Left Ventricle Of Human Heart Research Articles

Gut microbe-generated phenylacetylglutamine is an endogenous allosteric modulator of β2-adrenergic receptors

Allosteric modulation is a central mechanism for metabolic regulation but has yet to be described for a gut microbiota-host interaction. Phenylacetylglutamine (PAGln), a gut microbiota-derived metabolite, has previously been clinically associated with and mechanistically linked to cardiovascular disease (CVD) and heart failure (HF). Here, using cells expressing β1- versus β2-adrenergic receptors (β1AR and β2AR), PAGln is shown to act as a negative allosteric modulator (NAM) of β2AR, but not β1AR. In functional studies, PAGln is further shown to promote NAM effects in both isolated male mouse cardiomyocytes and failing human heart left ventricle muscle (contracting trabeculae). Finally, using in silico docking studies coupled with site-directed mutagenesis and functional analyses, we identified sites on β2AR (residues E122 and V206) that when mutated still confer responsiveness to canonical β2AR agonists but no longer show PAGln-elicited NAM activity. The present studies reveal the gut microbiota-obligate metabolite PAGln as an endogenous NAM of a host GPCR.

Non-Uniform Bioaccumulation of Lead and Arsenic in Two Remote Regions of the Human Heart's Left Ventricle: A Post-Mortem Study.

The extent of heavy-metal-induced cardiotoxicity is proportional to the levels of metal bioaccumulation, and it was previously assumed that heavy metals accumulate uniformly in the myocardium. Therefore, the aim of this study was to investigate concentrations of metals and metalloids in two distant regions of the left ventricle (LV), the base of the LV, and apex of the LV using inductively coupled plasma mass spectrometry (ICP-MS). We also examined the potential correlation between metal levels and the thickness of the interventricular septum in twenty LV specimens (ten from the base of LV and ten from the apex of LV) from 10 individuals (mean age 75 ± 6 years). We found significantly higher concentrations of arsenic and lead in the LV apex compared to the base of the LV. We also found a positive correlation between the concentrations of arsenic in the myocardium of LV and the thickness of the interventricular septum. Our results indicate that arsenic and lead accumulate to a higher extent in the apex of the LV compared to the base of the LV. Therefore, future studies designed to measure levels of metals in heart muscle should consider non-uniform accumulation of metals in the myocardium.

Catalytic antibodies in arrhythmogenic cardiomyopathy patients cleave desmoglein 2 and N-cadherin and impair cardiomyocyte cohesion

AimsArrhythmogenic cardiomyopathy (AC) is a severe heart disease predisposing to ventricular arrhythmias and sudden cardiac death caused by mutations affecting intercalated disc (ICD) proteins and aggravated by physical exercise. Recently, autoantibodies targeting ICD proteins, including the desmosomal cadherin desmoglein 2 (DSG2), were reported in AC patients and were considered relevant for disease development and progression, particularly in patients without underlying pathogenic mutations. However, it is unclear at present whether these autoantibodies are pathogenic and by which mechanisms show specificity for DSG2 and thus can be used as a diagnostic tool.Methods and ResultsIgG fractions were purified from 15 AC patients and 4 healthy controls. Immunostainings dissociation assays, atomic force microscopy (AFM), Western blot analysis and Triton X-100 assays were performed utilizing human heart left ventricle tissue, HL-1 cells and murine cardiac slices. Immunostainings revealed that autoantibodies against ICD proteins are prevalent in AC and most autoantibody fractions have catalytic properties and cleave the ICD adhesion molecules DSG2 and N-cadherin, thereby reducing cadherin interactions as revealed by AFM. Furthermore, most of the AC-IgG fractions causing loss of cardiomyocyte cohesion activated p38MAPK, which is known to contribute to a loss of desmosomal adhesion in different cell types, including cardiomyocytes. In addition, p38MAPK inhibition rescued the loss of cardiomyocyte cohesion induced by AC-IgGs.ConclusionOur study demonstrates that catalytic autoantibodies play a pathogenic role by cleaving ICD cadherins and thereby reducing cardiomyocyte cohesion by a mechanism involving p38MAPK activation. Finally, we conclude that DSG2 cleavage by autoantibodies could be used as a diagnostic tool for AC.

