Surface Plasmon Resonance for Measuring Exocytosis from Populations of PC12 Cells: Mechanisms of Signal Formation and Assessment of Analytical Capabilities

Publication year: 2017
Authors: Moreira B. 1, Tuoriniemi J. 1, Kouchak Pour N. 1, Mihalčíková L.1, Safina G. 1,2
Affiliations:

1 - Department of Chemistry and Molecular Biology, University of Gothenburg, Kemigården 4, 412 96 Gothenburg, Sweden
2 - Division of Biological Physics, Department of Physics, Chalmers University of Technology, Kemigården 1, 412 96 Gothenburg, Sweden

Published in: Analytical Chemistry, 2017, Vol. 89 (5), p. 3069–3077
doi: 10.1021/acs.analchem.6b04811

Because of cell to cell variation, it is difficult to obtain statistically significant data on the frequency of exocytosis events (Rexocytosis, t–1 m–2) with traditional single cell electrophysiological or fluorescence microscopy based methods. Here we take the first steps toward a rapid cost-effective surface plasmon resonance (SPR) based method for measuring the Rexocytosis for populations of PC12 cells. The conditions for culturing confluent monolayers on the sensor slides were optimized, and neurotransmitter exocytosis was evoked by injecting solutions with elevated [K+]. Exocytosis caused a shift of the resonance angle (Δθr) that was linearly proportional to Rexocytosis. The Δθr was mainly due to elevated concentration of secretory vesicles close to the cell membrane. The increased vesicle concentration thus acted as a proxy for the Rexocytosis that could not be measured directly. The Δθr was calibrated for Rexocytosis using single cell amperometry on parallel cell cultures. The cell populations were large enough for variation in responses between sensor slides to only reflect actual differences in biological condition. The applicability for drug screening is demonstrated by studying the effects of EGTA, reserpine, and prolonged stimulation by K+.


MP-SPR keywords: 37C, average response from ∼ 800 cells, cell monolayer growth on mouse collagen IV functionalized gold sensors, drug discovery, interaction of reserpine (enzyme inhibitor), K+ evoked catecholamine exocytosis, living cells, microscopy, neurotransmitters exocytosis, PC12, quantitative data