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Accessory Subunits in the Regulation and Dysregulation of Myocardial Sodium Channels

Mentor: Jeanne M. Nerbonne, PhD, Alumni Endowed Professor of Molecular Biology and Pharmacology, Departments of Medicine, Cardiovascular Division, and Developmental Biology

Lab description: A major goal of the research program in the Nerbonne lab is to define the physiological mechanisms that control the expression, properties, and functioning of myocardial Na+ (Nav) and K+ (Kv) channels, which are key determinants of myocardial membrane excitability, affecting the waveforms of action potentials in individual myocytes and the propagation of electrical activity through the heart. In addition, we are exploring the pathophysiological mechanisms contributing to the dysregulation of Nav and Kv channels in inherited and acquired cardiovascular diseases. A major focus of ongoing research in the lab is identifying the components of native myocardial Nav channels and defining the mechanisms involved in the regulation and modulation of these channels by accessory proteins. The following research project is presently accepting interested undergraduates to work under the direct oversight of a graduate student or postdoctoral fellow and with the guidance and mentorship of Dr. Nerbonne.

Project: Mechanisms linking voltage-gated sodium (Nav) channel accessory subunits to the regulation and dysregulation of membrane excitability in the mammalian heart: Considerable evidence suggests that native cardiac Nav channels function in macromolecular complexes of the Nav1.5 α subunit and multiple accessory proteins. The Nav1.5 α subunit, which forms the Na+ selective channel pore, has four homologous domains (DI-DIV), each with six transmembrane segments (S1-S6), connected by cytoplasmic linkers. The S1-S4 segments form the channel voltage-sensing domains, and the S5-S6 segments form the ion-selective pore. On depolarization, the voltage-sensing domains move and coupled to the pore through intracellular S4-S5 linkers, cause channel opening. Hydrophobic amino acids in the DIII-DIV linker, the IFM motif, are essential for inactivation. Native Nav channel function, however, is determined not only by the α subunit but also by the associations with intracellular and transmembrane accessory subunits. In collaboration with the laboratory of Jon Silva, Associate Professor of Biomedical Engineering, ongoing studies are detailing the molecular mechanisms underlying the regulation of myocardial Nav1.5-encoded channels by intracellular fibroblast growth factor 12 (iFGF12), the predominant iFGF expressed in the human heart. We are also testing the hypothesis that the iFGF12A variant, which we have found is upregulated in failing human hearts, has functional effects on the biophysical and pharmacological properties of myocardial Nav1.5-encoded channels distinct from the effects of iFGF12B. An additional goal is to test the hypothesis that another intracellular Nav channel auxiliary subunit, calmodulin (CaM), which binds to the C-terminus of Nav1.5 very near to the site of iFGF12 binding, modulates the effects of iFGF12B and/or iFGF12A on Nav1.5 channel gating. We are combining cellular electrophysiological and molecular genetic approaches to accomplish these goals. We are also developing a mathematical model of the Nav currents in native (mouse and human) myocytes and will use these in dynamic clamp experiments to manipulate the properties or the amplitudes of the Nav currents computationally in situ and in real-time during current-clamp recordings. In addition, biochemical and mass spectrometry-based proteomic approaches are being employed to identify the molecular components of native (mouse and human) myocardial Nav channel macromolecular protein complexes.

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Biomimetic Hydrogel Models of Early Development of the Placental Vasculature

Mentor: Michelle Oyen, Associate Professor, Department of Biomedical Engineering

Lab description: Our lab is developing engineering tools to understand and intervene into preterm birth, with a focus on the placenta and the maternal-fetal interface. The maternal and fetal vasculatures do not directly mix but the transport distances for oxygen and nutrients between them are small. One major cause of preterm birth is pre-eclampsia, when the placenta causes potentially fatally high blood pressure to develop in the mother. We use both experimental tools and computational modeling to try and understand the placenta’s role in this and related conditions.

Project: Both maternal and fetal outcomes in pregnancy are reliant on the first trimester development of the placenta, the maternal-fetal interface. Fetal trophoblast cells invade the maternal uterine lining and direct the remodeling of uterine spiral arteries. The signaling cues that direct this invasion are not fully known, nor are there diagnostic criteria to indicate that this process has been suboptimal. Poor outcomes in pregnancy, including pre-eclampsia and fetal growth restriction, occur when the placental vascular bed is insufficient for transport. Clinical signals of poor maternal and fetal outcomes may not appear until the third trimester, long after the early placental development has ceased. To study this process, we are developing assays using biomimetic hydrogels as an artificial extracellular matrix to model the physical and chemical environment during trophoblast invasion. The work is being conducted in collaboration with scientists in developmental biology, who derive trophoblast-like cells from induced pluripotent stem cells. The long-term goal of the project is to develop biomarker assays for signaling either poor or adequate trophoblast invasion, to allow for early diagnosis of future potential placental malfunction.