Selective blockade of AT1R and B1R modulates the expression of ACE2 ubiquitination partners

Angiotensin Converting Enzyme 2 (ACE2) is an important compensatory enzyme in the renin‐angiotensin system (RAS), converting Angiotensin (Ang)‐II to Ang‐(1‐7). We previously reported that Ang‐II mediates ACE2 internalization through its type 1 receptor (AT1R), followed by ubiquitination and degradation in lysosomes. We recently identified UBR1 and BRCC36, as potential ubiquitinase and deubiquitinase, respectively, modulating ACE2 expression in hypertensive conditions. Here, we aimed to evaluate the role of bradykinin B1R receptor in Ang‐II‐mediated ACE2 internalization and the enzyme’s ubiquitination partners in hypertension.C57BI/6J mice (3‐month‐old, n=3/group) were divided into 3 groups: control (Ctrl), B1R antagonist SSR240612 (SSR), AT1R antagonist Losartan (Los). Mice were implanted with blood pressure (BP) probes (telemetry) and upon recovery, baseline BP was recorded for 24 hours. Mice were then implanted subcutaneously with an osmotic pump containing Ang‐II (490 ng/Kg/min/4 weeks) or saline. BP was recorded weekly, for 4 weeks. After 2weeks of Ang‐II infusion, the mice were implanted with an additional osmotic pump containing Losartan (30 mg/Kg/min) or SSR240612 (10 mg/Kg/min) for 2 weeks. Animals were sacrificed and the heart was isolated from all groups and protein expression for UBR1, BRCC36 and ACE2 was assessed using capillary western. Clinical relevance was further assessed in human heart left ventricle tissues of normal and hypertensive donors.Ang‐II infusion induced a significant increase in mean BP in Ctrl group (159.2±1.5 vs. 100.6±0.5 mmHg; one‐way ANOVA, **P<0.001) after 4 weeks of infusion. This was significantly attenuated with blockade of B1R or AT1R (SSR: 98.8±0.6; Los: 99.6±0.8 mmHg; n=3: one‐way ANOVA, **P<0.001). Capillary western with mouse heart tissue confirmed that Ang‐II‐induced a significant 2‐fold increase (*P<0.05) in UBR1 expression and the SSR240612 treatment contributed to a significant 4‐fold increase (**P<0.001). However, losartan had no effect. BRCC36 expression in Ang‐II, losartan and SSR240612 treated group was significantly decreased by 2‐3‐fold (*P<0.05). However, Ang‐II induced decrease in ACE2 levels was significantly attenuated by losartan and SSR240612 (**P<0.001), indicating both B1R and AT1R receptors contribute to ACE2 internalization and degradation. In human’s, the UBR1 expression was 2 folds (*P<0.05) higher in hypertensive CA male and AA female and BRCC36 expression showed 2 folds decrease (*P<0.05) in CA hypertensive females. However, ACE2 expression showed 3 folds decrease in hypertensive CA males, indicating elevated levels of UBR1 mediated ACE2 ubiquitination and degradation in hypertensive conditions.Our data show that Ang‐II‐mediated hypertension was attenuated with inhibition of both AT1R and B1R receptor, while B1R contributed to upregulation of UBR1 and both AT1R and B1R contributes to downregulation of BRCC36. Therefore, inhibition of both AT1R and B1R modulates the expression of ACE2 ubiquitination partners and also inhibits ACE2 internalization and degradation. Therefore, developing a treatments strategy and simultaneously targeting AT1R, B1R, UBR1 and BRCC36, involved in the regulation of ACE2 expression may be effective for the treatment of cardiovascular disorders.

Morpho-functional comparison of differentiation protocols to create iPSC-derived cardiomyocytes.

ABSTRACTCardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs) offer an attractive platform for cardiovascular research. Patient-specific iPSC-CMs are very useful for studying disease development, and bear potential for disease diagnostics, prognosis evaluation and development of personalized treatment. Several monolayer-based serum-free protocols have been described for the differentiation of iPSCs into cardiomyocytes, but data on their performance are scarce. In this study, we evaluated two protocols that are based on temporal modulation of the Wnt/β-catenin pathway for iPSC-CM differentiation from four iPSC lines, including two control individuals and two patients carrying an SCN5A mutation. The SCN5A gene encodes the cardiac voltage-gated sodium channel (Nav1.5) and loss-of-function mutations can cause the cardiac arrhythmia Brugada syndrome. We performed molecular characterization of the obtained iPSC-CMs by immunostaining for cardiac specific markers and by expression analysis of selected cardiac structural and ionic channel protein-encoding genes with qPCR. We also investigated cell growth morphology, contractility and survival of the iPSC-CMs after dissociation. Finally, we performed electrophysiological characterization of the cells, focusing on the action potential (AP) and calcium transient (CT) characteristics using patch-clamping and optical imaging, respectively. Based on our comprehensive morpho-functional analysis, we concluded that both tested protocols result in a high percentage of contracting CMs. Moreover, they showed acceptable survival and cell quality after dissociation (>50% of cells with a smooth cell membrane, possible to seal during patch-clamping). Both protocols generated cells presenting with typical iPSC-CM AP and CT characteristics, although one protocol (that involves sequential addition of CHIR99021 and Wnt-C59) rendered iPSC-CMs, which were more accessible for patch-clamp and calcium transient experiments and showed an expression pattern of cardiac-specific markers more similar to this observed in human heart left ventricle samples.

