Insulin resistance in penile arteries from a rat model of metabolic syndrome

Abstract

Background and purpose: Metabolic and cardiovascular abnormalities accompanying metabolic syndrome, such as obesity, insulin resistance and hypertension, are all associated with endothelial dysfunction and are independent risk factors for erectile dysfunction. The purpose of the present study was to investigate the vascular effects of insulin in penile arteries and whether these effects are impaired in a rat model of insulin resistance and metabolic syndrome.

Experimental approach: Penile arteries from obese Zucker rats (OZR) and their counterpart, lean Zucker rats (LZR), were mounted on microvascular myographs and the effects of insulin were assessed in the absence and presence of endothelium and of specific inhibitors of nitric oxide (NO) synthesis, phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK). Insulin-induced changes in intracellular Ca(2+) concentration [Ca(2+)](i) were also examined. KEY RESULTS OZR exhibited mild hyperglycaemia, hypercholesterolemia, hypertryglyceridemia and hyperinsulinemia. Insulin induced endothelium- and NO-dependent relaxations in LZR that were impaired in OZR. Inhibition of PI3K reduced relaxation induced by insulin and by the beta-adrenoceptor agonist isoprenaline, mainly in arteries from LZR. Antagonism of endothelin 1 (ET-1) receptors did not alter insulin-induced relaxation in either LZR or OZR, but MAPK blockade increased the responses in OZR. Insulin decreased [Ca(2+)](i), a response impaired in OZR.

Conclusions and implications: Insulin-induced relaxation was impaired in penile arteries of OZR due to altered NO release through the PI3K pathway and unmasking of a MAPK-mediated vasoconstriction. This vascular insulin resistance is likely to contribute to the endothelial dysfunction and erectile dysfunction associated with insulin resistant states.

Figure 1

Relaxant responses to insulin were significantly impaired in penile arteries from OZR compared with LZR. (A, B) Representative traces showing the insulin-induced relaxations in penile arteries from (A) LZR and (B) OZR. (C) Average concentration–response curves for the relaxant effect of insulin in arteries from LZR and OZR. Data are shown as the mean ± SEM of 26 arteries (24 animals, LZR) and 28 arteries (25 animals, OZR). *P < 0.05, **P < 0.01 versus LZR. Vertical bars show force in mN and horizontal bars show time in min. ACh, acetylcholine; KPSS, high K⁺-physiological saline solution; LZR, lean Zucker rat; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 2

Effect of endothelial cell removal on the relaxant effects of insulin in penile arteries from LZR and OZR. (A, B) Average relaxant responses to insulin in endothelium-intact and endothelium-denuded penile arteries from (A) LZR and (B) OZR. (C, D) There was a significant linear correlation between insulin-dependent and ACh-dependent relaxations in both (C) LZR and (D) OZR. (E, F) Average ACh (10 µM) relaxant responses in endothelium-intact and endothelium-denuded penile arteries from (E) LZR and (F) OZR. Data are expressed as the mean ± SEM of 6–7 arteries (A, B, E, F) and 30 arteries (C, D), 1–2 per animal. *P < 0.05, **P < 0.01, ***P < 0.0001 versus endothelium-intact arteries. Unpaired Student's t-test. ACh, acetylcholine; LZR, lean Zucker rat; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 3

Effects of the NOS inhibitor, L-NNA (100 µM) on the relaxant responses to insulin (A, B) and (C, D) ACh (10 µM) in penile arteries from LZR and OZR. Data are shown as the means ± SEM of 6–8 arteries, 1–2 per animal. *P < 0.05, **P < 0.01, versus control arteries. Unpaired Student's t-test. ACh, acetylcholine; L-NNA, N^ω-nitro-L-arginine; LZR, lean Zucker rat; NOS, nitric oxide synthase; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 4

Effect of the PI3K inhibitor LY-294002 (3 µM) on the vasodilator responses induced by insulin (A, B) and ACh (C, D) in penile arteries from LZR and OZR. Data are shown as the means ± SEM of 7–8 arteries, 1–2 per animal. *P < 0.05, **P < 0.01, versus control arteries. Unpaired Student's t-test. ACh, acetylcholine; LZR, lean Zucker rat; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 5

The relaxant responses to the β-adrenoceptor agonist isoprenaline were partly dependent on the PI3K/Akt pathway in penile arteries from LZR but not from OZR. (A, B) Effect of the PI3K inhibitor LY-294002 (3 µM) on the vasodilator responses induced by isoprenaline in penile arteries from LZR (A) and OZR (B). (C, D) Relaxations induced by isoprenaline (C) and by the adenylate cyclase activator forskolin (D) in penile arteries from LZR and OZR. Data are shown as the means ± SEM of 10–15 arteries, 1–2 per animal. *P < 0.05, **P < 0.01, ***P < 0.0001. Unpaired Student's t-test. LZR, lean Zucker rat; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 6

Effect of the non-selective ET-1 receptor antagonist, bosentan (10 µM) on the relaxant responses to insulin (A, B) and on the contractions elicited by ET-1 (C, D) of penile arteries from LZR and OZR. Data are shown as the means ± SEM of 6–12 arteries, 1–2 per animal. ET-1, endothelin 1; KPSS, high K⁺-physiological saline solution; LZR, lean Zucker rat; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 7

Effect of the MAPK inhibitor, PD-98059 (5 µM) on the relaxant responses to insulin in penile arteries from LZR and OZR (A, B). Effect of the NOS inhibitor, L-NNA (100 µM) and the cyclooxygenase inhibitor indomethacin (1 µM) on the relaxant responses to insulin in penile arteries from LZR (C) and OZR (D) treated with PD-98059. Data are shown as the means ± SEM of 6–12 arteries, 1–2 per animal. *P < 0.05, **P < 0.01 versus control arteries. (A, B) Unpaired Student's t-test. (C, D) One-way anova. L-NNA, N^ω-nitro-L-arginine; LZR, lean Zucker rat; MAPK, mitogen-activated protein kinase; OZR, obese Zucker rat; Phe, phenylephrine.

Figure 8

Simultaneous recordings of [Ca²⁺]_i (A, C top) and tension (A, C bottom) showing the effects of a single dose of insulin (10⁻⁷ M) and ACh (10⁻⁵ M) on penile arteries from LZR (A) and OZR (C) pre-contracted with Phe (1 µM). Relaxations were accompanied by a substantial decrease of [Ca²⁺]_i. in LZR (A) and both relaxation and decrease in [Ca²⁺]_i were blunted in OZR (C,D). (B, D) Average effects of insulin and ACh on the increases in [Ca²⁺]_i and tension induced by Phe in penile arteries from LZR (B) and OZR (D). Data are shown as the means ± SEM of 5–7 arteries, 1 per animal. *P < 0.05, **P < 0.01 versus LZR. Unpaired Student's t-test. ACh, acetylcholine; KPSS, high K⁺-physiological saline solution; LZR, lean Zucker rat; OZR, obese Zucker rat; Phe, phenylephrine.