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Cardiac OEF or PAD/diabetes study

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Determining the biomechanics of perivascular tissues using an image-based system for in-situ arterial pressurization

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Drug development for Cantu Syndrome

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How sensitive hearts are to stimulation with ultrasound

Mentor: Christian Zemlin, Associate Professor of Surgery, Division of Cardiothoracic Surgery, Director of the Cardiothoracic Surgery Research Lab

The Zemlin laboratory studies cardiac arrhythmia mechanisms, particularly the mechanism of atrial fibrillation. We use both optical mapping with voltage-sensitive fluorescent probes and extracellular electrodes to monitor cardiac electrical activity. We are also interested in the surgical treatment of cardiac arrhythmias, and we are developing a new ablation modality for cardiac tissue based on electroporation with nanosecond pulsed electric fields.

Projects: One of our current projects that would be interesting for undergraduate students to participate in is sonogenetics. In this project, we study rat hearts that have been transfected with mechanosensitive ion channels, in particular, how sensitive these hearts are to stimulation with ultrasound. We have an existing optical mapping set up to monitor the electrical activity of rat hearts, which needs to be extended in several ways: 1) We need to extend our setup to allow for the placement and manipulation of different ultrasound probes; 2) We need to perform optical mapping experiments in which we use the ultrasound probes to stimulate the genetically modified hearts; 3) the movies that are recorded during or stimulation attempts need to be processed and evaluated to determine the efficacy of ultrasound stimulation. All these tasks offer great opportunities for undergraduate student participation.

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Identifying biological signals that change between sleep and wakefulness

Mentor: Yao Chen, Assistant Professor of Neuroscience, Cell Biology and Physiology, and Biomedical Engineering

Lab description: Neuromodulators such as dopamine, acetylcholine, and neuropeptides have profound effects on neural circuits and behavior. Altered neuromodulation is associated with most psychiatric disorders, major neurodegenerative disorders, and neuromodulatory systems are targets of almost all drugs of abuse. While specific behaviors have been linked to specific neuromodulators, and while many neuromodulator receptors and their downstream signaling pathways are known, how neuromodulators regulate behavior remains enigmatic.

The knowledge gap exists because our understanding of molecular signaling networks remains largely a static diagram of connections between molecules. Our laboratory attempts to fill the gap between molecular neuroscience and animal behavior by elucidating the spatial and temporal dynamics of biological signals, because their dynamics carry critical information that explain subsequent modifications of cells, circuits, and behavior.

Specifically, we aim to understand how the dynamics of neuromodulators and intracellular signals contribute to the function of neuromodulators, to learning, and to the function of sleep. We combine biosensor development, two photon fluorescence lifetime imaging microscopy, electrophysiology, as well as molecular, cellular, and biochemical approaches in mice to visualize molecular dynamics in action, perturb them with precise spatiotemporal control, and analyze the functional consequences. Our research promises to uncover principles of neuromodulator action, illuminate how molecular mechanisms produce behaviorally relevant features, and ultimately help treat psychiatric disorders.

Project: Despite the importance of sleep, it is unclear how sleep promotes many important functions (e.g. cardiovascular health, emotions, learning, immune functions). We will use fluorescence lifetime photometry combined with EEG/EMG analysis to identify biological signals that change between sleep and wakefulness. They will immediate generate hypotheses on the cellular functions of sleep. The student interested in this project needs to be comfortable handling live mice, detail oriented, and not afraid of new technology/software. The student will receive extensive mentoring and support.

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Imaging functional recovery after stroke

Mentor: Adam Bauer, Assistant Professor of Radiology, Assistant Professor of Biomedical Engineering

Lab description: Two major themes running through our lab’s research are mapping functional brain organization in the mouse and examining how changes in local neural activity are related to corresponding changes in blood flow. We are very interested in mapping functional network connectivity in the mouse brain, and how those connections evolve in healthy mice and models of disease, in particular stroke. To help us answer our questions, we use the latest advances mouse genetics and optogenetic targeting strategies.