Construction of a mechanical model of the human heart left ventricle in the process of its contraction

A detailed analysis of previous works on modeling the left ventricle (LV) of the human heart, starting with mechanical models in the form of the simplest three-dimensional figures (cylinder, sphere, ellipsoid of rotation) and ending with models using real contours of the human heart obtained by ultrasound examination of the human heart. A way of constructing a mechanical LV model based on the processing of its dynamic images obtained using computer and magnetic resonance imaging was proposed. Digitization of these images was carried out using numerical methods developed to create a finite element model were implemented in the CMISS system, which allows the use of finite element analysis methods to solve various complex problems. To avoid the need to operate with huge arrays of numbers (up to 10 thousand numerical values at each time of the MRI study) characteristic points of the three-dimensional LV image were selected and spline interpolation of the model framework written in C++ using the Cmgui module was performed. To describe the work of the mechanical model of LV reduction, the main components of this process were identified: transverse compression, longitudinal contraction and twisting. To describe them, simple mathematical characteristics were used.

Drift of Scroll Waves in a Mathematical Model of a Heterogeneous Human Heart Left Ventricle

Rotating spiral waves of electrical excitation underlie many dangerous cardiac arrhythmias. The heterogeneity of myocardium is one of the factors that affects the dynamics of such waves. In this paper, we present results of our simulations for scroll wave dynamics in a heterogeneous model of the human left ventricle with analytical anatomically based representation of the geometry and anisotropy. We used a set of 18 coupled differential equations developed by ten Tusscher and Panfilov (TP06 model) which describes human ventricular cells based on their measured biophysical properties. We found that apicobasal heterogeneity dramatically changes the scroll wave dynamics. In the homogeneous model, the scroll wave annihilates at the base, but the moderate heterogeneity causes the wave to move to the apex and then continuously rotates around it. The rotation speed increased with the degree of the heterogeneity. However, for large heterogeneity, we observed formation of additional wavebreaks and the onset of complex spatio-temporal patterns. Transmural heterogeneity did not change the dynamics and decreased the lifetime of the scroll wave with an increase in heterogeneity. Results of our numerical experiments show that the apex may be a preferable location of the scroll wave, which may be important for development of clinical interventions.

Human cardiomyocyte calcium handling and transverse tubules in mid-stage of post-myocardial-infarction heart failure.

AimsCellular processes in the heart rely mainly on studies from experimental animal models or explanted hearts from patients with terminal end‐stage heart failure (HF). To address this limitation, we provide data on excitation contraction coupling, cardiomyocyte contraction and relaxation, and Ca2+ handling in post‐myocardial‐infarction (MI) patients at mid‐stage of HF.Methods and resultsNine MI patients and eight control patients without MI (non‐MI) were included. Biopsies were taken from the left ventricular myocardium and processed for further measurements with epifluorescence and confocal microscopy. Cardiomyocyte function was progressively impaired in MI cardiomyocytes compared with non‐MI cardiomyocytes when increasing electrical stimulation towards frequencies that simulate heart rates during physical activity (2 Hz); at 3 Hz, we observed almost total breakdown of function in MI. Concurrently, we observed impaired Ca2+ handling with more spontaneous Ca2+ release events, increased diastolic Ca2+, lower Ca2+ amplitude, and prolonged time to diastolic Ca2+ removal in MI (P < 0.01). Significantly reduced transverse‐tubule density (−35%, P < 0.01) and sarcoplasmic reticulum Ca2+ adenosine triphosphatase 2a (SERCA2a) function (−26%, P < 0.01) in MI cardiomyocytes may explain the findings. Reduced protein phosphorylation of phospholamban (PLB) serine‐16 and threonine‐17 in MI provides further mechanisms to the reduced function.ConclusionsDepressed cardiomyocyte contraction and relaxation were associated with impaired intracellular Ca2+ handling due to impaired SERCA2a activity caused by a combination of alteration in the PLB/SERCA2a ratio and chronic dephosphorylation of PLB as well as loss of transverse tubules, which disrupts normal intracellular Ca2+ homeostasis and handling. This is the first study that presents these mechanisms from viable and intact cardiomyocytes isolated from the left ventricle of human hearts at mid‐stage of post‐MI HF.

