. METHODS: Theoretically, the model was validated for its ability to reproduce by computer simulation, experiments that described the pumping properties of the left ventricle--namely, elasticity, resistivity, deactivating and positive effect of ejection, and the behaviour of intracellular Ca2+. Experimentally, the model was tested to fit intraventricular pressure (P(t)) and volume (V(t)) of single ejective beats in nine open chest dogs fitted with a pressure microtransducer to measure intraventricular P(t) and an aortic flowprobe to measure ventricular outflow and calculate V(t). Parameters were estimated up to maximum negative dP/dt adjusting P(t) or V(t) data of the ejective beats, and the goodness of the fit was evaluated through the root mean square error normalised with respect to the corresponding mean P(t) or V(t) in the fitting interval (NE). RESULTS: Descriptive validation of the model showed that the mean NE for the ejective P(t) fit was 0.03(SD 0.005) and for the V(t) fit 0.014(0.003). Predictive validation of P(t) and V(t) data of beats with partial occlusion of the aorta was performed up to end ejection, with parameters estimated from the P(t) or V(t) fit of the preceding ejective beat. Results gave a mean NE equal to 0.05(0.02) for predicted P(t) and 0.02(0.007) for predicted V(t), from either source of estimated parameters. Explanative validation showed that all the estimated parameters were in the same range used in simulation and that derived indexes [isovolumic maximum pressure (Pmax) = 166(13) mm Hg, time to maximum pressure (TPmax) = 0.186(0.012) s and the slope of the end systolic pressure volume relation (Emax) = 5.45(1.5) mm Hg.ml-1] were within reported experimental values. Finally, the model responded to increased inotropic state [dobutamine (5-35 micrograms.kg-1.min-1)] causing the estimated Pmax and Emax to increase by 33% and 25%, respectively, and TPmax to decrease by 10%. CONCLUSION: This model represented an improvement over previous pump models because (1) the model was able to represent behaviours other than purely elastic-resistive ones, such as the deactivation and positive effect of ejection; (2) left ventricular properties were the response of model behaviour and not constitutive elements of its structure; and (3) it adequately fulfilled model validation procedures.

Differential effects of left anterior descending coronary occlusion on left and right ventricular anterior wall thickening in the conscious pig.

OBJECTIVE: In humans, the left anterior descending coronary artery supplies the left ventricular wall, anterior septum and the paraseptal part of the right ventricular anterior wall. Our aim was to study the effects of acute left anterior descending coronary occlusion on wall thickening in the regions of the left and right ventricular anterior walls supplied by the artery, and in remote, non-ischaemic regions of both ventricles. METHODS: Systolic wall thickening (defined as percent thickening with respect to end diastolic wall thickness) was studied in eight conscious pigs every 15 s during 1 min of acute left anterior descending coronary occlusion by a cuff occluder, and every 30 s during 4 min of reperfusion. Pigs were instrumented with ultrasonic microcrystals measuring wall thickness in the anterior walls (left anterior descending artery territory) and lateral walls (left circumflex or right coronary artery territory) of both ventricles, and a left ventricular pressure microtransducer. RESULTS: During control and reperfusion, both anterior walls displayed similar systolic thickening. During coronary occlusion, the left ventricular anterior wall showed paradoxical systolic thinning (dyskinesia) whereas the right ventricular anterior wall showed only hypokinesia. CONCLUSIONS: In the presence of equal blood flow deprivation, the right ventricular anterior wall supplied by the left anterior descending coronary artery displays a significantly lesser degree of functional impairment than the left ventricular anterior wall supplied by the same artery. This differential effect may be due to mechanical unloading of the right ventricular anterior wall resulting from left ventricular anterior wall ischaemia. This afterload reduction due to decreased mechanical interaction between the two walls would allow the right ventricular anterior wall to express its contractile reserve in the form of systolic thickening.

Immunoperoxidase tracing of Junin virus neural route after footpad inoculation.

To determine the pathway adopted by peripherally inoculated Junin virus (JV) to reach the CNS, rat tissues were serially harvested to trace the sequence of viral progression from right hind footpad to brain. Immunoperoxidase (PAP) labeling of viral antigen, concomitantly with infectivity assays and histological examination of each selected sample, were carried out. As from the 2nd week post-infection (pi), neurological disease inducing 100% mortality at 1 month was evident. At day 5 pi, viral antigen was first detected at footpad level in epidermic and dermic cells, as well as in neighbouring myocytes; labeled macrophages infiltrating small nerve branches were also disclosed. As from 10-15 days pi, viral antigen became apparent along ipsilateral sciatic nerve structures and within lumbar spinal ganglion neurons, followed by a fast viral spread throughout CNS neurons that involved spinal cord and brain. Concurrent histopathology featured minimal inflammatory reaction together with generalized astrocytic activation. Hematogenous viral transport was negligible, since JV was isolated much earlier and in higher infectivity titers in neural tissues than in blood. It may be concluded that after viral replication in footpad, JV neural route was demonstrated by its PAP labeling from peripheral nerves to cerebral cortex.

