Potassium-SelectiveNanoelectrode Arrays for Single-CellProfiling of Human iPSC-Derived Cardiomyocytes

Figures

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  1. Overview of the KINESIS platform for potassium ion-selective sensing and drug screening. (a) Schematic illustrating the operating principle of KINESIS. Drug-induced modulation of ion channels alters extracellular K+ concentrations at the cell–electrode interface. The released K+ ions are selectively detected by a valinomycin-based potassium-selective membrane (KSM) coated on platinum-black-functionalized nanopillar electrodes. (i) Inhibition of K+ efflux results in reduced extracellular K+ levels, whereas (ii) enhanced K+ release increases local extracellular K+ concentration. (b) Application of KINESIS for drug screening. Cardiomyocytes cultured on the nanopillar array are exposed to drug candidates, and drug-induced changes in extracellular K+ concentration are measured as shifts in electrode potential.

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  2. Characterization of the nanopillar electrode array (NEA) platform. (a) Photograph of the fabricated NEA device. (b) Scanning electron microscopy (SEM) images of the nanopillar array at increasing magnification showing (i) the electrode well, (ii) the nanopillar array, and (iii) an individual nanopillar. Insets show schematic representations of the corresponding structures. (c) SEM image of the cell–nanoelectrode interface, demonstrating close contact between a cardiomyocyte and the nanopillars. (d) Percent change in electrode impedance at 1 kHz measured over time in culture (Days on Device, DOD 4, 6, 10, and 13) relative to bare electrodes. (e) Representative extracellular recordings obtained before electroporation. (f) Representative intracellular recordings obtained after electroporation, following the formation of transient membrane pores in the plasma membrane (PM) that enable intracellular access.

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  3. Development and analytical characterization of the KINESIS platform. (a) Schematic illustration of the nanopillar functionalization workflow, including electrodeposition of an intermediate conductive layer followed by potassium-selective membrane (KSM) coating. (b) Cross-sectional schematic of the fully functionalized K+-selective nanopillar electrode. (c) Optical image showing KSM coverage on the device surface. (d, e) Potentiometric calibration curves of K+-selective nanopillar electrodes incorporating PANI:1.5DBSA (d) or PtB (e) as the intermediate transducer layer. Both configurations exhibit near-Nernstian K+ responses and minimal interference from Na+. (f) Long-term stability assessment of a representative electrode before and after repeated biological exposure, showing preservation of K+ sensing performance following cardiomyocyte culture and device reuse.

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  4. Validation of KINESIS for monitoring drug-induced potassium dynamics in human iPSC-derived cardiomyocytes. (a) Photograph of a functionalized KINESIS device. (b) LIVE/DEAD fluorescence images of cardiomyocytes cultured on the device, demonstrating cell viability and monolayer formation. (c) Quantification of cell viability on unmodified and KINESIS-functionalized nanoelectrode arrays, showing no significant effect of device modification on cell viability (n = 3 devices per condition). (d, g) Schematic illustrations of the proposed mechanisms underlying drug-induced potassium flux. Caffeine stimulates intracellular Ca2+ release and promotes K+ efflux (d), whereas ouabain inhibits Na+/K+-ATPase activity and alters membrane-proximal K+ dynamics (g). (e, h) Representative potentiometric recordings showing KINESIS responses to caffeine (e) and ouabain (h) treatment. (f, i) Quantification of drug-induced potential shifts measured across independent recording trials, demonstrating significant differences between control and drug-treated conditions. Together, these results establish that KINESIS enables label-free, selective detection of pharmacologically induced potassium dynamics at…