Enalapril Normalizes Endothelium-Derived Hyperpolarizing Factor-Mediated Relaxation in Mesenteric Artery of Adult Hypertensive Rats Prenatally Exposed to Testosterone

Abstract

Prenatal exposure to elevated testosterone levels induces adult life hypertension associated with selective impairments in endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation in mesenteric arteries. We tested whether the angiotensin-converting enzyme inhibitor enalapril restores EDHF function through regulating the activities of small (Kcnn3) and intermediate (Kcnn4) conductance calcium-activated potassium channels in mesenteric arteries. Pregnant Sprague-Dawley rats were injected subcutaneously with vehicle or testosterone propionate (0.5 mg/kg/day from Gestation Day 15 to 19), and their 6-mo-old adult male offspring were examined. A subset of rats in these two groups was given enalapril (40 mg/kg/day) for 2 wk through drinking water. Blood pressures were assessed through carotid arterial catheter and endothelium-dependent mesenteric arterial EDHF relaxation, using wire myography. Ace and Kcnn3 and Kcnn4 channel expression levels were also examined. Renal and vascular Ace expression and plasma angiotensin II levels were increased in testosterone offspring. Blood pressure levels were significantly higher in testosterone offspring than in controls, and treatment with enalapril significantly attenuated blood pressure in testosterone offspring. EDHF relaxation in testosterone offspring was reduced compared to that in controls, and it was significantly restored by enalapril treatment. Kcnn4 channel expression and function were similar between control and testosterone rats, but it was not affected by enalapril treatment. Relaxation mediated by Kcnn3 was impaired in testosterone offspring, and it was normalized by enalapril treatment. Furthermore, enalapril treatment restored expression levels of Kcnn3 channels. These findings suggest that enalapril has a positive influence on endothelial function with improvement in EDHF relaxation through normalization of Kcnn3 expression and activity.

Keywords: EDHF; Kcnn3 channel; blood pressure; enalapril; endothelium; mesenteric arteries; pregnancy; testosterone.

FIG. 1

Ace mRNA expression in kidney and mesenteric arteries of control and T rats. Real-time RT-PCR was used to assess vascular Ace mRNA expression. Quantitation of Ace was normalized relative to that of β-actin levels. Values are means  ±  SEM of 5 rats in each group. *P ≤ 0.05 versus control.

FIG. 2

Effect of ACE inhibition on hypertension. ACE inhibitor (enalapril, 0.25 g/L through drinking water) was administered for 2 wk to 6-mo-old offspring, and changes in MAP were measured using a carotid catheter. Values are means ± SEM of 6–7 animals in each group of animals.*P < 0.05 versus control (C); ^#P < 0.05 versus T counterpart.

FIG. 3

Effect of enalapril treatment on endothelium-dependent EDHF-mediated vascular relaxation in mesenteric arteries. Arterial rings were isolated from control (C) and T rats with and without enalapril treatment, contracted with PE, and examined for relaxation to ACh in the presence of the NOS inhibitor L-NAME (10⁻⁴ M) and with the PGI₂ blocker indomethacin (10⁻⁵ M). Values are means ± SEM (n = 6 rats, 2 vessel segments/rat).

FIG. 4

Effect of the Kcnn3 channel inhibitor apamin on EDHF-mediated vascular relaxation in mesenteric arteries from control (A) and T (B) rats. ACE inhibitor enalapril was administered for 2 wk to control (C) and T (D) rats, and the changes in EDHF-mediated vascular relaxation were determined. Arterial rings were contracted with PE and examined for relaxation to ACh in the presence of apamin (10⁻⁷ M), the NOS inhibitor L-NAME (10⁻⁴M), and the PGI₂ blocker indomethacin (10⁻⁵M). Values are means ± SEM (n = 6–7 rats, 2 vessel segments/rat). C, control.

FIG. 5

Effect of the Kcnn4 channel inhibitor TRAM-34 on EDHF-mediated vascular relaxation in mesenteric arteries from control (A) and T (B) rats. ACE inhibitor enalapril was administered for 2 wk to control (C) and T (D) rats, and the changes in EDHF-mediated vascular relaxation were determined. Arterial rings were contracted with PE and examined for relaxation to ACh in the presence of TRAM-34 (10⁻⁶ M), the NOS inhibitor L-NAME (10⁻⁴M), and the PGI₂ blocker indomethacin (10⁻⁵M). Values are means ± SEM (n = 6–7 rats, 2 vessel segments/rat). C, control.

FIG. 6

Effect of enalapril treatment on endothelium-independent EDHF relaxation. Arterial rings were isolated from control (C) and T rats with and without enalapril treatment, contracted with PE, and examined for relaxation to cumulative additions of levcromakalim. Values are means ± SEM (n = 5 rats from each group, 2 vessel segments/rat).

FIG. 7

Effect of enalapril treatment on Kcnn3 and Kcnn4 channel mRNA expression in mesenteric arteries of control (C) and T rats with and without enalapril treatment. Real-time RT-PCR was used to assess vascular Kcnn3 and Kcnn4 mRNA expression. Quantitation of vascular Kcnn3 and Kcnn4 components was normalized relative to β-actin levels. Values are means  ±  SEM of five rats in each group. *P ≤ 0.05 versus control. ^#P < 0.05 versus T counterpart.