Angiotensin II-induced hypertension increases plasma membrane Na pump activity by enhancing Na entry in rat thick ascending limbs

Abstract

Thick ascending limbs (TAL) reabsorb 30% of the filtered NaCl load. Na enters the cells via apical Na-K-2Cl cotransporters and Na/H exchangers and exits via basolateral Na pumps. Chronic angiotensin II (ANG II) infusion increases net TAL Na transport and Na apical entry; however, little is known about its effects on the basolateral Na pump. We hypothesized that in rat TALs Na pump activity is enhanced by ANG II-infusion, a model of ANG II-induced hypertension. Rats were infused with 200 ng·kg(-1)·min(-1) ANG II or vehicle for 7 days, and TAL suspensions were obtained. We studied plasma membrane Na pump activity by measuring changes in 1) intracellular Na (Nai) induced by ouabain; and 2) ouabain-sensitive oxygen consumption (QO2). We found that the ouabain-sensitive rise in Nai in TALs from ANG II-infused rats was 12.8 ± 0.4 arbitrary fluorescent units (AFU)·mg(-1)·min(-1) compared with only 9.9 ± 1.1 AFU·mg(-1)·min(-1) in controls (P < 0.024). Ouabain-sensitive oxygen consumption was 17 ± 5% (P < 0.043) greater in tubules from ANG II-treated than vehicle rats. ANG II infusion did not alter total Na pump expression, the number of Na pumps in the plasma membrane, or the affinity for Na. When furosemide (1.1 mg·kg(-1)·day(-1)) was coinfused with ANG II, no increase in plasma membrane Na pump activity was observed. We concluded that in ANG II-induced hypertension Na pump activity is increased in the plasma membrane of TALs and that this increase is caused by the chronically enhanced Na entry occurring in this model.

Dietary Fructose Enhances the Ability of Low Concentrations of Angiotensin II to Stimulate Proximal Tubule Na+ Reabsorption

“…This explanation is supported by the literature in which stimulation of proximal tubules with Ang II triggers transient increases in Na i of up to 8 mmol/L [ 32 , 55 ]. Such an increase in Na i would be expected to double the resting levels of about 10 mmol/L [ 32 ], and elevate Na/K-ATPase activity near to its maximum velocity [ 49 ].…”

“…Oxygen Consumption: QO 2 was measured using methods similar to those we reported [ 49 ]. Briefly, 2 to 4 mg of protein from proximal tubule suspensions were taken to a final volume of 6 mL in the chamber of a YSI Model 5301B bath assembly (Yellow Springs Instruments, Yellow Springs, OH, USA).…”

Dietary Fructose Increases the Sensitivity of Proximal Tubules to Angiotensin II in Rats Fed High-Salt Diets

“…Oxygen consumption (QO 2 ): changes in QO 2 by proximal tubules are a surrogate for changes in net transport rates because this segment produces ATP via aerobic metabolism, and the majority of this ATP (60–70%) is used to drive Na + across the basolateral membrane by the Na + /K + -ATPase. QO 2 was measured using methods similar to those we reported [ 38 , 54 ]. Briefly, 2 to 4 mg of protein suspended in bicarbonate-buffered physiological saline were taken to a final volume of 6 mL in the chamber of a YSI Model 5301B bath assembly (Yellow Springs Instruments, Yellow Springs, OH, USA).…”

TRPV4 activation mediates flow-induced nitric oxide production in the rat thick ascending limb

“…Three days after the adenovirus injections were performed medullary thick ascending limb suspensions of the left (TRPV4shRNA-transduced) and the right (nontransduced) kidney from the same rat were obtained by collagenase digestion as we previously described (20,54,62). Suspensions were lysed in a buffer containing 20 mmol/l HEPES (pH 7.4), 2 mmol/l EDTA, 0.3 mmol/l sucrose, 1.0% NP-40, 0.1% sodium dodecyl sulfate, 5 g/ml antipain, 10 g/ml aprotinin, 5 g/ml leupeptin, 4 mmol/l benzamidine, 5 g/ml chymostatin, 5 g/ml pepstatin A, and 0.105 mmol/l pf-block.…”