Abstract
A battery of genetically encoded calcium indicators (GECIs) with different binding kinetics and calcium affinities was developed over the recent years to permit long-term calcium imaging. GECIs are calcium buffers and therefore, expression of GECIs may interfere with calcium homeostasis and signaling pathways important for neuronal differentiation and survival. Our objective was to investigate if the biolistically induced expression of five commonly used GECIs at two postnatal time points (days 14 and 22–25) could affect the morphological maturation of cortical neurons in organotypic slice cultures of rat visual cortex. Expression of GCaMP3 in both time windows, and of GCaMP5G and TN-XXL in the later time window impaired apical and /or basal dendrite growth of pyramidal neurons. With time, the proportion of GECI transfectants with nuclear filling increased, but an only prolonged expression of TN-XXL caused higher levels of neurodegeneration. In multipolar interneurons, only GCaMP3 evoked a transient growth delay during the early time window. GCaMP6m and GCaMP6m-XC were quite “neuron-friendly.” Since growth-impaired neurons might not have the physiological responses typical of age-matched wildtype neurons the results obtained after prolonged developmental expression of certain GECIs might need to be interpreted with caution.
Highlights
In eukaryotic cells, calcium controls virtually all fundamental processes including motility, metabolism, secretion, gene transcription, and even cell death
Employing organotypic slice cultures we aimed to investigate whether the prolongedexpression of these genetically encoded calcium indicators (GECIs) in two postnatal time windows could affect the morphological maturation of cortical pyramidal cells and multipolar interneurons
We have assessed the transfectants for symptoms of cellular degeneration in Organotypic cultures (OTCs) transfected at DIV3/4 and stained at DIV 14
Summary
Calcium controls virtually all fundamental processes including motility, metabolism, secretion, gene transcription, and even cell death. Calcium is of particular importance for the proper development and functioning of the nervous system. Crucial functions such as migration, morphofunctional development, synaptic transmission up to learning, and memory consolidation require a precise spatial and temporal regulation of calcium signals. Neural cells have evolved an intricate and complex toolkit to process calcium-related signals (for review, Brini et al, 2014). Activity causes the levels to increase up to 100-fold (for review, Berridge et al, 2000) with so-called calcium blips, puffs, and waves occurring in all compartments of a neuron. Calcium transients are induced by ionotropic and metabotropic signaling and are mediated by voltage-, ligand-, or store-operated channels at the cell membrane or in internal organelles
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