Abstract 077: In Vivo Evidence of AT1a Receptor-Mediated Uptake of Angiotensin II by the Proximal Tubule Visualized by Intravital Multiphoton Imaging
The development of all forms of angiotensin II (ANG II)-dependent hypertension is associated with higher levels of intrarenal ANG II, which are greater than can be explained on the basis of circulating ANG II and suppressed cortical renin expression. In the present study, we used intravital multiphoton imaging to test the hypothesis that AT1 (AT1a) receptor-mediated uptake of ANG II by the proximal tubules of the kidney plays a major role in the underlying mechanisms. Adult male Munich-Wistar rats were anesthetized with Inactin and continuously infused with a pressor dose of Alexa 488-conjugated ANG II (50 ng/min, i.v.) for 2 hr. Time-dependent proximal tubular uptake responses of Alexa 488-ANG II were studied with mean arterial blood pressure maintained at ~150 mmHg throughout the experiment. After 30 min infusion, very low levels of Alexa 488-ANG II were visualized within proximal tubules and cortical collecting ducts (CCDs) (p<0.05). However, high levels of Alexa 488-ANG II were accumulated in the lumen of CCDs, but not that of proximal tubules. After 1 hr infusion, moderate levels of Alexa 488-ANG II were visualized in the proximal tubules, but not in the glomeruli and CCDs. The most striking uptake responses were visualized in all segments of proximal tubules after 2 hr infusion. Internalized Alexa 488-ANG II was predominantly localized to the basolateral side, where it was colocalized with tetramethyl rhodamine methyl ester (TMRM), a mitochondrial membrane potential-dependent dye. TMRM is a lipophilic cationic dye that is primarily accumulated in the mitochondria of proximal tubules. Some internalized Alexa 488-ANG II was visualized around and over the nuclei. Furthermore, the uptake of Alexa 488-ANG II by proximal tubules was significantly attenuated in caveolin 1-knockout mice (p<0.01), and blocked in AT1a receptor-knockout mice (p<0.01). Our results provide strong intravital multiphoton microscopic evidence that circulating ANG II is primarily taken up by the proximal tubules of the kidney via an AT1a receptor-mediated mechanism, and that internalized ANG II may be transported to the mitochondria and the nucleus, where it may alter mitochondrial and nuclear function in the proximal tubules of the kidney.
Author Disclosures: X.C. Li: None. R.M. Sandoval: None. B.A. Molitoris: None. J.L. Zhuo: None.
- © 2015 by American Heart Association, Inc.