CELLULAR DISTRIBUTION OF THE RENAL BUMETANIDE‐SENSITIVE Na–K–2Cl COTRANSPORTER BSC‐1 IN THE INNER STRIPE OF THE OUTER MEDULLA DURING THE DEVELOPMENT OF HYPERTENSION IN THE SPONTANEOUSLY HYPERTENSIVE RAT

Abstract

1. The renal bumetanide-sensitive Na-K-2Cl cotransporter (BSC-1) is expressed only in the thick ascending limb and selectively traffics from intracellular vesicles (IVs) to apical plasma membranes (PMs), where BSC-1 regulates sodium reabsorption. We showed previously that in kidneys from adult spontaneously hypertensive rats (SHR; model of essential hypertension) total protein expression of BSC-1 was higher compared with kidneys from normotensive Wistar-Kyoto (WKY) rats. However, whether this change is associated with an increased trafficking of BSC-1 from IVs to PMs is unknown. The goal of the present study was to test the hypothesis that the increase in total renal BSC-1 protein expression in SHR is accompanied by an augmented distribution of BSC-1 from IVs to PMs. 2. To test the hypothesis, we obtained renal tissue from the inner stripe of the outer medulla (ISOM; enriched in thick ascending limbs) and isolated IVs and PMs from this tissue by differential centrifugation. Total BSC-1 protein expression in ISOM and BSC-1 protein expression in ISOM IVs and PMs were measured by semiquantitative western blotting in SHR and aged-matched WKY rats at different ages and stages of hypertension. 3. At 5 weeks of age, SHR were prehypertensive (mean arterial blood pressure (MABP) 97 mmHg). At this age, both the total abundance and cellular distribution of BSC-1 were similar in ISOM from SHR and WKY rats. 4. As SHR aged, their hypertension progressed (MABP 137 and 195 mmHg at 8 and 14 weeks of age, respectively). Associated with the increase in MABP was an increase in both steady state protein levels of ISOM BSC-1 and the distribution of ISOM BSC-1 to PMs (four- and sixfold increases at 8 and 14 weeks of age, respectively, compared with age-matched WKY rats; P < 0.001). 5. Using semiquantitative reverse transcription-polymerase chain reaction, BSC-1 mRNA was measured and was found not to differ between SHR and WKY rat ISOM at any age or level of MABP. 6. We conclude that as SHR transition from prehypertensive to established hypertension, there is a marked increase in the total expression of BSC-1 in ISOM that is not related to increases in steady state levels of BSC-1 mRNA and therefore unlikely to be due to changes in either the rate of BSC-1 gene transcription or the stability of BSC-1 mRNA. This suggests changes in either translational efficiency or BSC-1 protein stability in SHR. 7. We also conclude that the age/hypertension-related increase in BSC-1 protein levels in ISOM is accompanied by an equally marked increased trafficking of BSC-1 to PMs in SHR ISOM.

NKCC2 is activated in Milan hypertensive rats contributing to the maintenance of salt-sensitive hypertension

“…For instance, progression from pre-hypertensive to hypertensive state in spontaneously hypertensive rats (SHR) is accompanied by an increase in steady-state protein levels of NKCC2 and its distribution to plasma membrane compared to control rats [37,38]. Interestingly, the same amount of NKCC2 mRNA has been found in both SHR and control rats suggesting that the NKCC2 overexpression was due to a greater translational efficiency or an enhanced membrane stability of the cotransporter in SHR rats.…”

Angiotensin II and Anti Diuretic Hormone Exert Synergistic Effects on Thick Ascending Limb Transport in Spontaneously Hypertensive Rats

“…This was studied by Sonalker et al [7] who showed that Na/K/2Cl co-transporter expression is higher in outer medullas of hypertensive rats [9]. These data suggest a potential role for the Ang II/ADH interaction in the TAL in this model of hypertension, implying a concomitant enhancement of the Na/K/2Cl co-transporter membrane expression.…”

“…Although the mechanism of interaction remains the same, the additive or superimposed hormonal effects are observed only in hypertension. This could be due to the fact that V2 receptors and Ang II receptors might be upregulated [5 ]or because the genetic model we used has an enhanced tendency to increase transport [7], as has been shown by other investigators in other medullar segments of the nephron.…”

“…Indeed, during water deficit, both ADH and Ang II releases rise in an attempt to generate positive sodium balance [7]. This is important because hypertension is a condition characterized by chronic positive sodium balance.…”

“…For instance, in spontaneously hypertensive rats (SHR), there is not only increased basal collecting duct water absorption but also increased basal ENaC apical expression in connecting tubules [5,6]. Although it has been reported that SHRs have constitutive increased apical expression of the Na/K/2Cl co-transporter [7], synergic effects of hormones that regulate transport are yet to be evaluated in this model.…”