LUMIN: an automated graphical analysis toolbox for high-throughput calcium imaging ofin vitro neuronal cultures

Human pluripotent stem cell-derived in vitro models, combined with advances in high-content live-cell calcium imaging, provide a powerful platform for disease modeling and drug screening. However, most calcium imaging analysis tools have been developed for in vivo applications with suboptimal features for processing data from cultured neurons. Here, we present LUMIN ( L ive-cell U ser M odule for I maging and analysis of N euronal activity), a scalable and broadly applicable software that integrates a graphical user interface with automated data analysis encompassing single-cell segmentation, signal extraction, and activity quantification. Two analysis modules for transiently active or quiescent stimuli-evoked neurons make it broadly applicable for cultures with distinct functional properties. Its efficient data processing and linear time complexity enable processing thousands of cells within hours (~ 7000 cells from ~ 70 recordings per 30 min) on a standard laptop. We demonstrate LUMIN’s performance on human stem cell-derived ventral midbrain neurons through two applications: (1) modulation of spontaneous activity with well-characterized pharmacological compounds, and (2) evoking responses in quiescent neurons upon compound stimulation.