Force-frequency Relationship Research Articles

Phospholamban and cardiac function: a comparative perspective in vertebrates

Phospholamban (PLN) is a small phosphoprotein closely associated with the cardiac sarcoplasmic reticulum (SR). Dephosphorylated PLN tonically inhibits the SR Ca-ATPase (SERCA2a), while phosphorylation at Ser16 by PKA and Thr17 by Ca(2+) /calmodulin-dependent protein kinase (CaMKII) relieves the inhibition, and this increases SR Ca(2+) uptake. For this reason, PLN is one of the major determinants of cardiac contractility and relaxation. In this review, we attempted to highlight the functional significance of PLN in vertebrate cardiac physiology. We will refer to the huge literature on mammals in order to describe the molecular characteristics of this protein, its interaction with SERCA2a and its role in the regulation of the mechanic and the electric performance of the heart under basal conditions, in the presence of chemical and physical stresses, such as β-adrenergic stimulation, response to stretch, force-frequency relationship and intracellular acidosis. Our aim is to provide the basis to discuss the role of PLN also on the cardiac function of nonmammalian vertebrates, because so far this aspect has been almost neglected. Accordingly, when possible, the literature on PLN will be analysed taking into account the nonuniform cardiac structural and functional characteristics encountered in ectothermic vertebrates, such as the peculiar and variable organization of the SR, the large spectrum of response to stresses and the disaptive absence of crucial proteins (i.e. haemoglobinless and myoglobinless species).

The Fast Skeletal Troponin Activator, CK-2017357, Increases Muscle Function and Survival in SOD1 (G93A) Mice; a Model of ALS (P05.169)

Objective: To examine the effects of CK-2017357 in a mouse model of ALS. Background CK-2017357 is a small molecule activator of fast skeletal troponin that sensitizes the sarcomere to calcium and amplifies the muscle9s response to motor neuron input at sub-tetanic forces. Design/Methods: The potential effects of CK-2017357 were examined on muscle force, respiratory function and survival in SOD1 G93A mutant transgenic mice, a model of amyotrophic lateral sclerosis (ALS) in humans. Results: Extensor digitorum longus (EDL) muscle was stimulated via the peroneal nerve over a range of frequencies (10-200 Hz, 350 ms trains) to establish the force-frequency relationship. EDL tetanic force (26.3 ± 4.4 N/cm 2 ) was reduced by 44% ± 3.9% in 90-100 day-old SOD1 vs. WT and 66% ± 7.7% in 100-110 day-old SOD1 vs. WT, reflecting progressive loss of muscle force with disease progression. Infusion of CK-2017357 resulted in dose-dependent increases (mean ± SEM) in force at 30 Hz compared to baseline in WT (206% ± 29%), 90-100 day-old (200% ± 52%) and 100-110 day-old (153% ± 55%) SOD1 G93A mice. In female SOD1 G93A mice fed chow containing CK-2017357 (400 ppm) beginning at 11 weeks of age, animals survived to a humane endpoint longer compared to control chow-fed mice (p=0.024; hazard ratio=0.66). Body weight was similar in the two groups. There were no differences in respiratory parameters (breathing frequency, tidal volume and ventilation during hypercapnic challenge) in CK-2017357 chow-fed SOD1 G93A mice compared to WT mice. SOD1 G93A mice fed 400 ppm CK-2017357 chow exhibited a significant increase in hindlimb grip strength compared to control chow fed SOD1 G93A mice, while forelimb grip strength was not altered. Conclusions: CK-2017357 increased isometric in situ muscle force and hindlimb grip strength in SOD1 G93A mice and improved survival. These results indicate that CK-2017357 has the potential to improve muscle function in patients suffering from ALS. Supported by: Cytokinetics, Inc. Disclosure: Dr. Malik has received personal compensation for activities with Cytokinetics, Inc. Dr. Malik holds stock and/or stock options in Cytokinetics, Inc. Dr. Malik has received research support from Cytokinetics, Inc. Mr. Russell holds stock and/or stock options in Former Cytokinetics, Inc. which sponsored research in which Mr. Russell was involved as an investigator. Dr. Pannirselvam holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Pannirselvam was involved as an investigator.Dr. Pannirselvam has received research support from Cytokinetics, Inc. Dr. Hinken holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Hinken was involved as an investigator.Dr. Hinken has received research support from Cytokinetics, Inc. Dr. Thomsen holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Thomsen was involved as an investigator. Dr. Thomsen has received research support from Cytokinetics, Inc. Dr. Ardiana holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Ardiana was involved as an investigator. Dr. Ardiana has received research support from Cytokinetics, Inc. as an employee. Dr. Godinez holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Godinez was involved as an investigator. Dr. Jia holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Jia was involved as an investigator. Dr. Saikali holds stock and/or stock options in Cytokinetics, Inc. Dr. Saikali has received research support from Cytokinetics, Inc. Dr. Chen holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Chen was involved as an investigator. Dr. Chen has received research support from Cytokinetics, Inc. Dr. Morgans holds stock and/or stock options in Cytokinetics, Inc. Dr. Morgans has received research support from Cytokinetics, Inc. Dr. Jasper holds stock and/or stock options in Cytokinetics, Inc., which sponsored research in which Dr. Jasper was involved as an investigator.Dr. Jasper has received research support from Cytokinetics, Inc..

