Upregulated Renal Mineralocorticoid Receptor and 11β-Hydroxysteroid Dehydrogenase Type 2 Are Associated with K+ Retention During MR Antagonism in Female Spontaneously Hypertensive Rats

Figures

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  Figure 1. Spironolactone increases serum potassium in female but not male SHRs. Summary graphs showing serum K+, Na+, and Cl− levels in male (a,c,e) and female (b,d,f) SHRs receiving vehicle (M, F) or spironolactone (MS, FS). Data were analyzed using repeated measures two-way ANOVA with Geisser–Greenhouse correction and Holm–Sidak post hoc test. *—denotes p < 0.05 for pairwise comparisons. Each group included n ≥ 6 rats.

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  Figure 2. Sex-specific effects of spironolactone on urinary electrolyte excretion. Summary graphs showing urinary K+ (UKV), Na+ (UNaV) and Cl− (UClV) excretion in male (a,c,e) and female (b,d,f) SHRs receiving vehicle (M, F) or spironolactone (MS, FS). Data were analyzed using repeated measures two-way ANOVA with Geisser–Greenhouse correction and Holm–Sidak post hoc test. *—denotes p < 0.05 for pairwise comparisons. Each group included n = 6 rats.

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  Figure 3. Spironolactone elevates circulating aldosterone in female SHRs, while PRA and serum corticosterone levels are lower in females compared to males. Summary graphs showing endpoint serum aldosterone concentrations (a), PRA (b) and serum corticosterone levels (c) in male and female SHRs. Data were analyzed using two-way ANOVA with the Holm–Sidak post hoc test. *—denotes p < 0.05 for pairwise comparisons. Each group included n ≥ 7 rats.

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  Figure 4. Elevated renal MR and 11β-HSD2 in female vs. male SHRs. (a,b) Relative renal expression of Nr3c2 mRNA, encoding MR (a), and Hsd11b2 mRNA, encoding 11β-HSD2 protein (b), in SHRs treated with vehicle (M, F) or spironolactone (MS, FS) normalized to Gapdh and expressed relative to the levels in vehicle-treated male SHRs. (c,d) Representative western blots showing the MR-reporting band at ~107 kDa (c), the 11β-HSD2-reporting band at ~40 kDa (d), and total protein loading (Ponceau red) in kidney homogenates from SHRs. (e,f) Quantification of relative renal MR (e) and 11β-HSD2 (f) abundance normalized to the total protein signal visualized using Ponceau S staining. Data were analyzed using two-way ANOVA with Holm–Sidak post hoc test. n ≥ 6 per group.

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  Figure 5. Spironolactone does not attenuate hypertension in SHRs. Mean arterial pressure (MAP) averages recorded by tail-cuff plethysmography at baseline, day 7, day 14 and day 21 in male (a) and female (b) SHRs treated with vehicle, M, F, or 30 mg·kg−1·day−1 spironolactone, MS, FS (n = 8 per group). Data were analyzed using repeated measures two-way ANOVA with the Geisser-Greenhouse correction. n = 8 per group.

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  Figure 6. ENaC activity and amiloride-sensitive urinary electrolyte excretion are not affected by spironolactone administration in SHRs. (a) Representative western blot showing the α-ENaC-reporting band and total protein loading (Ponceau red) in kidney homogenates from male and female SHRs treated with vehicle (M, F) or spironolactone (MS, FS). (b) Quantification of the relative renal α-ENaC abundance in SHRs. (c) Total ENaC activity (fNPo) measured by patch-clamp in isolated cortical collecting duct segments. (d–f) Endpoint amiloride-sensitive urinary excretion of sodium, UNaV (d), potassium, UKV (e), and chloride, UClV (f), normalized to body weight. Data were analyzed using two-way ANOVA. n ≥ 6 rats per group.

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  Figure 7. Experimental design and timeline.