Abstract
Currently, cellular action potentials are detected using either electrical recordings or exogenous fluorescent probes that sense the calcium concentration or transmembrane voltage. Ca imaging has a low temporal resolution, while voltage indicators are vulnerable to phototoxicity, photobleaching, and heating. Here, we report full-field interferometric imaging of individual action potentials by detecting movement across the entire cell membrane. Using spike-triggered averaging of movies synchronized with electrical recordings, we demonstrate deformations up to 3 nm (0.9 mrad) during the action potential in spiking HEK-293 cells, with a rise time of 4 ms. The time course of the optically recorded spikes matches the electrical waveforms. Since the shot noise limit of the camera (~2 mrad/pix) precludes detection of the action potential in a single frame, for all-optical spike detection, images are acquired at 50 kHz, and 50 frames are binned into 1 ms steps to achieve a sensitivity of 0.3 mrad in a single pixel. Using a self-reinforcing sensitivity enhancement algorithm based on iteratively expanding the region of interest for spatial averaging, individual spikes can be detected by matching the previously extracted template of the action potential with the optical recording. This allows all-optical full-field imaging of the propagating action potentials without exogeneous labels or electrodes.
Highlights
Modern methods for detecting electrical activity in cells rely on either electrical or optical recordings, both of which are invasive
One proposed explanation was a change in the refractive index associated with ions flowing into the cell during the action potential or cell swelling due to water molecules accompanying those ions[24,29]
The number of Na+ ions entering the cell during the depolarization phase of an action potential is N = CmAΔVm/e, where ΔVm ∼ 100 mV is the transmembrane potential rise, Cm is the specific membrane capacitance (~0.5 μF cm−2, ref. 15), A is the cell membrane surface area, and e is the elementary charge
Summary
Modern methods for detecting electrical activity in cells rely on either electrical or optical recordings, both of which are invasive. Optical measurements rely on exogenous fluorescent probes, such as calcium indicators[6] or transmembrane voltage sensors[7,8,9]. The large changes in transmembrane voltage that take place during action potentials have long been hypothesized to induce changes in the shape of biological cells, which is primarily determined by the balance of intracellular hydrostatic pressure, membrane tension, and strain exerted by the cytoskeleton[12,13,14]. 15), which increases the force exerted on the membrane of a 10 μm cell by 0.3 nN. This is expected to deform the cell by decreasing its surface area while preserving volume, thereby making it more spherical. The movement of the cell membrane, called electromotility[15,18], is expected to follow the voltage change nearly instantaneously, since the magnitude of this force is very significant at the cellular scale: if it were not counteracted by the cytoskeleton, such a force would accelerate the cell by ~600 m/s2—much more than what is
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