Label-free real-time imaging of extracellular Ca~(2+) uptake in the hippocampal slice using Ca-PVC membrane based on charge-transfer-type potentiometric sensor arrays

摘要

An elevation of intracellular Ca~(2+) concentration during neuronal excitation is widely known, and much research has already been done. However, the changes in "extracellular" Ca~(2+) concentration ([Ca~(2+)]o) have received only limited attention, and its imaging and spatiotemporal analysis has not been achieved. Thus, we develop a [Ca~(2+)]o imaging sensor to clarify the role(s) of [Ca~(2+)]o in regulation of physiological and pathophysiological neurons. 23.55-μm-pitch and 128×128 pixels Ca~(2+) image sensor using plasticized polyvinyl chloride (PVC) as an ion sensitive membrane was fabricated and the diffusion image of Ca~(2+) concentration in sample solution was successfully obtained. The sensitivity was 23.3 mV/decade. The mouse hippocampal slice was placed on the sensor, and stimulated with glutamate and its related agonists. When stimulated, the output image was clearly changed, i.e., glutamate decreased [Ca~(2+)]o via NMDA receptor. We also found that such [Ca~(2+)]o changes were obvious in CA3 and dentate gyrus (DG) of the hippocampus, and were not observed in the slice-free sensor area. Taken together, we succeeded in the development of the [Ca~(2+)]o imaging device, allowing us to monitor and analyze real-time imaging of the spatiotemporal changes in [Ca~(2+)]o in the brain slices.