Effect of the form and anisotropy of the left ventricle on the drift of spiral waves

Three-dimensional spiral waves of electrical excitation in the myocardium are sources of dangerous cardiac arrhythmias. In this work, the dynamics of spiral waves of electrical excitation were studied in a symmetric anatomical model of the human heart left ventricle and a realistic ionic cell model of the human ventricular myocardium. Three factors that affect the drift waves in the heart were compared for the first time: the geometry of the heart wall, myocardial anisotropy, and wave chirality. Cardiac anisotropy was identified as a main factor in determining the drift of spiral waves. In the isotropic case, the dynamics were determined by the wall thickness, but did not depend on the wave chirality. In the anisotropic case, chirality was found to play a crucial role.

Morphological characteristics of the atypically located chordae of the human heart left ventricle

Morphological characteristics of the atypically located chordae of the human heart left ventricle

Three-dimensional structure of the flow inside the left ventricle of the human heart

The laboratory models of the human heart left ventricle developed in the last decades gave a valuable contribution to the comprehension of the role of the fluid dynamics in the cardiac function and to support the interpretation of the data obtained in vivo. Nevertheless, some questions are still opened and new ones stem from the continuous improvements in the diagnostic imaging techniques. Many of these unresolved issues are related to the three-dimensional structure of the left ventricular flow during the cardiac cycle. In this paper, we investigated in detail this aspect using a laboratory model. The ventricle was simulated by a flexible sack varying its volume in time according to a physiologically shaped law. Velocities measured during several cycles on series of parallel planes, taken from two orthogonal points of view, were combined together in order to reconstruct the phase-averaged, three-dimensional velocity field. During the diastole, three main steps are recognized in the evolution of the vortical structures: (1) straight propagation in the direction of the long axis of a vortex ring originated from the mitral orifice; (2) asymmetric development of the vortex ring on an inclined plane; and (3) single vortex formation. The analysis of three-dimensional data gives the experimental evidence of the reorganization of the flow in a single vortex persisting until the end of the diastole. This flow pattern seems to optimize the cardiac function since it directs velocity towards the aortic valve just before the systole and minimizes the fraction of blood residing within the ventricle for more cycles.

A 2‐D gel reference map of the basic human heart proteome

We have undertaken the identification of basic proteins (pH 6-11) of the human heart using 2-DE. Tissue from the left ventricle of human heart was lysed and proteins were separated in the first dimension on pH 6-11 IPG strips using paper-bridge loading followed by separation on 12% SDS polyacrylamide gels in the second dimension. Proteins were then identified by mass spectrometry and analysed using Proline, a proteomic data analysis platform that was developed in-house. The proteome map contains 176 identified spots with 151 unique proteins and has been made available as part of the UCD-2DPAGE database at http://proteomics-portal.ucd.ie:8082. The associated mass spectrometry data have been submitted to PRIDE (Accession number ♯10098). This reference map, and the other heart reference maps available through the UCD-2DPAGE database, will aid further proteomic studies of heart diseases such as dilated cardiomyopathy and ischaemic heart disease.

CD117-positive Cells of the Heart: Progenitor Cells or Mast Cells?

Human cardiac stem/progenitor cells and their potential for repair of heart injury are a current hot topic of research. CD117 has been used frequently as a marker for identification of stem/progenitor cells in the heart. However, cardiac mast cells, which are also CD117(+), have not been excluded by credible means when selecting putative cardiac progenitors by using CD117 as a marker. We evaluated the relationship between CD117(+) cells and mast cells in the left ventricle of human hearts (n=5 patients, ages 1 week-75 years) with the well-established mast cell markers tryptase, toluidine blue, and thionine. A large number (85-100%) of CD117(+) cells in the human heart were specifically identified as mast cells. In addition, mast cells showed weak or moderate CD45 immunostaining signals. These results indicate that the majority of CD117(+) cells in the heart are mast cells and that these cells are distinctly positive for CD45, although staining was weak or moderate. These results strongly suggest that the newly reported CD117(+)/CD45(dim/moderate) putative cardiac progenitor cells are mast cells. The significance of this observation in stem cell research of the heart is discussed.