Chronic Hypobaric Hypoxia Increases Isolated Rat Fast-Twitch and Slow-Twitch Limb Muscle Force and Fatigue

Chronic hypoxia alters respiratory muscle force and fatigue, effects that could be attributed to hypoxia and/or increased activation due to hyperventilation. We hypothesized that chronic hypoxia is associated with phenotypic change in non-respiratory muscles and therefore we tested the hypothesis that chronic hypobaric hypoxia increases limb muscle force and fatigue. Adult male Wistar rats were exposed to normoxia or hypobaric hypoxia (PB=450 mm Hg) for 6 weeks. At the end of the treatment period, soleus (SOL) and extensor digitorum longus (EDL) muscles were removed under pentobarbitone anaesthesia and strips were mounted for isometric force determination in Krebs solution in standard water-jacketed organ baths at 25 °C. Isometric twitch and tetanic force, contractile kinetics, force-frequency relationship and fatigue characteristics were determined in response to electrical field stimulation. Chronic hypoxia increased specific force in SOL and EDL compared to age-matched normoxic controls. Furthermore, chronic hypoxia decreased endurance in both limb muscles. We conclude that hypoxia elicits functional plasticity in limb muscles perhaps due to oxidative stress. Our results may have implications for respiratory disorders that are characterized by prolonged hypoxia such as chronic obstructive pulmonary disease (COPD).

Transient Analysis of Force–Frequency Relationships in Rat Hearts Perfused by Krebs-Henseleit and Tyrode Solutions with Different [Ca2+

Healthy hearts respond to enhancement of pacing rate by an increase of contractility exhibiting positive force–frequency relations (FFR). In failing hearts rhythm acceleration leads to the progressive decline in force that reflects negative FFR. Our goal is to estimate the effect of [Ca2+]o on the force transient dynamics and resulting FFR in perfused rat hearts. We study in details the transient force responses of Langendorff perfused hearts of rats to step changes in pacing period from 200 to 100 ms and back with a step of 20 ms. We find that the transient responses for both directions are biphasic and can last for more than 3–5 min, indicating that one must be careful in the interpretation of traditional FFR measurements. Also, we study the effects of different [Ca2+]o in Tyrode and Krebs-Henseleit solutions on FFR and show that FFR in rat heart can be changed from positive at 1 mM of [Ca2+]o in Tyrode solution to negative at 4 mM of [Ca2+]o. Furthermore, we find that perfusion with lower [Ca2+]o will give bigger at fast rates and longer lasting transient responses. On the other hand, transients are smaller and less sensitive to pacing period at higher [Ca2+]o. Force–frequency relationships can be analyzed not only upon steady state conditions; the structure of force transient after step changes in pacing period is dependent on extracellular calcium. Our data should be useful in the construction of detailed model of force generation in a cardiac tissue; they can be used to test the validity of models.

Skeletal muscle force is preserved in NOX4 deficient mice

NADPH oxidase levels are elevated in various cardiovascular diseases. The NOX4 isoform of NADPH oxidase generates reactive oxygen species that promotes calcium release in skeletal muscle. NOX4 knockdown depresses maximal tetanic force in mouse EDL muscle. We tested the hypotheses that NOX4 deficient mice would have decreased muscle force, rightward shift in the force‐frequency relationship, and accelerated fatigue in vitro. We studied intact soleus muscle from genetic control (C57BL/6J) and NOX 4 knockout mice (n = 7/group; 2 males/group) at 95%O2 and 37°C. Our initial experiments suggest no difference in maximal soleus tetanic force (Po) between groups (in N/cm2; Control 32 ± 6, NOX 4−/− 36 ± 4). Neither the stimulus frequency that elicited 50% Po (Control 53 ± 3 Hz; NOX4−/− 53 ± 8 Hz) nor muscle fatigue (not shown) was affected by NOX4 deficiency. Our study suggests that skeletal muscle force is preserved in mice lacking NOX4 isoform of NADPH oxidase.

