Probing physical limits in TRPV1-magnetogenetics with micro-magnetic devices

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Ion channels are gates that control the flow of ions across the cell membrane. Activation (opening) of ion channels triggers cascades of critical signaling processes in cells and tissues. Most ion channels are activated in response to chemical or electrical stimuli, though some called mechanosensitive channels specifically respond to mechanical forces such as stretch or shear stress. "Magnetogenetics" is a method that uses magnetism to remotely and noninvasively control mechanosensitive channels to affect gene expression. This project will engineer a magnetic system to amplify and spatially target magnetogenetic responses. Magnetic nanobeads or iron-sequestering proteins will be attached to the mechanosensitive channels, and engineered magnetic fields will be applied to manipulate channel behavior. This new system of ion channel control will be used to study the response of mechanosensitive channels to a range of force. The research is expected to yield new understanding on the biophysics of mechanosensitive ion channels and inform new strategies on how to target or activate such channels remotely. The methods developed could potentially be applied to a variety of biological processes that depend on mechanical signals. The education program is focused on teaching broad skills among younger (and often underrepresented) students, encompassing design, fabrication, simulation, and biotechnology. The investigators seek to create long-term interest in STEM by introducing younger students to integrative projects that link from design and fabrication in engineering, to studying and analyzing the response of biological systems. A core goal of modern biomedical engineering remains the development of tools that can engage and control the underlying machinery of living systems. This project will develop integrated magnetic devices to spatially-control and amplify biomagnetic response in TRPV1-magnetogenetic cells. These tools will first be used to comprehensively characterize magnetogenetic Ca2+ signaling. This will be accomplished by: (1) Interfacing micro-magnetic amplifier devices to such cells (that would behave like massively-parallel, tunable magnetic traps), and (2) Varying the magnetic volume of nano-magnets tagged to TRPV1. The magnetic systems will create the highest static or time-varying magnetic field / magnetic-field gradients ever elicited in magnetogenetic systems, and will generate channel-tagged mechanical forces that exceed the theoretical force required to open TRPV1 channels. In addition, detailed studies on how static or time-varying magnetic field, field gradient, pull-rate, and nano-magnet volume interact over 2D space to polarize cellular Ca2+ signaling will help clarify (or refute) magnetogenetic behavior. The project will address emerging controversy on the viability of, and physical mechanisms behind magnetogenetics. Magnetic approaches to manipulate ion channels carry tremendous, as of yet unrealized potential to enable non-invasive tools to control biological behavior. The investigations of this project are a critically important step to both laying out the limits of magnetogenetic control and clarifying how such strategies may be practically utilized. Cells will be comprehensively probed at both the single cell and massively-parallel scales in-vitro, using modern techniques in calcium imaging, patch-clamping, and combined single-cell imaging/probing. It is anticipated that these systems will yield new insights on the physics of mechanosensitive channels under highly-localized stimuli and realize a new platform for in-vitro neuroengineering, as TRPV1 ion channels are abundant in the nervous system and play an important role in many functions, e.g., the regulation of pain. Such tools may additionally uncover new insights on the mechanotransductive nature of a wide variety of proteins due to their scalability in data collection, protein targeting, and mechanical force generation.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.