A new view of macula densa cell microanatomy

Macula densa (MD) cells are well known as chief regulators of renal and glomerular hemodynamics and renin secretion. However, their new, non‐traditional function as master regulators of renal tissue remodeling is emerging. The present study aimed to explore the microanatomical basis and regulation of the secretory phenotype of MD cells by directly visualizing the morphological features of living MD cells in male and female mice on low, normal, and high salt diet. A new mouse model (MD‐GFP mice) was generated with inducible and conditional MD‐specific, membrane‐targeted expression of the fluorescence reporter EGFP by crossing nNOS‐CreERT2 and mTmG‐floxed mice. MD cells were freshly isolated from digested kidneys of MD‐GFP mice using FACS sorting, and high resolution time‐lapse multiphoton imaging was performed to visualize single, living MD cells. Also, suboptimal tamoxifen induction was performed in MD‐GFP mice, which resulted in the membrane‐targeted expression of EGFP only in a single MD cell per MD plaque and allowed high resolution imaging of the MD ultrastructure on fixed histological sections, including both the plasma and intracellular membranes. MD cells featured very long (~10 μm) single primary cilia at the apical membrane, and a dense network of thin cell processes (up to 20 μm long) at the base of cells projecting into the extraglomerular mesangium. The basal cell processes were more detailed in female mice and in response to low salt diet. Time‐lapse imaging of living MD cells visualized a high number of membrane vesicles trafficking within these cell processes from the cell center to the periphery. These results are consistent with the highly regulated paracrine signaling function and secretory phenotype of MD cells, and suggest the high rate of synthesis and release of novel, yet to be identified secreted MD proteins and their actions on target cell types at the glomerular entrance. Support or Funding Information NIH R01 DK64324