Age‐dependent changes in contractile function and passive elastic properties of myocardium from mice lacking muscle LIM protein (MLP)

Muscle LIM protein (MLP) null mice are often used as a model for human dilated cardiomyopathy. So far, little is known about the time course and pathomechanisms leading to the development of the adult phenotype. We systematically analysed the contractile phenotype, myofilament calcium (Ca(2)(+)) responsiveness, passive myocardial mechanics, histology, and mRNA expression in mice aged 4 and 12 weeks. In 4-week-old animals, there was no significant difference in the force-frequency relationship (FFR) and catecholamine response of intact isolated papillary muscles between wild-type (WT) and MLP null myocardium. In 12-week-old animals, WT myocardium exhibited a significantly positive FFR, while that of MLP null mice was significantly negative, and the inotropic response to catecholamines was significantly reduced in MLP null mice. This time course of decline in contractile function was confirmed in vivo by echocardiography. Whereas at 4 weeks of age MLP null mice and WT littermates showed similar levels of SERCA2a (sarcoplasmic reticulum Ca(2+) ATPase) expression, the expression was significantly lower in 12-week-old MLP null mice compared with littermate controls. Myofilament Ca(2)(+) responsiveness was not affected by the lack of MLP, irrespective of age. Whereas in 4-week-old animals MLP null myocardium showed a trend to an increased compliance compared with the WT, myocardium of 12-week-old MLP null mice was significantly less compliant than WT myocardium. Parallel to the decrease in compliance there was an increase in fibrosis in the MLP null animals. Our data suggest that MLP deficiency does not primarily influence myocardial contractility. A lack of MLP leads to an age-dependent impairment of excitation-contraction coupling with resulting contractile dysfunction and secondary fibrosis.

Effects of increased preload on the force-frequency response and contractile kinetics in early stages of cardiac muscle hypertrophy

Numerous studies have aimed to elucidate markers for the onset of decompensatory hypertrophy and heart failure in vivo and in vitro. Alterations in the force-frequency relationship are commonly used as markers for heart failure with a negative staircase being a hallmark of decompensated cardiac function. Here we aim to determine the functional and molecular alterations in the very early stages of compensatory hypertrophy through analysis of the force-frequency relationship, using a novel isolated muscle culture system that allows assessment of force-frequency relationship during the development of hypertrophy. New Zealand white male rabbit trabeculae excised from the right ventricular free wall were utilized for all experiments. Briefly, muscles held at constant preload and contracting isometrically were stimulated to contract in culture for 24 h, and in a subset up to 48 h. We found that, upon an increase in the preload and maintaining the muscles in culture for up to 24 h, there was an increase in baseline force produced by isolated trabeculae over time. This suggests a gradual compensatory response to the impact of increased preload. Temporal analysis of the force-frequency response during this progression revealed a significant blunting (at 12 h) and then reversal of the positive staircase as culture time increased (at 24 h). Phosphorylation analysis revealed a significant decrease in desmin and troponin (Tn)I phosphorylation from 12 to 24 h in culture. These results show that even very early on in the compensatory hypertrophy state, the force-frequency relationship is already affected. This effect on force-frequency relationship may, in addition to protein expression changes, be partially attributed to the alterations in myofilament protein phosphorylation.

Computational Analysis of Contractility in Engineered Heart Tissue

Engineered heart tissue (EHT) is a potential therapy for heart failure and the basis of functional in vitro assays of novel cardiovascular treatments. Self-organizing EHT can be generated in fiber form, which makes the assessment of contractile function convenient with a force transducer. Contractile function is a key parameter of EHT performance. Analysis of EHT force data is often performed manually; however, this approach is time consuming, incomplete and subjective. Therefore, the purpose of this study was to develop a computer algorithm to efficiently and objectively analyze EHT force data. This algorithm incorporates data filtering, individual contraction detection and validation, inter/intracontractile analysis and intersample analysis. We found the algorithm to be accurate in contraction detection, validation and magnitude measurement as compared to human operators. The algorithm was efficient in processing hundreds of data acquisitions and was able to determine force-length curves, force-frequency relationships and compare various contractile parameters such as peak systolic force generation. We conclude that this computer algorithm is a key adjunct to the objective and efficient assessment of EHT contractile function.

Activation of fast skeletal muscle troponin as a potential therapeutic approach for treating neuromuscular diseases

Limited neuromuscular input results in muscle weakness in neuromuscular disease either because of a reduction in the density of muscle innervation, the rate of neuromuscular junction activation or the efficiency of synaptic transmission1. We developed a small molecule fast skeletal troponin activator, CK-2017357, as a means to increase muscle strength by amplifying the response of muscle when neuromuscular input is diminished secondary to a neuromuscular disease. Binding selectively to the fast skeletal troponin complex, CK-2017357 slows the rate of calcium release from troponin C and sensitizes muscle to calcium. As a consequence, the force-calcium relationship of muscle fibers shifts leftwards as does the force-frequency relationship of a nerve-muscle pair. In vitro and in vivo, CK-2017357 increases the production of force at sub-maximal stimulation rates. Importantly, we show that sensitization of the fast skeletal troponin complex to calcium improves muscle force and grip strength immediately after single doses of CK-2017357 in a model of neuromuscular disease, myasthenia gravis. Troponin activation may provide a new therapeutic approach to improve physical activity in diseases where neuromuscular function is compromised.

