Structural and Functional Determinants of Cardiac Impulse Propagation and Arrhythmias.

Each year, sudden cardiac death (SCD) attributed to ventricular fibrillation (VF) kills approximately 200,000 people in the United States. However, the mechanisms responsible for VF, and therefore VF-related SCD, are incompletely understood. My PhD studies focused on two major topics directly related to the mechanisms of reentry in VF. My general approach was based on the use of neonatal cardiac cell monolayers, gene transfer, immunolocalization, patch clamping and optical mapping techniques. First, I examined how a delayed rectifier potassium channel gene (hERG) involved in cardiac repolarization affects reentry frequency in a ventricular myocyte monolayer model of reentry. The results provided strong evidence for a role of hERG in controlling the frequency and stability of reentry. The mechanisms underlying the acceleration in reentry frequency were shown to be action potential duration (APD) shortening and a transient hyperpolarization after each action potential. APD shortening reduced reentry wavelength which prevented wave front-wave tail interactions and increased reentry stability. The transient hyperpolarization enhanced sodium channel availability and excitability of tissue ahead of the propagating electrical wave front. Together they set the stage for fast and stable reentry that maintains VF. Second, I examined the principle of whether rescuing normal electrical impulse propagation in damaged or fibrotic myocardium using cell therapy would be an effective approach to alter reentry behavior. Electrically excitable cardiac fibroblasts were generated using viral constructs encoding Kir2.1, NaV1.5 and Cx43 proteins. Excitable fibroblasts were able to form monolayers and conduct electrical waves at high velocity. When used to replace normal fibroblasts in heterocellular monolayers, they significantly increased conduction velocity to values similar to those of pure myocytes monolayers. Moreover, during reentry, propagation was faster and more organized, with a significantly lower number of wavebreaks. Altogether, the work accomplished in my dissertation should lead to a better understanding of VF and to the development of novel therapeutic approaches for the prevention of SCD.PHDMolecular and Integrative PhysiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/98057/1/houl_1.pd

Electrical Coupling Between Micropatterned Cardiomyocytes and Stem Cells

To understand how stem cells functionally couple with native cardiomyocytes is crucial for cell-based therapies to restore the loss of cardiomyocytes that occurs during heart infarction and other cardiac diseases. Due to the complexity of the in vivo environment, our knowledge of cell coupling is heavily dependent on cell-culture models. However, conventional in vitro studies involve undefined cell shapes and random length of cell-cell contacts in addition to the presence of multiple homotypic and heterotypic contacts between interacting cells. Thus, it has not been feasible to study electrical coupling corresponding to isolated specific types of cell contact modes. To address this issue, we used microfabrication techniques to develop different geometrically-defined stem cell-cardiomyocyte contact assays to comparatively and quantitatively study functional stem cell-cardiomyocyte electrical coupling. Through geometric confinements, we will construct a matrix of identical microwells, and each was constructed as a specific microenvironment. Using laser-guided cell micropatterning technique, individual stem cells or cardiomyocytes can be deposited into the microwells to form certain contact mode. In this research, we firstly constructed an in-vivo like cardiac muscle fiber microenvironment, and the electrical conductivity of stem cells was investigated by inserting stem cells as cellular bridges. Then, the electrical coupling between cardiomyocytes and stem cells was studied at single-cell level by constructing contact-promotive/-preventive microenvironments

Multiscale Modeling of Cardiac Electrophysiology: Adaptation to Atrial and Ventricular Rhythm Disorders and Pharmacological Treatment

Multiscale modeling of cardiac electrophysiology helps to better understand the underlying mechanisms of atrial fibrillation, acute cardiac ischemia and pharmacological treatment. For this purpose, measurement data reflecting these conditions have to be integrated into models of cardiac electrophysiology. Several methods for this model adaptation are introduced in this thesis. The resulting effects are investigated in multiscale simulations ranging from the ion channel up to the body surface

Doctor of Philosophy

dissertationDoes strain induce changes in the electrical properties of the heart? Does strain affect the microstructure of cardiac myocytes? Others have considered these questions, but have been limited in their findings. I addressed the first question by measuring conduction velocity in papillary muscles in rest conditions and during applied strain. I also applied streptomycin, a nonselective stretch ion channel blocker, in the above conditions. In control, conduction velocity increased with strain before conduction block occurred. When streptomycin was applied conduction velocity peaked at a higher strain, but conduction block remained unchanged. Changes in electrical properties of papillary muscle allowed for changes in conduction velocity. Although streptomycin did not alter the strain at which conduction block occurred, it did shift the peak conduction velocity to a higher strain. The second question was addressed by imaging isolated cardiac ventricular myocytes in varying degrees of contraction and strain using confocal microscopy. The length of transverse tubules (t-tubules), along with cross-section ellipticity, and orientation in myocytes were analyzed for cells in 16% contraction, rest, and 16% strain. Cells in contraction showed an increase in length of t-tubules with less elliptical cross-sections compared to cells in rest. Strained cells showed a decrease in length of t-tubules with less elliptical cross-sections than cells at rest. The orientation of t-tubule cross-sections changed in a similar manner when comparing contracted and strained cells with cells at rest. The transfer of strain to the t-tubule system supports the hypothesis that the motion of t-tubules during contraction and stretch may constitute a mechanism for pumping extracellular fluid