gms | German Medical Science

Objective: Bradykinin (Bk) the active substrate of the kallikrein-kinin-system (KKS) may induce relaxation in cerebral arteries by activating B2 receptors constitutively expressed on the endothelium. Furthermore, a de novo expression of a B1 receptor may occur under pathological conditions. Here we have addressed Bk-induced vasomotor effects following brain ischemia with special respect to the underlying receptor subtypes using functional and gene expression assays.

Method: Isometric force was measured in ring segments of the middle cerebral artery (MCA) from Sprague-Dawley rats. Relaxation was elicited after precontraction induced by the thromboxane A2 mimetic U46619 (3E-7 M). Focal ischemia was induced by right-sided MCA occlusion using the intraluminal filament approach (MCAO, 2h) followed by reperfusion (22h). Gene expression was studied by means of real time semi-quantitative PCR (rt-PCR) methodology using elongation factor 1 as reference gene.

Results: In control rats Bk did not elicit any significant vasomotor effects. Following MCAO Bk induced relaxation of the previously occluded MCA with a maximum effect of 89% of precontraction. Using rt-PCR B2 receptor mRNA was present in normal MCAs throughout. After MCAO a significant upregulation of the B2 message (14 fold after 8h, 3 fold after 24h; each value p<0.05 vs. control), and a de novo expression of the B1 receptor was found. Relaxation after MCAO was not affected by the B1 receptor antagonist Lys-(Des-Arg9, Leu8)-Bk (1E-6 M), and the B1 receptor specific agonist Lys-(Des-Arg9)-Bk did not induce any relaxation. However, relaxation was largely inhibited by the selective B2 receptor antagonist, Hoe140 (1E-7 M). The same effects albeit at a somewhat lower level were observed in the contralateral MCA. Inhibiting endothelial NO synthase by Nitro-L-arginine (L-NNA, 1E-6M) decreased Bk-induced relaxation by a 50%. Charybdotoxin (CTx, 1E-8 M) a blocker of large and medium conductance Ca²⁺ dependent K⁺ channels also decreased Bk-induced relaxation by approximately 50%. Moreover, L-NNA and CTx exerted partially additive effects on Bk-induced relaxation.

Conclusions: The results suggest that focal brain ischemia results in activation of the KKS in the cerebroarterial wall indicated by a gain of vasodilator function of Bk. This effect which is also present in the contralateral MCA is mediated by the B2 receptor after upregulation of gene expression. Despite de novo expression the B1 receptor does not contribute to post-ischemic Bk-induced relaxation.

Note: Youhai Li and Natalia Lapina contributed equally.