Measuring Calcium Responses in Cells

Cytosolic calcium responses were indicated as the increase in green fluorescence intensity. The raw images (Tagged Image File Format) were exported and analyzed using image analysis software (ImageJ, National Institutes of Health, Bethesda, MD, USA ImageJ.nih.gov/ij/">https://ImageJ.nih.gov/ij/ accessed on 2 May 2022) [27 (link),28 (link)] and LIF (Leica Microsystems GmbH). An experimenter traced the activated cells manually to determine cell size and fluorescence intensity. Fluorescence intensity at the baseline level was taken as F₀. Evoked calcium response was expressed as the ratio of the post-treatment fluorescence intensity (F) to the basal level, as shown in our previous studies [10 (link),25 (link)]. Activation in neurons was defined as an increase in F/F₀ ≥ 1.5 fold.