IMPROVED VSF ALGORITHM FOR SMOOTH SURFACE RECONSTRUCTION FROM SPARSE MEDICAL DATA

This paper presents a Modified Variational Splines Fitting (MVSF) algorithm for surface reconstruction using thin plate splines on scattered patches or points of originally smooth surfaces. In particular, a more accurate derivation of the discrete equations for the energy corresponding to the thin plate model is introduced. The results obtained on simulated data show that the proposed algorithm converges faster than the original VSF algorithm. Additionally, we discuss an approach for choosing the algorithm’s parameters using a cross validation technique. Results obtained with the modified algorithm are compared to those using a Frequency Fourier-based 3D Harmonic modelling (3DHM) algorithm and show that the proposed algorithm gives an improved performance under the small sample size condition. The developed model has been successfully applied for real biomedical data; in particular for the reconstruction of left ventricle of human heart.

The human heart proteome: Two‐dimensional maps using narrow‐range immobilised pH gradients

The analysis of complex proteomes is undertaken using a variety of techniques and technologies such as 2-DE, surface-enhanced laser desorption ionisation, and various types of MS. In order to overcome the complexities of protein expression in discrete proteomes, sample fractionation has become an important aspect of proteomic experiments. The use of narrow-range IPGs (nrIPGs) is of special importance using the 2-DE proteomics workflow, since an enhanced visualisation of a given proteome is achieved through an improved physical separation and resolution of proteins. The work described in this paper presents a series of protein maps of the human heart left ventricle proteome that have been generated using nrIPGs for the first, IEF, dimension of 2-DE. A total of 374 gel spots were excised from seven different pH gradients, covering the range pH 3-10, giving rise to a total of 388 identifications from 110 unique proteins. Using Gene Ontologies (GOs), the identified proteins were found to be associated with 97 types of GO Process, 144 types of GO Function, and 54 types of GO Component. It is hoped that the maps presented in this paper will be of use to other researchers for reference purposes.

Age-Related and Individual Variability of Parameters of Work of the Human Heart Left Ventricle

Age-Related and Individual Variability of Parameters of Work of the Human Heart Left Ventricle

Vasoactive intestinal peptide has a direct positive inotropic effect on isolated human myocardial trabeculae.

The aim of the present study was to assess the inotropic effects of vasoactive intestinal peptide (VIP) on isolated myocardial trabeculae from the right atrium and the left ventricle of human hearts. Furthermore, using reverse transcriptase-PCR, we wanted to determine the presence of mRNAs encoding the three cloned human VIP receptors, VPAC(1), VPAC(2) and PAC(1). The trabeculae were paced at 1.0 Hz in tissue baths, and changes in isometric contractile force upon exposure to agonist were studied. VIP had a potent positive inotropic effect in some of the atrial and ventricular trabeculae tested. This effect was almost completely blocked by the VIP-receptor antagonist VIP-(6-28). mRNAs encoding the human VPAC(1), VPAC(2) and PAC(1) receptors were detected in human myocardial trabeculae from both the right atrium and the left ventricle. In conclusion, VIP has a direct positive inotropic effect in both the atria and the ventricles of the human heart. The presence of mRNAs for the VPAC(1), VPAC(2) and PAC(1) receptors suggest that VIP may mediate its effect via these receptors.

Vasoactive intestinal peptide has a direct positive inotropic effect on isolated human myocardial trabeculae

The aim of the present study was to assess the inotropic effects of vasoactive intestinal peptide (VIP) on isolated myocardial trabeculae from the right atrium and the left ventricle of human hearts. Furthermore, using reverse transcriptase-PCR, we wanted to determine the presence of mRNAs encoding the three cloned human VIP receptors, VPAC1, VPAC2 and PAC1. The trabeculae were paced at 1.0Hz in tissue baths, and changes in isometric contractile force upon exposure to agonist were studied. VIP had a potent positive inotropic effect in some of the atrial and ventricular trabeculae tested. This effect was almost completely blocked by the VIP-receptor antagonist VIP-(6-28). mRNAs encoding the human VPAC1, VPAC2 and PAC1 receptors were detected in human myocardial trabeculae from both the right atrium and the left ventricle. In conclusion, VIP has a direct positive inotropic effect in both the atria and the ventricles of the human heart. The presence of mRNAs for the VPAC1, VPAC2 and PAC1 receptors suggest that VIP may mediate its effect via these receptors.

