Na⁺/H⁺ exchange activity and NHE-3 expression in renal tubules from the spontaneously hypertensive rat

Background. The NHE-3 isoform of the Na⁺/H⁺ antiporter, in the apical membrane of renal proximal tubule, is responsible for the bulk transport of Na⁺ and fluid reabsorption. Studies have reported that apical NHE-3 translocates to internal pools, thereby facilitating natriuresis when blood pressure increases abruptly. Methods. The present study examined Na⁺/H⁺ exchange activity and NHE-3 expression in renal cortical tubules from the spontaneously hypertensive rat (SHR) and WKY rats before and after the development of hypertension. SHR 4 to 6 weeks of age were pre-hypertensive, 6 to 7 weeks old had mild hypertension, and 8 to 13 weeks old had severe hypertension. Renal proximal tubules (PTs) were isolated and purified by Percoll gradient centrifugation. NHE-3 protein and mRNA levels were determined by Western and Northern blots, respectively. Apical brush border membrane vesicles (BBMV) were prepared using the MgSO₄ aggregation method and Na⁺/H⁺ exchange activity assessed using the acridine orange method. Results. Na⁺/H⁺ exchange activity, determined as the rate of Na⁺-dependent intracellular pH (pH_i) recovery assessed using BCECF after an acute acid load, was significantly greater in PTs from SHR than in WKY rats at all age groups (4 to 6 weeks, 0.30 ± 0.04 vs. 0.24 ± 0.02 pH U/30 sec, P < 0.05; 6 to 7 weeks, 0.42 ± 0.07 vs. 0.29 ± 0.05 pH U/30 sec, P < 0.05; and 8 to 13 weeks, 0.48 ± 0.07 vs. 0.40 ± 0.07 pH U/30 sec, P < 0.05). The Na⁺-dependent recovery in BBMV was also greater in SHR than WKY rats (1464 ± 62 vs. 1042 ± 79 fluorescence. U/5 sec, P < 0.001) and was unaffected by cariporide, a specific NHE-1 inhibitor. NHE-3 protein levels also were significantly higher in SHR than age-matched WKY rats at all stages during the development of hypertension (pre-hypertensive 1.8-fold; early onset hypertension twofold; established hypertension 1.5-fold; each P < 0.05). By contrast, NHE-3 mRNA levels were not different between SHR and WKY rats at each age group. Conclusions. Na⁺/H⁺ exchange activity and NHE-3 protein abundance in renal proximal tubules from the SHR are increased while NHE-3 mRNA is not. A post-transcriptional event(s) best explains the increase in NHE-3 protein expression since mRNA levels were not increased. The alterations in the SHR antedate the development of hypertension and fail to decrease as blood pressure increases with age in the SHR, which likely results in inappropriate renal sodium retention in the face of a chronic rise in blood pressure.