Levo-carnitine supplementation normalizes skeletal muscle functions in type 2 diabetic rats

Levo-carnitine is known to improve skeletal muscle contractile functions in healthy humans and rats. It has also been claimed to correct metabolic derangements in type 2 diabetes mellitus (T2DM). This randomized control trial was designed to determine the effect of levo-carnitine on isometric contraction, force frequency relationship and fatigue of skeletal muscles along with oxidative stress in T2DM. Sprague-dawley rats (n=90) were randomly divided in three equal groups. Control group remained healthy while T2DM was induced in the diabetic and carnitine groups. Then, carnitine group was administered levo-carnitine 200mg/kg/day intraperitonealy for 6 days. At 28th day, extensor digitorum longus (EDL) muscles were removed and their functions were assessed using iWorx data acquisition unit (AHK/214). Blood obtained by intra-cardiac sampling at 28th day was used for estimation of serum malondialdehyde (MDA) levels. Regarding muscle parameters, no significant difference was found in maximum isometric twitch tension, time-to-peak twitch tension and time-to-relax to 50% of the peak twitch tension amongst the three groups. However, carnitine group showed significant improvement in maximum fused tetanic tension, maximum fused tetanic tension after fatigue protocol and recovery from fatigue after 5 minutes of rest period when compared with diabetic group. Serum MDA levels were reduced in carnitine group as compared to the diabetic group. These findings suggest that levo-carnitine supplementation reduces oxidative stress, improves work capacity and delays fatigue in T2DM.

Feed Forward Modeling. Fixing the Force Frequency Relationship

The force frequency relationship has intrigued researchers since its discovery by Bowditch in 1871. Several attempts were made to construct mathematical descriptions of this phenomenon; from the simple formulation of Koch-Wesser & Blinks in 1963 to the most sophisticated ones of present days. This property of the cardiac muscle is amplified by the β adrenergic stimulation. In a coordinated way the neurohumoral state alters both: frequency (acting on the SA node) and force generation (modifying the ventricular myocytes). This synchronized tuning is needed to meet new metabolic demands. Failure to do so has deleterious consequences. We implemented this physiological coordination in a new version of LabHEART (v5.5) where the cell parameters are updated according to the frequency of stimulation in a sigmoidal fashion (namely: ICaL, IKs, SERCA pump and myofilaments’ Ca-sensitivity). This feed forward modeling helps to reproduce a more realistic cell behavior and complement the information obtained by experiments where frequency has been altered but without adding β-adrenergic agonists. Plots like rate-adaptation or APD vs cycle length can, now, be mapped into the neurohumoral state to provide insights about arrhythmias (or antiarrhythmic drugs). Cardiac modelers have already linked the mechanical and the electrical activity in their formulations and showed how those activities feedback on each other. Feed forward modeling is a step towards reproducing the neurological control and thus, to have a complete description of the heart performance, particularly when changes in frequency are involved. The presumptive sigmoidal dependence of parameters on pacing frequency used here can be refined but is a valid starting point to implement the convergence of cardiac chronotrophy and inotropy.

Normal and Abnormal Ca Cycling during Rapid Pacing and the Development of Triggered Waves in Dog Left Atrial Myocytes

Although there have been a number of studies of Ca cycling in atrial myocytes, few have investigated the effects of rapid pacing on Ca cycling, as would occur during atrial fibrillation (AF). We investigated subcellular Ca cycling during rapid pacing in left atrial myocytes using high resolution confocal imaging in order to study rate-dependent changes in Ca cycling that might occur during AF. Myocytes were isolated from dog left atrium, loaded with fluo-4AM and studied during pacing at basic cycle lengths ranging from 5000-200msec (36oC). Myocytes showed a positive force-frequency relationship and normal rapid restitution. However, at BCL≤300, Ca transient magnitude abruptly decreased[GLA1] and multiple Ca waves developed during rapid pacing. Many waves were very large in magnitude and comparable to that inducible by caffeine. These triggered waves only develop during stimulation and are absent at rest. Triggered wave frequency and velocity both increased with rate and with increasing diastolic Ca levels. We also found this highly unusual Ca wave behavior in myocytes of intact dog left atrium during rapid pacing, underscoring the fact that this form of Ca wave behavior is not unique to isolated myocytes and occurs in normal atrium. Our results demonstrate that Ca cycling becomes highly abnormal at high heart rates and is characterized by highly fragmented Ca waves that occur at a high frequency. It is likely that these forms of Ca dysregulation could contribute to Ca overload and triggered activity during AF.

