Beyond Cardiac Function: Microstructure & Metabolism
Jürgen E. Schneider1
1University of Leeds, United Kingdom

Synopsis

The cardiac microarchitecture is a key determinant of the relevant functions of the heart, including elec­trophysiological properties and mechanical activity, affecting both systemic blood flow and coronary perfusion. Thus, it also fundamentally determines metabolism of the heart muscle as a constant supply of substrates is crucial for the heart’s ability to continuously pump blood through the body. The aim of this presentation is to discuss two techniques beyond routinely applied assessments of cardiac function, namely the use of Diffusion Magnetic Resonance Imaging to investigate the microstructure of the heart and MR Spectroscopy to interrogate metabolic processes in the myocardium.

Introduction

The cardiac microarchitecture is a key determinant of (patho-) physiologically relevant functions of the heart, including elec­trophysiological properties underlying normal conduction [1, 2], arrhythmias [3-6], and mechanical activity [7, 8] affecting not only systemic blood flow but also coronary perfusion [9, 10]. Thus, it also fundamentally determines metabolism of the heart muscle (‘myocardium’), as a constant supply of substrates through the metabolic network is paramount for the heart’s ability to continuously pump blood through the body. The aim of this presentation is to discuss two fields in preclinical MR research, which go beyond routinely applied assessments of cardiac function, namely the use of Diffusion Magnetic Resonance Imaging (DMRI) to investigate the microstructure of the heart and MR Spectroscopy (MRS) to interrogate metabolic processes in the myocardium, respectively. While either technique utilises fundamentally different measurement approaches, they both provide information at very small length scales (i.e. cellular information – DMRI; molecular information – MRS) without physically resolving the underlying structures: the observed signal is the average over ~102-105 cardiomyocytes.

Diffusion as a surrogate measure of tissue microstructure

The central role of the heart’s microstructure in both, health and disease has motivated the development of non-invasive diffusion-based imaging techniques, which measure the diffusion of water molecules in biological tissue due to Brownian motion. Water molecules therefore act as a sensitive marker of tissue integrity and cellular orientation. Diffusion is measured quantitatively by applying two magnetic field gradients. In biological tissue, this signal attenuation will not only depend on the degree of diffusion weighting (DW), but also on the direction of the applied gradients. Diffusion data can be modelled using the Diffusion Tensor, which requires at least six measurements with non-collinear DW plus one measurement with no / low DW. Diagonalising the tensor allows for determining the principal directions of diffusion and the corresponding diffusivities: the primary eigenvector indicates the cardiomyocyte orientation, the secondary eigenvector reflects local sheetlet orientations, whilst the third eigenvector describes the sheetlet-normal direction [11]. The many measurements combined with the requirement for sufficient signal-to-noise ratio (SNR) result in long scan times. Cardiac DMRI (cDMRI) in vivo is further confounded by motion of the beating heart, strain and perfusion. The ex vivo setting provides an opportunity to optimize and apply dMRI protocols under idealized conditions avoiding these confounds.

Application of cardiac DMRI

cDMRI has been used ex vivo on fixed hearts of sheep, pigs, rabbits, rats and mice [12-15] as well as on fresh cadaveric human hearts [16], describing a transition of cardiomyocyte orientation from epi- to endocardium as characterized by the helix-angle (HA). The HA is positive (left-handed) subepicardally, near-zero in the midwall, and negative (right-handed) subendocardially. The perfused heart setting for rodent and rabbit hearts allowed for investigation of microstructural changes due to cardiac motion [17-20]. Ex vivo cDMRI with histological validation has also been used to study remodelling in cardiac disease, such as myocardial infarction (MI) and dilated cardiomyopathy (DCM) [21, 22]. Changes specifically in fractional anisotropy (FA) in murine hearts with MI was consistent with lengthening of myocytes in the region of infarction [22]. Conversely, a decrease in mean diffusivity (MD) in DCM samples correlated with the appearance of calcium depositions in the myocardium, and may indicate hindered diffusion [21]. Only very few studies have applied cDMRI in animal models in vivo demonstrating microstructural changes between systole and diastole [15], benefits of higher-order moment-nulling of diffusion gradients [23], and the microstructural impact of ischemia and bone marrow-derived cell therapy [24].

MRS as a window into metabolism

MRI uses the signal of water or fat protons, which are present in abundance in biological tissue: the concentration of water protons in the myocardium is about ~85 mol/l. In MRS, however, the signals of metabolites, which are present in many orders of magnitude lower concentration (typically around 5-20 mM), are interrogated. Additionally, nuclei other than protons, such as 13C or 31P, have a lower MR sensitivity. Taking the natural abundance into account, the absolute sensitivity is a factor of 1.8 · 10-4 for 13C and of 6.6 · 10-2 for 31P lower compared to protons. Therefore, low metabolite concentrations in the presence of cardiac and respiratory motion pose major challenges for preclinical implementation of cardiac MRS. 1H-MRS is additionally hampered by the presence of a dominating water signal, which needs to be suppressed sufficiently in order to detect the weak metabolite signals. Thus, the signals detectable with MRS are fundamentally much weaker than those of MRI, making it a relatively low-resolution technique.

Examples of cardiac MRS

Cardiac 1H-MR spectroscopy detects several metabolites such as lactate, lipids and creatine that can be used as indicators for the physiological condition of myocardial tissue. 1H-MRS studies in dogs have previously been reported [25, 26], and we and others have used this technique to investigate creatine [27] and lipid [28, 29] metabolism in the murine heart.
31P-MRS, which measures the signals of phosphocreatine (PCr), ATP and inorganic phosphate allows investigation of cardiac energy metabolism. The intracellular pH can be calculated from the frequency shift between the resonances of PCr and inorganic phosphate. Single-voxel [30, 31] and CSI [32-35] techniques have been used to characterize the high-energy phosphate metabolism in mice.
The signal of sodium represents an intrinsic marker of myocardial viability: A cellular ion-pump maintains a transmembrane concentration gradient between the intracellular and the extracellular space (intracellular sodium concentration 10-15 mM; extracellular 145 mM [36]). This energy-consuming process is compromised during periods of ischemia, leading to elevated intracellular sodium concentrations. Furthermore, the extra-cellular space increases in scar tissue after myocardial infarction. Thus, changes in the detectable sodium signal may be an indicator of cell viability, as shown in the isolated rat heart model (e. g. [36, 37]), or in vivo in mice [38], rabbits [39], dogs [40, 41].
Dynamic Nuclear Polarization (DNP) has been demonstrated to increase the sensitivity of 13C-MRS more than 10,000-fold [42, 43]. The application in animal models in vivo such as mouse [44], rat [45, 46] and pig [47-51] has enabled a greater understanding of the metabolic changes that occur as a consequence of cardiac diseases such as myocardial infarction and / or hypertrophy.

Acknowledgements

J.E.S would like to acknowledge funding from the British Heart Foundation (PG/17/28/32943) and the Medical Research Council (MR/M008991/1).

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Proc. Intl. Soc. Mag. Reson. Med. 28 (2020)