In vivo confocal neuroimaging (ICON): non‐invasive,functional imaging of the mammalian CNS with cellular resolution

With in vivo confocal neuroimaging (ICON), single retinal ganglion cells (RGCs) can be visualized non‐invasively, repeatedly, in real‐time and under natural conditions. Here we report the use of ICON to visualize dynamic changes in RGC morphology, connectivity and functional activation using calcium markers, and to visualize nanoparticle transport across the blood–retina barrier by fluorescent dyes. To document the versatility of ICON, we studied the cellular response to optic nerve injury, and found evidence of reversible soma swelling, recovery of retrograde axonal transport and a difference in calcium activation dynamics between surviving and dying RGCs. This establishes ICON as a unique tool for studying CNS physiology and pathophysiology in real‐time on a cellular level. ICON has potential applications in different research fields, such as neuroprotection/regeneration, degeneration, pharmacology, toxicity and drug delivery.