Mild metabolic acidosis impairs the β-adrenergic response in isolated human failing myocardium

IntroductionPronounced extracellular acidosis reduces both cardiac contractility and the β-adrenergic response. In the past, this was shown in some studies using animal models. However, few data exist regarding how the human end-stage failing myocardium, in which compensatory mechanisms are exhausted, reacts to acute mild metabolic acidosis. The aim of this study was to investigate the effect of mild metabolic acidosis on contractility and the β-adrenergic response of isolated trabeculae from human end-stage failing hearts.MethodsIntact isometrically twitching trabeculae isolated from patients with end-stage heart failure were exposed to mild metabolic acidosis (pH 7.20). Trabeculae were stimulated at increasing frequencies and finally exposed to increasing concentrations of isoproterenol (0 to 1 × 10-6 M).ResultsA mild metabolic acidosis caused a depression in twitch-force amplitude of 26% (12.1 ± 1.9 to 9.0 ± 1.5 mN/mm2; n = 12; P < 0.01) as compared with pH 7.40. Force-frequency relation measurements yielded no further significant differences of twitch force. At the maximal isoproterenol concentration, the force amplitude was comparable in each of the two groups (pH 7.40 versus pH 7.20). However, the half-maximal effective concentration (EC50) was significantly increased in the acidosis group, with an EC50 of 5.834 × 10-8 M (confidence interval (CI), 3.48 × 10-8 to 9.779 × 10-8; n = 9), compared with the control group, which had an EC50 of 1.056 × 10-8 M (CI, 2.626 × 10-9 to 4.243 × 10-8; n = 10; P < 0.05), indicating an impaired β-adrenergic force response.ConclusionsOur data show that mild metabolic acidosis reduces cardiac contractility and significantly impairs the β-adrenergic force response in human failing myocardium. Thus, our results could contribute to the still-controversial discussion about the therapy regimen of acidosis in patients with critical heart failure.

Strain, Strain Rate, and the Force Frequency Relationship in Patients with and without Heart Failure

The aim of this study was to examine the effect of heart rate (HR) on indices of deformation in adults with and without heart failure (HF) who underwent simultaneous high-fidelity catheterization of the left ventricle to describe the force-frequency relationship. Right atrial pacing to control HR and high-fidelity recordings of left ventricular (LV) pressure were used to inscribe the force-frequency relationship. Simultaneous two-dimensional echocardiographic imaging was acquired for speckle-tracking analysis. Thirteen patients with normal LV function and 12 with systolic HF (LV ejection fraction, 31 ± 13%) were studied. Patients with HF had depressed isovolumic contractility and impaired longitudinal strain and strain rate. HR-dependent increases in LV+dP/dt(max), the force-frequency relationship, was demonstrated in both groups (normal LV function, baseline to 100 beats/min: 1,335 ± 296 to 1,564 ± 320 mm Hg/sec, P < .0001; HF, baseline to 100 beats/min: 970 ± 207 to 1,083 ± 233 mm Hg/sec, P < .01). Longitudinal strain decreased significantly (normal LV function, baseline to 100 beats/min: 18.0 ± 3.5% to 10.8 ± 6.0%, P < .001; HF: 9.4 ± 4.1% to 7.5 ± 3.4%, P < .01). The decrease in longitudinal strain was related to a decrease in LV end-diastolic dimensions. Strain rate did not change with right atrial pacing. Despite the inotropic effect of increasing HR, longitudinal strain decreases in parallel with stroke volume as load-dependent indices of ejection. Strain rate did not reflect the modest HR-related changes in contractility; on the other hand, the use of strain rate for quantitative stress imaging is also less likely to be confounded by chronotropic responses.

Abstract 16766: A Human Induced Pluripotent Stem Cell-Derived Engineered Cardiac Tissue Model of Pompe Disease

