Using 2-Photon Microscopy to Quantify the Effects of Chronic Unilateral Ureteral Obstruction on Glomerular Processes

Abstract

Applying novel microscopy methods to suitable animal disease models to explore the dynamic physiology of the kidney remains a challenge. Rats with surface glomeruli provide a unique opportunity to investigate physiological and pathophysiological processes using intravital 2-photon microscopy. Quantification of glomerular capillary blood flow and vasoconstriction and dilatation in response to drugs, permeability, and inflammation are just some of the processes that can be studied. In addition, transgenic rats, i.e., podocytes labeled with fluorescent dyes and other molecular biomarker approaches, provide increased resolution to directly monitor and quantify protein-protein interactions and the effects of specific molecular alterations. In mice, which lack surface glomeruli after four weeks of age, unilateral ureteral obstruction (UUO) for several weeks has been used to induce surface glomeruli. As this induction model does not allow for baseline studies, we quantified the effects of UUO on glomerular processes in the UUO model in Munich Wistar Frömter (MWF) rats, which have surface glomeruli under physiologic conditions. The UUO model for five weeks or more induced significant alterations to gross renal morphology, the peritubular and glomerular microvasculature, as well as the structure and function of tubular epithelia. Glomerular and peritubular red blood cell (RBC) flow decreased significantly (p < 0.01), probably due to the significant increase in the adherence of white blood cells (WBCs) within glomerular and peritubular capillaries. The glomerular sieving coefficient of albumin increased from 0.015 ± 0.002 in untreated MWFs to 0.045 ± 0.05 in 5-week-old UUO MWF rats. Twelve weeks of UUO resulted in further increases in surface glomerular density and glomerular sieving coefficient (GSC) for albumin. Fluorescent albumin filtered across the glomeruli was not reabsorbed by the proximal tubules. These data suggest that using UUO to induce surface glomeruli limits the ability to study and interpret normal glomerular processes and disease alterations.