The spinal cord is the main pathway connecting brain and peripheral nervous system.
Its functionality relies on the orchestrated activity of both neurons and glial cells. To
date, most advancement in understanding the spinal cord inner mechanisms has been
made either by in vivo exposure of its dorsal surface through laminectomy or by
acute ex vivo slice preparation, likely affecting spinal cord physiology in virtue of the
necessary extensive manipulation of the spinal cord tissue. This is especially true of cells
immediately responding to alterations of the surrounding environment, such as microglia
and astrocytes, reacting within seconds or minutes and for up to several days after the
original insult. Ca2+ signaling is considered one of the most immediate, versatile, and
yet elusive cellular responses of glia. Here, we induced the cell-specific expression of
the genetically encoded Ca2+ indicator GCaMP3 to evaluate spontaneous intracellular
Ca2+ signaling in astrocytes and microglia. Ca2+ signals were then characterized in
acute ex vivo (both gray and white matter) as well as in chronic in vivo (white matter)
preparations using MSparkles, a MATLAB-based software for automatic detection and
analysis of fluorescence events. As a result, we were able to segregate distinct astroglial
and microglial Ca2+ signaling patterns along with method-specific Ca2+ signaling
alterations, which must be taken into consideration in the reliable evaluation of any result
obtained in physiological as well as pathological conditions. Our study revealed a high
degree of Ca2+ signaling diversity in glial cells of the murine spinal cord, thus adding
to the current knowledge of the astonishing glial heterogeneity and cell-specific Ca2+
dynamics in non-neuronal networks