Development of a label-free Ca²⁺ image sensor to visualize extracellular Ca²⁺ dynamics in the biological samples

Calcium ion (Ca²⁺) regulates various biological functions. Most studies, however, have focused on intracellular Ca²⁺ (Ca²⁺i) and its role in cellular responses. Extracellular Ca²⁺ ion (Ca²⁺o) also plays a crucial role in physiology serving as a key ion pool for entry into cells through various plasma membrane channels. Changes in Ca²⁺o occur under both physiological and pathological conditions and can influence biological responses. Despite its importance, Ca²⁺o has received little attention due to the lack of easy-to-use imaging devices. Here, we show the development of a label-free Ca²⁺ image sensor (CIS) for monitoring Ca²⁺o. The CIS was developed by fabricating a charge transfer type CMOS image sensor with Ca²⁺-selective membrane and is highly selective for Ca²⁺o without detecting other ions. We used CIS to monitor glutamate-induced Ca²⁺o changes in the whole hippocampal slices. CIS detected different spatiotemporal patterns of Ca²⁺o decrease across the hippocampal slice, with the largest decrease at the CA1 pyramidal cell layer. This Ca²⁺o decrease was inhibited by the NMDA receptor antagonist but not by AMPA receptor antagonist and was mimicked by NMDA suggesting that Ca²⁺ entry into neurons through NMDA receptors decreases Ca²⁺o. These results show that the CIS is a valuable tool for analyzing spatiotemporal dynamics of Ca²⁺o in normal physiology and pathological states and has tremendous potential for elucidating the biological function of Ca²⁺o.