Retinal plasma extravasation in streptozotocin-diabetic rats mediated by kinin B(1) and B(2) receptors

Abstract

Background and purpose: We investigated whether or not kinin receptors play a role in diabetic blood-retinal barrier breakdown, which is a leading cause of vision loss.

Experimental approach: Blood-retinal barrier breakdown was quantified using Evans blue, and expression of kinin B(1) receptor mRNA was measured using quantitative reverse transcrition-PCR. Diabetic rats (streptozotocin (STZ), 65 mg kg(-1)) received a single intraocular injection of bradykinin (BK) or des-Arg(9)-BK, alone, or in combination with antagonists for B(1) (des-Arg(10)-Hoe140, R-715) and/or B(2) (Hoe140) receptors, given intraocularly or intravenously (i.v.).

Key results: In control rats, BK (0.1-10 nmol) dose-dependently increased plasma extravasation, which was inhibited by Hoe140 (0.2 nmol), whereas des-Arg(9)-BK (0.1 and 1 nmol) was without effect. B(1) receptor mRNA was markedly increased in retinas of diabetic rats, and this was prevented by N-acetyl-L-cysteine (1 g kg(-1) day(-1) for 7 days). Plasma extravasation in retinas of STZ-diabetic rats was higher than in controls and enhanced by des-Arg(9)-BK. Response to des-Arg(9)-BK was inhibited by intraocular or i.v. injection of B(1) receptor antagonists. Diabetes-induced plasma extravasation was inhibited only by a combination of des-Arg(10)-Hoe140 and Hoe 140 (100 nmol kg(-1), i.v. 15 min earlier) or by R-715 (1 micromol kg(-1), i.v.) injected daily for 7 days.

Conclusions and implications: Kinin B(1) receptors are upregulated in retinas of STZ-diabetic rats through a mechanism involving oxidative stress. Both kinin B(1) and B(2) receptors contribute to increased plasma extravasation in diabetic retinopathy. Chronic inhibition of both kinin receptors, possibly with antioxidant adjuvants, may be a novel therapeutic strategy for diabetic retinopathy.

Figure 1

Bradykinin (BK) induces plasma extravasation in the retinas of healthy rats. (a) Dose–response effect of BK or des-Arg⁹-BK on plasma extravasation after intravitreous injection. (b) B₂ receptors mediate BK-induced retinal plasma extravasation. The intravitreous injection of Hoe140 was followed 10 min later by the intravitreous injection of BK. Responses are expressed as optical density measured at 620 nm wavelength (n=4 or 5). Statistical significance are set at P<0.05, where ^* shows differences between rats treated with and without agonist and + shows differences between rats treated with and without antagonist.

Figure 2

Des-Arg⁹-BK induces retinal plasma extravasation in streptozotocin (STZ)-diabetic rats. (a) Plasma extravasation was quantified in the retina of STZ-diabetic rat and STZ-nondiabetic rat, and after an intravitreous injection of increasing doses of des-Arg⁹-BK in STZ-diabetic rats. (b) Retinal plasma extravasation was quantified after an intravitreous injection of des-Arg¹⁰-Hoe140 followed 10 min later by the intravitreous injection of des-Arg⁹-BK. (c) Retinal plasma extravasation was quantified in intravitreous-injected des-Arg⁹-BK STZ-diabetic rat before and after an i.v. injection of des-Arg¹⁰-Hoe140 or R-715. Responses are expressed as optical density measured at 620 nm (n=3–5). Statistical significance are set at P<0.05, where † shows differences between rats with and without STZ-diabetes, ^* shows differences between diabetic rats treated with and without agonist and + shows differences between diabetic rats treated with and without antagonist.

Figure 3

Effect of treatment with N-acetyl-L-cysteine (NAC) for 7 days on B₁ kinin receptor mRNA overexpression induced by streptozotocin (STZ) in the retina. Control rats received the vehicle only. NAC (1 g kg⁻¹ day⁻¹) was given by gavage 3 h after STZ and daily until the end of the experiment. Data are means±s.e.m. of 5–7 rats per group. Statistical comparison with controls (^*) and STZ-diabetic rats (+) is indicated at P<0.05.

Figure 4

Kinin receptors induce diabetes-evoked retinal plasma extravasation. (a) Retinal plasma extravasation was quantified in streptozotocin (STZ)-diabetic rat retina before and after an intravitreous injection of Hoe140 or des-Arg¹⁰-Hoe140. (b and c) Retinal plasma extravasation was quantified in STZ-diabetic rat retina before and after i.v. injection of Hoe140 (b) or des-Arg¹⁰-Hoe140 or R-715 (c). (d) Retinal plasma extravasation was quantified in STZ-diabetic rat retina before and after i.v. injection of a combination of Hoe140 and des-Arg¹⁰-Hoe140. Responses are expressed as optical density measured at 620 nm (n=3–5). Statistical significance are set at P<0.05, where † shows differences between STZ-diabetic rats versus healthy control rats, + shows differences between rats treated with and without antagonist.