Dynamics of astrocytes Ca signaling: a low-cost fluorescence customized system for 2D cultures.

In an effort to help reduce the costs of fluorescence microscopy and expand the use of this valuable technique, we developed a low-cost platform capable of visualising and analysing the spatio-temporal dynamics of intracellular Ca signalling in astrocytes. The created platform, consisting of a specially adapted fluorescence microscope and a data analysis procedure performed with Imagej Fiji software and custom scripts, allowed us to detect relative changes of intracellular Ca ions in astrocytes. To demonstrate the usefulness of the workflow, we applied the methodology to several astrocyte preparations, specifically immortalised human astrocyte cells and wild-type mouse cells. To demonstrate the reliability of the procedure, analyses were conducted by stimulating astrocyte activity with the agonist dihydroxyphenylglycine (DHPG), alone or in the presence of the antagonist 2-methyl-6-phenylethyl-pyridine (MPEP).