Fig. 4.

Example of using the 2-photon laser as a micromanipulator. (a–d) Intravital MPM images of a Ren1d-GCaMP5 mouse glomerulus before (a) and after laser-induced stimulation (b–d) of a single parietal epithelial cell (PEC, crosshair). Renin lineage cells co-express tdTomato (red) and the fluorescent calcium indicator GCaMP5G (cyan), which indicates real-time elevations of intracellular calcium [Ca²⁺] by increased GCaMP5 fluorescence. Focusing high laser power on a single PEC cell induced a robust and sustained elevation of PEC [Ca²⁺] at the site of injury, which promoted a cell-to-cell propagating PEC [Ca²⁺] wave along the Bowman’s capsule, suggesting that PEC cells are functionally coupled. (e) GCaMP5G fluorescence intensity of regions of interest (ROI) 1–3 (b) over time. (f) Maximal GCaMP5G/tdTomato fluorescence intensity ratios of ROI 1–3 before and after laser-induced PEC cell stimulation. (g, h) Intravital MPM images of a wild-type mouse glomerulus. Plasma is stained after iv injection of Alexa 680-conjugated albumin (gray). Red blood cells are excluded from the staining and appear as dark bands in the lumen of the renal capillaries. Laser-induced stimulation of a glomerular capillary loop induces thrombus formation and severe alterations of glomerular hemodynamics (arrows). EA efferent arteriole, AA afferent arteriole, Bar = 20 μm in each panel