Patrick K. K. Leong

Mechanisms of Pressure Natriuresis

Abstract: An acute increase in blood pressure provokes a rapid decrease in proximal tubule salt and water reabsorption that is central to tubuloglomerular feedback regulation of renal blood flow and glomerular filtration rate and contributes to pressure natriuresis. The molecular mechanisms responsible for this critical homeostatic adjustment were studied. When blood pressure is acutely elevated, apical proximal tubule NHE3 are rapidly redistributed out of the microvilli to intermicrovillar clefts and then endosomal pools, and Na,K‐ATPase activity is suppressed. Depressing apical Na+ entry without hypertension is not sufficient to decrease Na,K‐ATPase activity, and depressing Na,K‐ATPase activity alone is not sufficient to decrease proximal tubule Na+ and water reabsorption; thus, it appears that coordinated decreases in both NHE3 surface distribution and Na,K‐ATPase activity may be important for the response to acute hypertension. Clamping plasma angiotensin II levels blunts the retraction of NHE3 from the cell surface to endosomal pools. The increased volume flow of salt and water to the loop of Henle stimulates Na,K‐ATPase activity in this region and provides evidence for a downstream shift in sodium transport during acute hypertension. These same responses in the proximal tubule and loop develop and persist in the spontaneously hypertensive rat. These studies demonstrate that sodium transporters along the nephron are very dynamic, responding quickly to normal fluctuations of blood pressure, and are key to generating the macula densa tubuloglomerular feedback signal and for accommodating increased volume flow through the loop of Henle.

Angiotensin II stimulates trafficking of NHE3, NaPi2, and associated proteins into the proximal tubule microvilli

Angiotensin II (ANG II) stimulates proximal tubule (PT) sodium and water reabsorption. We showed that treating rats acutely with the angiotensin-converting enzyme inhibitor captopril decreases PT salt and water reabsorption and provokes rapid redistribution of the Na(+)/H(+) exchanger isoform 3 (NHE3), Na(+)/Pi cotransporter 2 (NaPi2), and associated proteins out of the microvilli. The aim of the present study was to determine whether acute ANG II infusion increases the abundance of PT NHE3, NaPi2, and associated proteins in the microvilli available for reabsorbing NaCl. Male Sprague-Dawley rats were infused with a dose of captopril (12 microg/min for 20 min) that increased PT flow rate approximately 20% with no change in blood pressure (BP) or glomerular filtration rate (GFR). When ANG II (20 ng x kg(-1) x min(-1) for 20 min) was added to the captopril infusate, PT volume flow rate returned to baseline without changing BP or GFR. After captopril, NHE3 was localized to the base of the microvilli and NaPi2 to subapical cytoplasmic vesicles; after 20 min ANG II, both NHE3 and NaPi2 redistributed into the microvilli, assayed by confocal microscopy and density gradient fractionation. Additional PT proteins that redistributed into low-density microvilli-enriched membranes in response to ANG II included myosin VI, DPPIV, NHERF-1, ezrin, megalin, vacuolar H(+)-ATPase, aminopeptidase N, and clathrin. In summary, in response to 20 min ANG II in the absence of a change in BP or GFR, multiple proteins traffic into the PT brush-border microvilli where they likely contribute to the rapid increase in PT salt and water reabsorption.

Redistribution of Myosin VI from Top to Base of Proximal Tubule Microvilli during Acute Hypertension

During acute hypertension, Na(+)/H(+) exchangers (NHE3) retract from top to base of proximal tubule microvilli (MV) and Na(+) reabsorption decreases in proximal tubule. This study aimed to determine whether the actin-based motor myosin VI coordinately retracts with NHE3 in response to acute hypertension. BP was raised approximately 50 mmHg in rats for 20 to 30 min or sham treated, and kidneys were analyzed by subcellular fractionation or microscopy. During acute hypertension, myosin VI redistributed from low density apical MV-enriched membranes (from 23 +/- 2.4 to 11.4 +/- 2.2%) into higher density membranes (from 23.2 +/- 0.7 to 36.9 +/- 2.6%). By confocal microscopy, myosin VI was detected over the whole length of the MV in controls, then became completely focused at the base of MV during acute hypertension. For electron microscopic analysis using immunogold labeling, MV were divided into five zones from top (z1) to base (z5). In controls, myosin VI was evenly distributed through the five MV zones. In acute hypertension, myosin VI decreased in z1 (from 20.6 +/- 1.9 to 10.5 +/- 2.3%) and z2 (from 21.0 +/- 2.0 to 13.2 +/- 1.4%) and increased in z5 (from 21.1 +/- 3.3 to 38.6 +/- 3.0%). These results provide the first observation that acute hypertension causes myosin VI redistribution and support the idea that myosin VI may serve as the molecular motor for NHE3 retraction from top to base of MV during acute hypertension.