1) Mapping brain activity and behavior in mice following focal ischemia.
2) Using deep learning to characterize functional vs. compensatory recovery after stroke.

For either project, the student would be involved in experimental design, data collection (e.g. functional neuroimaging) and image analysis. Depending on interests, students would also be able to take on a larger role in hardware development and data processing/analysis.

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Mapping cerebral hemodynamics during pediatric critical care

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Methodology for culturing organotypic cardiac slices from atria and ventricles from explanted human donor hearts rejected from transplantation

Mentor: Stacey Rentschler, M.D., Ph.D., Associate Professor, Department of Medicine, Cardiovascular Division

Research in the Rentschler lab is centered around three broad themes: defining the developmental and molecular mechanisms underlying congenital and acquired arrhythmias, defining the transcriptional and epigenetic mechanisms that regulate the programming and reprogramming of cellular electrophysiological phenotypes, and harnessing the power of developmental signaling cascades to treat conduction disorders. Specifically, we focus on the transcriptional and epigenetic pathways activated in response to Notch and Wnt signaling during development and in response to cardiac injury, which mediate changes in ion channel gene expression and cellular electrophysiology. Our group previously demonstrated that reactivation of developmental signaling pathways including Notch and Wnt could electrically remodel cardiomyocytes or “reprogram” them to adopt a new electrical phenotype in animal models. In heart failure, reactivation of the Notch signaling pathway may contribute to transcriptional changes influencing ion channel gene expression that may predispose to the development of lethal arrhythmias. In contrast, reactivation of distinct signaling networks within the adult myocardium may provide new avenues for regenerative medicine approaches for the treatment of arrhythmias, such as in the development of a biological pacemaker.

Projects: Current projects in the Rentschler lab include the development of a methodology for culturing organotypic cardiac slices from atria and ventricles from explanted human donor hearts rejected from transplantation. Functional readouts include optical mapping with voltage-sensitive dyes to visualize and quantify electrophysiological properties, as well as microelectrode recordings to assess changes in cellular physiology. In addition, we are performing genome-wide gene expression and epigenetic studies on mouse and human cardiac tissue to decipher the complex regulatory mechanisms that govern the expression of important regulators of electrophysiology within regions of the heart in both health and disease states. This methodology will allow us to test the response of human cardiac tissue to selected therapeutics, with the goal of expediting the prolonged process involved in getting new therapeutics validated for use in humans. More recently, Dr. Rentschler has built a team-science approach and led a multidisciplinary group of pathologists, cancer biologists, radiation oncologists, and cardiac electrophysiologists at Washington University to understanding mechanisms of cardiac irradiation to treat ventricular tachycardia first observed in the ENCORE-VT clinical trial.

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Predicting whether patients will respond to anti-arrhythmic drug therapies

Mentor: Jonathan Silva, Associate Professor, Department of Biomedical Engineering

Lab description: We want to develop precision approaches to arrhythmia therapy by identifying biophysical features of ion channels that predict patient drug response.

Project: We have developed an automated protocol to assess anti-arrhythmic drug response in patients with different genetics. Students involved with this summer project would help with the collection of this data and working on predictive methods to connect it to patient response.

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Reagents to increase elastic fiber assembly and prevent elastic fiber degradation

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Role of lysosomes in homeostasis and disease

Mentor: Abhinav Diwan, MD, Professor, Department of Medicine, Cardiovascular Division

Lab description: The Diwan lab is investigating the regulation of lysosome function in physiology and disease. Their program has discovered evidence for acquired lysosome dysfunction in cardiac myocytes and macrophages in the setting of myocardial infarction and cardiomyopathies and in protein aggregate induced cardiomyopathy and heart failure. His lab employs molecular, genetic, and surgical modeling techniques in animal models to investigate lysosome biology in cardiac stress and homeostasis using basic research approaches.