A triple staining procedure to evaluate phagocytic role of differentiated astrocytes.

To determine whether phagocytic activity is affected by a viral infection known to induce astrocyte differentiation, a triple procedure (PAP labeling for GFAP, PAS reaction for added baker's yeast cells and hematoxylin for nuclear staining of the whole monolayer) was applied to Junin virus-inoculated cultures, as well as matched controls. The three-step staining simplified yeast cell count for subsequent statistical analysis, which discerned preferential uptake by differentiated rather than immature astrocytes. Accordingly, greater cell maturation induced by Junin virus was concomitant with early enhancement of phagocytic activity.

Murine acinar pancreatitis preceding necrotizing myocarditis after Coxsackievirus B3 inoculation.

Balb/C weanling mice were inoculated intraperitoneally with a myocarditic variant of coxsackie-virus B3, with the aim of characterizing more fully the cell damage induced in the heart as well as in other organs. During the first week postinfection (pi), all animals developed acinar pancreatitis, followed by focal myocarditis. In accordance with the increasing infectivity titers, such progressive histopathological changes correlated with local viral replication. From day 4 pi, acinar degeneration accompanied by diffuse inflammatory exudate was observed in the pancreas, followed by fatty tissue replacement by day 8. In the heart, focal necrosis rather than inflammatory reaction first appeared at 4 days pi and became widespread by 6-8 days pi. Necrotic foci usually presented calcium deposits, with absence of myofibrils in the affected fibers. The fact that both periodic acid Schiff (PAS) and Best carmine staining remained positive even after diastase treatment ruled out basophilic necrosis. In summary, the pancreas appeared to be the site of primary viral replication leading to viremia.

Interventricular coronary steal induced by stenosis of left anterior descending coronary artery in exercising pigs.

BACKGROUND: In pigs and humans, the left anterior descending coronary artery (LAD) supplies the left ventricular anterior wall (LVAW), anterior septum, and paraseptal band of the right ventricular anterior wall (RVAW). The purposes of our study were 1) to study the LAD flow distribution in these walls during preexercise, exercise, and exercise with LAD stenosis and 2) to analyze regional wall motion under these conditions. METHODS AND RESULTS: Nine pigs were instrumented with sonomicrometers for measuring percent wall thickening (%WTh) in LVAW, RVAW, and lateral (control) walls of both ventricles, a hydraulic occluder at the LAD origin, an LV pressure transducer, and catheters for radioactive microsphere injection (left atrium) and blood withdrawal (aorta). One month later, regional %WTh and flows were measured during preexercise, exercise, and continuing exercise with LAD stenosis resulting in more than 50% reduction in systolic LVAW %WTh with regard to exercise. LAD stenosis caused a dramatic decrease in total mean +/- SD LVAW subendocardial flow with regard to exercise (28.7 +/- 8 to 9.1 +/- 3.2 ml.min-1, p less than 0.0001) but not significant changes in either LVAW subepicardial flow or RVAW flow. The transmural distribution of flows within the LAD bed (as percentages of the total LAD flow in each experimental condition) showed that LAD stenosis redistributed flows with regard to exercise such that the LVAW subendocardial flow decreased from 26.4 +/- 4.2% of the total LAD flow to 11.8 +/- 4.3% (p less than 0.0001), whereas LVAW subepicardial flow increased from 32.9 +/- 2.3% of the total LAD flow to 45.5 +/- 7.9% (p less than 0.0001) and RVAW increased from 12 +/- 4.9% of the total LAD flow to 18.7 +/- 7.2% (p less than 0.0005). With exercise plus LAD stenosis, LVAW %WTh decreased from 43.2 +/- 8.4% to 17.2 +/- 9.7% (p less than 0.0001), but RVAW %WTh did not change. CONCLUSIONS: In the LAD bed of exercising pigs, LAD stenosis induces, in addition to transmural steal, an interventricular steal favoring the RVAW at the expense of the LVAW subendocardium. This steal results in preserved RVAW thickening despite severe LVAW hypokinesia.

Effect of long-term oral nisoldipine on infarct size and collateral development in pigs undergoing gradual occlusion of the left circumflex artery.