Background: Infantile-onset Pompe disease is a lethal autosomal recessive lysosomal storage disorder caused by the complete loss of acid α-glucosidase activity resulting in glycogen accumulation within lysosomes. Skeletal and cardiac muscle pathology develop due to aberrant glycogen metabolism and lysosomal dysfunction. This work presents the first in vitro human cardiac tissue model of Pompe disease documenting the classic Pompe cellular phenotype within a multicellular engineered cardiac tissue preparation. Methods: Pompe disease-specific human induced pluripotent stem (iPS) cell derived cardiomyocyte (iPS-CMs) were seeded into a polymerizing fibrin mixture to form mechanically integrated multicellular engineered cardiac tissue (ECT) constructs that were evaluated over a one-month period in parallel with healthy human iPS cell derived ECTs. Weekly measurements of isometric force and intracellular calcium transients were recorded under varying work load imposed by electrical pacing and β1 adrenergic stimulation. Structural assessment was performed using electron microscopy and immunohistochemistry. Results: Healthy and Pompe iPS-CMs can form cylindrically shaped ECTs of 1-2 mm diameter and 1.5-2 cm in length composed of coupled CMs that generate force in the milli-newton range along the cylindrical axis. We report structural changes in the ECT model consistent with the hallmarks of Pompe disease including progressive intracellular glycogen accumulation, lysosomal disruption, and myofibrillar disarray. Both healthy and Pompe ECTs generate higher twitch forces in response to mechanical stretch, have a negative force-frequency relationship characteristic of immature myocardium, and have an inotropic response to dobutamine stimulation. Conclusion: Human iPS-CM ECTs obey the Frank-Starling law and produce calcium cycling and force kinetics that respond to β1 adrenergic stimulation. The Pompe ECT provides a novel cardiac tool in which to further study the pathophysiology of this disease and test novel therapeutic approaches.

Incidence and Outcome of Gastrointestinal Complications After Cardiac Surgery—A Ten-Year Experience of a Single Australian Centre

[2] Lus F, Hagl C, Haverich A, Pichlmaier M. Elephant trunk procedure 27 years after Borst: what remains and what is new? European Journal of Cardio-thoracic Surgery 2011;40:1–12. doi:10.1016/j.hlc.2011.08.064 2011 Poster Presentation/Panel 26 Incidence and Outcome of Gastrointestinal Complications After Cardiac Surgery—A Ten-Year Experience of a Single Australian Centre Fabiano Viana ∗, Yi Chen, Aubrey Almeida, Andrew Cochrane, Adrian Pick, Randall Moshinsky, Cliff Choong, Jacob Goldstein, Julian Smith Monash Medical Centre, Southern Health, Melbourne, Australia Introduction: Gastrointestinal (GI) complications after cardiac surgery are uncommon, but are associated with highmortality and prolonged hospital stay. The incidence, patient profile and outcome of GI complications after cardiac surgery in Australia have not been previously reported. Methods: We analysed a single-centre prospectively collected database containing all adult cardiac surgery procedures performed from July 2001 to March 2011 at Monash Medical Centre and Jessie McPherson’s Private Hospital. Patients with post-operative GI complications Conclusions: GI complications after cardiac surgery remain an uncommon but dreadful complication associated with high mortality. It is hoped that our findings will prompt a high degree of clinical vigilance and early diagnosis especially in selected high risk patients. Further studies aiming to identify independent predictors for GI complications after cardiac surgery in our population would be warranted. doi:10.1016/j.hlc.2011.08.065 2011 Poster Presentation/Panel 27 A Remote Ischaemic Preconditioning Protocol Does not Alter the Function of Human Right Atrium In Vitro Yasmin Whately 1,∗, Peter Molenaar 1, Katherine Gillette 2, Bronwyn Pearse1, John Fraser 1 1 Critical Care Research Group, Prince Charles Hospital, University of Queensland, Brisbane, Australia 2 Queensland University of Technology, Brisbane, Australia Introduction:Remote ischaemicpreconditioning (RIPC) is the phenomenon in which non-lethal ischaemia and reperfusion of one organ or tissue confers resistance to subsequent ischaemia reperfusion injury in a remote organ. Early clinical trials have shownpotential benefits in humans undergoing cardiac surgery.We aimed to investigate the effects of an RIPC protocol on human right atrial were compared with patients without GI complications who were operated in the same period. Statistical analyses were carried out in SPSS using t-test and chi-square test where appropriate. A p-value≤ 0.05 is considered statistically significant. Results: The incidence of GI complications was 1.1% (60 out of 5393 patients) with an overall 30-day mortality of 25.8%. Themost common complications involved GI bleeding, perforatedduodenal ulcer andbowel ischaemia. Preoperatively, patients that had GI complications were significantly older (69 vs 64 years), had higher creatinine levels (155 vs 108 mol/l) and higher incidence of chronic lung disease (21 vs 11%), anticoagulation therapy (24 vs 11%), shock (9 vs 2%) and inotrope usage (6 vs 2%). Emergency, combined CABG and valve, aortic dissection and anti-arrhythmic surgery cases occurred more frequently in theGI complication group (14 vs 5%, 21 vs 10%, 11 vs 3%, 14 vs 2%, respectively). Aortic cross-clamp and CPB times were significantly higher in the GI complication group (88 vs 77, 137 vs 105min). Regarding outcomes, the GI complication group had higher incidence of return to theatre (42 vs 5%), renal failure (36 vs 6%),myocardial infarction (8 vs 0.6%), cardiogenic shock (9 vs 0.5%), stroke (12 vs 1%), prolonged ventilation (51 vs 12%), septicaemia (16 vs 1%) and multi-organ failure (24 vs 0.5%). In the GI complication group, those who died had a higher CPB time (197 vs 116min) and higher incidence of emergency cases (24 vs 10%), return to theatre (65 vs 35%), renal failure (70 vs 24%), cardiogenic shock (18 vs 6%), stroke (35 vs 4%), prolonged ventilation (70 vs 45%) and septicaemia (29 vs 12%) (p≤ 0.05 for all comparisons). function in vitro. Methods: Samples of right atrial appendage were obtained from 10 patients undergoing coronary artery bypass grafting. Patients were randomised to receive the RIPC protocol (four cycles of inflation of a blood pressure cuff placed on the upper limb inflated to 200mmHg for 5min followedby5minof reperfusion) or shamplacement of the cuff (control). Immediately prior to termination of cardiopulmonary bypass, the right atrial appendage was removed. Trabeculae were isolated, set up on electrode tissue blocks and electrically stimulated. Force–frequency relationships (6–120 bpm)and concentration–effect curves for 5HTwere established followed bymaximal concentrations of isoprenaline (200mM) and calcium (7mM) were established. Serial troponinswere obtained preandpostoperatively. Results: The force–frequency relationships for both groupswerepositive.Therewerenodifferences inpotency to inotropic (−logEC50 RIPC protocol 6.88± 0.08, n= 15 trabeculae/5 hearts; control 6.69± 0.16, n= 12 trabeculae/5 hearts; P= 0.27 or maximal effects (P= 0.17) of 5HT. Peak troponins were not significantly different (P= 0.86). Conclusions:Our results show that RIPC does not alter the function of human right atrium in vitro. doi:10.1016/j.hlc.2011.08.066