Projects: Specific research projects in the Diwan lab that will offer interdisciplinary training opportunities for undergraduate students are as follows. Project 1: The first project focuses on elucidating the role of specific genes (n=6) uncovered in a genetic screen of starvation survival in lysosome biogenesis transcription factor-deficient worms in regulating the starvation response in worms and mammalian cells. Students will test the role of individuals genes in mutant worm lines and cells targeted for loss of function of these genetic pathways to evaluate their role in starvation stress. Project 2: will focus on uncovering the redundancy of three members of the lysosome biogenesis transcription factor family, namely TFEB, TFE3, and Mitf, in genetically targeted murine embryonic fibroblasts. These data will be correlated with murine models with cardiac myocyte-specific deficiency of these transcription factors to understand their role in cardiac homeostasis. Project 3: is to evaluate the role of genetic mutations and stress-induced post-translational modifications in CRYAB, a cardiac enriched chaperone protein, in causing aberrant phase separation and protein aggregation, leading to proteotoxic cardiomyopathy. The undergraduates will work with targeted phase separation assays, as well as generic tool compounds to disrupt phase separation to assess the mechanisms whereby CRYAB becomes aggregate-prone. Project 4: A fourth project is focused on zinc-induced lysosomal remodeling in worms and mammalian cells. This is based on preliminary observations that toxic metals are sequestered within lysosomes to attenuate cellular toxicity. Students will evaluate worm models of gain and loss of function of the lysosome biogenesis program and quantify metals in the lysosomal compartment using state-of-the-art organelle isolation strategies and mass spectroscopy-based chemical assays.

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Small animal ECG device for cardiac gated micro-CT

Mentor: Tiezhi Zhang, Associate Professor of Radiation Oncology

Lab description: We are developing novel x-ray imaging technologies for image guided radiotherapy and diagnostic imaging. Cardiac ablation is a hot topic in radiotherapy. We are developing micro-CT with cardiac gating for small animal studies.

Project: Micro-CT is of importance for preclinical research. We are developing a novel inverse-geometry micro-CT that can improve the radiation dose efficiency and super-high resolution. However, cardiac motion causes artifacts in the images. We will need a device to monitor the cardiac cycles of the mice for cardiac gated micro-CT imaging. The summer fellow will work togher with the PI and graduate students to develop an ECG device for mice and interface with micro-CT system.

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Trojan vs Hand Drivers: Effectiveness in Optogenetic Pacing of Drosophila

Mentor: Chao Zhou, Associate Professor, Department of Biomedical Engineering

Lab description: We are interested in developing novel optical imaging technologies for biomedical applications, especially in developing optical coherence tomography (OCT) and microscopy (OCM) technologies to perform “optical biopsy” and generate 3D in situ images of tissue morphology, function and pathological status in real-time without the need to remove and process specimens.

We are also interested in applying these technologies to a variety of biological and clinical applications, including cancer research, neuroscience, developmental biology and tissue engineering.

Project: In order to effectively place a gene of interest into a tissue of interest, a driver gene must be used. In optogenetics, this gene of interest encodes for an opsin that, such as channelrhodopsin or halorhodopsin, and in our lab, we design crosses so that these opsins are specific to the heart region, utilizing the GAL4-UAS system. Previous work in our group has shown that the Hand driver is effective at producing flies with heart specific opsins that are able to respond to heart pacing experiments. As we explore ways to further optimize our cardiac optogenetic pacing experiments, we would like to explore more drivers, comparing their effectiveness at delivering the gene of interest to the heart for optogenetic pacing experiments, both through looking at tissue specificity, using tools such as multiphoton imaging and fluorescent markers, and by comparing the results of pacing experiments at different laser power levels.

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Use of Human Pluripotent Stem Cell-derived Macrophages to Improve Tissue Integration

Mentor: Kory Lavine, Assistant Professor of Internal Medicine

Lab description: Our laboratory is interested in understanding how the immune system can be manipulated to reduce inflammation, improve tissue healing, and leveraged to develop regenerative approaches.

Project: This project will explore the functions of human pluripotent stem cells-derived macrophages within the mouse heart. Key areas of interest will include how transplanted macrophages influence the organization, morphology, and transcriptional signatures of native endothelial cells, fibroblasts, and cardiomyocytes.

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Using image processing to study physiology in iPSC-derived micro-heart muscle arrays

Mentor: Nathaniel Huebsch, Associate Professor, Department of Biomedical Engineering

Lab description: We use induced pluripotent stem cells, micro-fabrication and image processing technologies to study how mechanical stress combines with genetics to trigger the pathophysiology and molecular signatures of genetically inherited cardiomyopathy.

Project: The student will work with Huebsch lab team members to form arrays of cardiac micro-tissues derived from human induced pluripotent stem cells, perform high speed video microscopy of the tissues and use Matlab coding routines to convert these videos into information on tissue-level contractility, voltage and calcium handling. He/she will also assist in structural and biochemical analysis of the tissues and gain proficiency in these techniques.