We examined the effects of nisoldipine on infarct size and collateral development in pigs, whose coronary circulation is similar to that of humans, using an experimental protocol reproducing as closely as possible the usual clinical setting. Fifteen pigs undergoing left circumflex Ameroid-occlusion were randomized into a control group (n = 8) and a group (n = 7) treated with oral nisoldipine 0.03 mg/kg every 6 h for 1 month starting on the second postoperative day. Infarct size (tetrazolium red) was 37.2 +/- 9.2% of the circumflex distribution in the control group and 10 +/- 3.2% in the treated group (p less than 0.01). Endocardial and transmural blood flows (microspheres) in the circumflex distribution were significantly higher (p less than 0.05) in the treated group (control endocardial 1.25 +/- 0.1 mg/g/min, treated endocardial 1.77 +/- 0.26 ml/g/min; control transmural 1.39 +/- 0.08 ml/g/min; treated transmural 1.78 +/- 0.23 ml/g/min). Epicardial flow and the ratio of subendocardial to subepicardial blood flow (endo/epi) were nonsignificantly higher in treated pigs. No differences were observed in heart rate (HR) and aortic pressure (AP). We conclude that in pigs undergoing left circumflex Ameroid-occlusion, long-term oral nisoldipine reduces infarct size and enhances collateral circulation to the ischemic myocardium.

Astrocytic reaction predominance in chronic encephalitis of Junin virus-infected rats.

Junin virus antigen distribution and astrocytic reaction to prolonged infection were characterized in rat brain by the PAP technique. During the acute stage of neurologic disease following intracerebral inoculation, Junin antigen was detected in 100% of animals, strongly in most neurons but also to a much lesser degree in scattered astrocytes, dropping to 20% of rats at 540 days postinfection. Initially labeled in all brain areas, viral antigen gradually disappeared from hippocampus but persisted irregularly in cerebral cortex, basal ganglia, Purkinje cells, pons, and medulla oblongata. Such a pattern suggests that specific neuronal subpopulations, in spite of apparently unaltered cell morphology, may persistently harbor the virus, leading on occasion to a delayed neurologic syndrome. During both the acute and chronic stages of disease, a mild inflammatory exudate was observed, characterized by the presence of T and B lymphocytes, as well as macrophages and unidentified round cells. GFAP immunostaining showed increased astrocytic reaction as infection lapsed into chronicity. Corpus callosum, hippocampus, and cerebellum exhibited the sharpest reactive astrocytosis, followed by basal ganglia, pons, and medulla oblongata, whereas in cerebral cortex it was considerably less. Astrocyte activation, which failed to correlate with viral antigen presence in neurons, seems to result from a generalized condition, possibly including diffusible brain factors triggered by viral infection. Such widespread astroglial reaction may thus contribute to the outcome of the late neurologic syndrome.

Single-beat evaluation of left ventricular inotropic state in conscious dogs.

Two competing left ventricular elastic-resistive (ER) models were used to predict parameter values from pressure, volume, and time data of a single ejective beat in conscious dogs during control, enhanced (dobutamine), and decreased (propranolol) inotropic states. The animals were instrumented with three pairs of microcrystals and a transducer to measure intraventricular volume and pressure. Results showed that with the ER nonlinear model (ERNL), parameter values in all animals lay within the physiological range. These were the slope (Emax) and the intercept (V0) of the isovolumic end-systolic pressure-volume relationship (ESPVR), the slope of the end-diastolic pressure-volume relationship (Ed), the time to Emax (Tmax), the normalized time to end of activation (A), and the resistive constant (K). In the two models, the normalized SE of the estimate of data fitting was below 0.2 Emax, as estimated from a single beat, responded to changes in contractility in a significantly more consistent fashion than the slope of ESPVRs (Ees) generated by preload maneuvers in conscious dogs. Single-beat estimated Tmax and K with the ERNL model did also respond consistently to contractility changes, whereas with the elastic resistive linear (ERL) model, K did not reproduce the experimental findings with decreased inotropic state. We conclude that 1) the ERNL model can be employed to assess contractility changes in conscious dogs from data of a single ejective beat, and 2) these changes are better indicated by single-beat estimated Emax than by Ees calculated from conventional ESPVRs.

A controlled silver impregnation method to characterize cultured cardiomyocytes.

A morphological characterization of cultured cardiomyocytes was attempted using a modification of a silver impregnation technique originally described for connective tissue. Cardiac cells, obtained from newborn rats and grown as dissociated cultures on plastic surfaces, were fixed in methanol plus 5% glacial acetic acid, treated with potassium permanganate, decolorized in oxalic acid, sensitized with potassium bichromate, impregnated with a silver-ammonium complex, reduced in gelatin-formalin preparation, toned with gold chloride and fixed in sodium thiosulfate. The cultured cardiac cells tended to form a monolayer, although many myocytes remained isolated. Spherical nuclei, sharply stained with silver, were centrally located and surrounded by relatively plentiful cytoplasm packed with well delineated myofibrils. Contaminating fibroblasts were readily distinguished by their spindle-shaped nuclei and the presence of overstained collagen fibers, as well as the absence of myofibrils. In the absence of specific antibody for immunocytochemical identification of cardiomyocytes, morphological characterization of cell type and degree of differentiation by the controlled silver impregnation procedure described here provides a viable alternative, both in short- and long-term studies.