Influence of Mild Metabolic Acidosis on Cardiac Contractility and Isoprenaline Response in Isolated Ovine Myocardium

The postoperative course after major surgical procedures such as cardiothoracic operations is often accompanied by acute metabolic abnormalities due to large volume and temperature shifts. In general, those intervention-induced trauma might cause the use of catecholamines to stabilize hemodynamics. Within the cardiac community, there are still controversial discussions about standardized medical therapy to treat postoperative acidosis, for example, buffering versus nonbuffering for improving catecholaminergic response of myocardial contractility. The aim of this study was to investigate the influence of mild (and thus clinically relevant) acidosis on myocardial contractility and catecholamine response in explanted trabeculae of ovine hearts. Intact trabeculae (n = 24) were isolated from the right ventricle of healthy sheep hearts. Two different groups (group 1: pH = 7.40, n = 9 and group 2: pH = 7.20, n = 13) were investigated, and force amplitudes were measured at frequencies between 30 and 180 beats per minute and increasing catecholamine concentrations (isoprenaline 0-3 × 10(-6) mM). Force-frequency relation experiments in the presence of a physiological and/or mild acidotic pH solution showed no significant differences. Mean force amplitudes normalized to the lowest frequency showing no significant differences in force development between 0.5 and 3 Hz (n = 9 vs. 13, P = n.s.) (0.5 Hz absolute values 3.1 ± 2.6 for pH = 7.40 vs. 3.8 ± 2.6 mN/mm(2) for pH = 7.20, P = n.s.). Moreover, there was no significant difference in relaxation kinetics between the two groups. Furthermore, the experiments showed similar catecholamine responses in both groups. Force amplitudes normalized to baseline and maximum force showed no significant differences in force development between baseline and maximum isoprenaline concentrations (n = 6 vs. 9, P = n.s.) (baseline absolute values 4.3 ± 4.0 for pH = 7.40 vs. 3.9 ± 1.2 mN/mm(2) for pH = 7.20, P = n.s.). Additionally, relaxation kinetics did not show differences after catecholamine stimulation. The presented experiments revealed no significant negative inotropic effects on isometrically contracting ovine trabeculae with mild metabolic acidosis (pH = 7.2) compared with physiological pH (7.4). Additionally, similar catecholamine responses were seen in both groups. Further investigations (e.g., in vivo and/or in failing hearts with reduced compensatory reserves) will be necessary to examine optimal medical treatment for metabolic abnormalities after cardiac surgery.

