A CALCIUM IMAGING PIPELINE TO DETECT AND QUANTIFY COMPOUND-SPECIFIC EFFECTS IN HUMAN AND MOUSE ASTROCYTES

Astrocytes are crucial mediators of diverse aspects of brain function, including energy metabolism and synapse formation and maturation. They form a vast network in the brain and signal both with each other and with neurons. Calcium is the primary information carrier in astrocytes and can be measured using fluorescent indicators. However, it is unclear how calcium signals differ between mouse astrocytes, human astrocytes derived from forward programming, and astrocytes from brain organoids. Here we compare and characterise astrocyte
calcium dynamics in mouse astrocytes, human astrocytes derived from two iPSC- lines, and human astrocytes from dissociated cortical brain organoids. Astrocytic calcium signals were measured using GFAP-driven GCaMP6f in mouse astrocytes and the calcium dye Cal520-AM human astrocyte cultures. We demonstrate reliable activation using the purinergic receptor agonist ATP, as well as decreased activity following treatment with the ER calcium pump blocker CPA across all culture models. Furthermore, we identify similarities and differences in event-based characteristics such as amplitude, area, and duration. We then tested serotonergic receptor function using lysergic acid (LSD) and observed increased calcium activity. Furthermore, human recombinant tau
aggregates, an in vitro model of Alzheimer´s disease pathology, decreased activity in human astrocytes. We show that properties of calcium signals are sensitive to species and model system. Despite limited knowledge regarding the interpretation of astrocyte calcium data, this pipeline provides valuable insight about cell health and
compound-specific effects that could support drug candidate screening in the future.