Investigation of Local Ion Transport with Potentiometric Scanning Ion Conductance Microscopy

Tuesday, October 13, 2015
West Hall 1 (Phoenix Convention Center)
L. Zhou, Y. Zhou (Indiana University-Purdue University), C. Chen (Indiana University-Purdue University), and L. A. Baker (Department of Chemistry, Indiana University)
Investigation of the fundamental properties of ion transport at nanometer scale is very important in understanding physiological processes. Scanning ion conductance microscopy (SICM) is an ideal tool in studies of ion transport in biological samples due to the high spatial resolution and the ability to simultaneously monitor local ion current and map surface topography in a non-contact manner. We have modified the conventional SICM instrument to allow local potentiometric measurements by utilizing double barrel nanopipet as the probe and by introducing additional bulk electrodes to apply transmembrane perturbations to the sample. This new technique is termed potentiometric scanning ion conductance microscopy (P-SICM). With nanoporous membranes, we have demonstrated the use of P-SICM to differentiate local conductive pathways in the sample with enhanced signal-to-noise ratio as compared to current-based measurements. Para- and transcellular pathways for ion transport in epithelial cell monolayers can be differentiated with P-SICM and local conductance value for individual cell-cell junctions can be obtained which cannot be done with any other available techniques. Moreover, we were able to capture the conductance difference in tricellular vs. bicellular cell junctions, which is useful in studying the functions of different junctional proteins that regulate ion transport at specific locations within epithelial cell layers. To achieve higher sensitivity and wider time scales of P-SICM, we have developed a new method, alternating current potentiometric scanning ion conductance microscopy (AC-PSICM). In AC-PSICM, AC transmembrane perturbations are applied across the sample while phase and amplitude of the measured local potential deflections were analyzed for a range of frequencies (5 Hz – 50 kHz). With nanoporous membranes, phase was found to be a sensitive signal in distinguishing local conductive pathways and was also used to quantify the resistance/conductance of nanopores. The sensitivity of using phase in differentiation of heterogeneous conductive pathways is dependent on frequency and the optimal frequency can be determined from Bode plot. AC-PSICM shows promise for a phase mapping technique, which can be utilized to visualize heterogeneity of conductive pathways in biological systems such as cell layers and tissues.