The neonatal heart has a relatively high content of total collagen and type I collagen, a condition that may explain the less compliant state

Objectives. This study evaluated the extent of the collagen network in neonatal heart muscle and whether the type I/type III collagen ratio is the same as in the adult heart.Background. The functional integrity and the stress-strain relation of heart muscle depends largely on the extracellular collagen matrix. The question therefore arises whether the altered compliance of the neonatal heart could relate to the developmental state of collagen and, in particular, the distribution of types I and III collagen. Type I collagen mainly provides rigidity and type III collagen elasticity.Methods. Specimens from the left lateral wall of the left ventricle of human hearts (immature to full term, n = 23; 3 weeks to 12 years, n = 17) were used to determine the total collagen amount, using the hydroxyproline assay. Similar left ventricular specimens of human hearts (fetal to mature, n = 20; 2 months to 1.5 years, n = 6) were fixed in formalin, paraffin embedded and stained with Sirius red F3BA for total collagen. The ratio of total collagen to total protein was quantified spectrophotometrically. Frozen sections of left ventricular myocardium (immature to mature, n = 17; 4 months to 12 years, n = 10) were stained with antibodies raised against types I and III collagen. Antibody titration was done on human leiomyoma tissue with a known type I/type III collagen ratio. The endomysial collagen types were quantified using a spectrophotometer and expressed as a ratio. Adult human myocardium (n = 10) was used as reference.Results. The study showed that the total amount of collagen increases with age. However, the ratio of total collagen to total protein and the ratio of type I to type III collagen were very high in hearts of the very young. During development, a gradual decrease occurred, with the total collagen/total protein ratio reaching normal levels at ~5 months after birth and the type I/type III collagen ratio stabilizing at a much later age.Conclusions. These findings suggest that the relative high content of collagen, related to the myocytes, and the high ratio of type I to type III collagen provide the substrate for a rigid, less compliant heart in neonates.

No upregulation of digitalis glycoside receptor (Na,K-ATPase) concentration in human heart left ventricle samples obtained at necropsy after long term digitalisation

The aim was to evaluate the hypothesis that digitalis glycosides increase the concentration of their specific receptor (Na,K-ATPase) in human myocardial tissue, thereby possibly reducing the inotropic effect of long term digitalis treatment. Intact samples of left ventricle were obtained at necropsy from patients who had been on long term treatment with digoxin and from patients not previously given digoxin. Digitalis glycoside receptors were quantified using vanadate facilitated 3H-ouabain binding before and after washing samples in buffer containing excess digoxin antibody fragments for 16 h at 30 degrees C. This washing procedure has previously been shown to reduce prior specific digoxin binding in human left ventricle by 95% and to allow subsequent vanadate facilitated complete quantification of 3H-ouabain binding sites. In this context it was performed to reduce occupancy of digitalis glycoside receptors by digoxin, caused by digitalisation before 3H-ouabain binding. 11 patients who had been on long term treatment with digoxin and eight who had not previously been given digoxin were studied. Left ventricle samples were obtained at necropsy at around 15 h after death. Standard 3H-ouabain binding was 39% less in samples from digitalised than from undigitalised subjects (p less than 0.001). Washing samples in buffer containing excess digoxin antibody fragments induced an increase in 3H-ouabain binding from 174(SEM 10) to 265(20) pmol.g-1 wet weight (n = 11, p less than 0.001) in samples from digitalised patients. After washing, the digitalis glycoside receptor concentration in left ventricle samples showed a tendency to a lower value (14%, p greater than 0.10) in patients exposed to digoxin compared to left ventricle samples from individuals unexposed to digitalis glycoside treatment. Calculating 3H-ouabain binding relative to dry ventricular muscle weight confirmed the results obtained using wet weight as reference. The results suggest that digoxin treatment in life is associated with a 34% occupancy of digitalis glycoside receptors with digoxin. In the human heart there was no evidence for upregulation of digitalis glycoside receptor concentration due to long term digitalisation. Thus at receptor level there was no evidence for development of tolerance to digoxin therapy. The lower digitalis glycoside receptor concentration in the left ventricle observed in the heart failure patients may support the report of a relationship between Na,K-ATPase concentration as evaluated by 3H-ouabain binding and left ventricular function.