Procedures for Rat &lt;em&gt;in situ&lt;/em&gt; Skeletal Muscle Contractile Properties

There are many circumstances where it is desirable to obtain the contractile response of skeletal muscle under physiological circumstances: normal circulation, intact whole muscle, at body temperature. This includes the study of contractile responses like posttetanic potentiation, staircase and fatigue. Furthermore, the consequences of disease, disuse, injury, training and drug treatment can be of interest. This video demonstrates appropriate procedures to set up and use this valuable muscle preparation.To set up this preparation, the animal must be anesthetized, and the medial gastrocnemius muscle is surgically isolated, with the origin intact. Care must be taken to maintain the blood and nerve supplies. A long section of the sciatic nerve is cleared of connective tissue, and severed proximally. All branches of the distal stump that do not innervate the medial gastrocnemius muscle are severed. The distal nerve stump is inserted into a cuff lined with stainless steel stimulating wires. The calcaneus is severed, leaving a small piece of bone still attached to the Achilles tendon. Sonometric crystals and/or electrodes for electromyography can be inserted. Immobilization by metal probes in the femur and tibia prevents movement of the muscle origin. The Achilles tendon is attached to the force transducer and the loosened skin is pulled up at the sides to form a container that is filled with warmed paraffin oil. The oil distributes heat evenly and minimizes evaporative heat loss. A heat lamp is directed on the muscle, and the muscle and rat are allowed to warm up to 37°C. While it is warming, maximal voltage and optimal length can be determined. These are important initial conditions for any experiment on intact whole muscle. The experiment may include determination of standard contractile properties, like the force-frequency relationship, force-length relationship, and force-velocity relationship.With care in surgical isolation, immobilization of the origin of the muscle and alignment of the muscle-tendon unit with the force transducer, and proper data analysis, high quality measurements can be obtained with this muscle preparation.

Procedures for Rat &lt;em&gt;in situ&lt;/em&gt; Skeletal Muscle Contractile Properties

There are many circumstances where it is desirable to obtain the contractile response of skeletal muscle under physiological circumstances: normal circulation, intact whole muscle, at body temperature. This includes the study of contractile responses like posttetanic potentiation, staircase and fatigue. Furthermore, the consequences of disease, disuse, injury, training and drug treatment can be of interest. This video demonstrates appropriate procedures to set up and use this valuable muscle preparation. To set up this preparation, the animal must be anesthetized, and the medial gastrocnemius muscle is surgically isolated, with the origin intact. Care must be taken to maintain the blood and nerve supplies. A long section of the sciatic nerve is cleared of connective tissue, and severed proximally. All branches of the distal stump that do not innervate the medial gastrocnemius muscle are severed. The distal nerve stump is inserted into a cuff lined with stainless steel stimulating wires. The calcaneus is severed, leaving a small piece of bone still attached to the Achilles tendon. Sonometric crystals and/or electrodes for electromyography can be inserted. Immobilization by metal probes in the femur and tibia prevents movement of the muscle origin. The Achilles tendon is attached to the force transducer and the loosened skin is pulled up at the sides to form a container that is filled with warmed paraffin oil. The oil distributes heat evenly and minimizes evaporative heat loss. A heat lamp is directed on the muscle, and the muscle and rat are allowed to warm up to 37°C. While it is warming, maximal voltage and optimal length can be determined. These are important initial conditions for any experiment on intact whole muscle. The experiment may include determination of standard contractile properties, like the force-frequency relationship, force-length relationship, and force-velocity relationship. With care in surgical isolation, immobilization of the origin of the muscle and alignment of the muscle-tendon unit with the force transducer, and proper data analysis, high quality measurements can be obtained with this muscle preparation.

Impaired Diastolic Function and Disruption of the Force-Frequency Relationship in the Right Ventricle of Newborn Pigs Resuscitated with 100% Oxygen

Background: Resuscitation with 100% oxygen increases oxidative stress and is detrimental for organ function. Objective: To study the effects of resuscitation with 100% oxygen compared to room-air on myocardial function. Methods: Twenty-eight newborn pigs underwent global hypoxia (8% oxygen/N₂) until base excess reached –20 mmol/l. The animals were randomized into two groups and resuscitated with either 100% or room air for 30 min. Myocardial tissue Doppler velocities and acceleration of the mitral and tricuspid valve annuli during systole and diastole were assessed before global hypoxia and after resuscitation together with troponin I. Results: Peak early diastolic velocity (E′) and acceleration (pEac) in the septum and pEac in the lateral tricuspid valve annulus were lower after resuscitation with 100% oxygen, suggesting impaired diastolic relaxation in the right ventricle. Lower systolic velocities and acceleration in the right ventricle relative to heart rate indicate disruption of the right ventricular force-frequency relationship after resuscitation with 100% oxygen. Troponins were higher in the 100% oxygen group, suggesting increased myocardial damage in this group. Conclusion: Resuscitation with 100% oxygen compared to room air induces diastolic dysfunction, disrupts the systolic force-frequency relationship and increases myocardial damage in the newborn pig.