Soluble guanylate cyclase alpha1beta1 limits stroke size and attenuates neurological injury

Abstract

Background and purpose: Nitric oxide mediates endothelium-dependent vasodilation, modulates cerebral blood flow, and determines stroke outcome. Nitric oxide signals in part by stimulating soluble guanylate cyclase (sGC) to synthesize cGMP. To study the role of sGC in stroke injury, we compared the outcome of cerebral ischemia and reperfusion in mice deficient in the alpha(1) subunit of sGC (sGCalpha(1)(-/-)) with that in wild-type mice.

Methods: Blood pressure, cerebrovascular anatomy, and vasoreactivity of pressurized carotid arteries were compared in both mouse genotypes. Cerebral blood flow was measured before and during middle cerebral artery occlusion and reperfusion. We then assessed neurological deficit and infarct volume after 1 hour of occlusion and 23 hours of reperfusion and after 24 hours of occlusion.

Results: Blood pressure and cerebrovascular anatomy were similar between genotypes. We found that vasodilation of carotid arteries in response to acetylcholine or sodium nitroprusside was diminished in sGCalpha(1)(-/-) compared with wild-type mice. Cerebral blood flow deficits did not differ between the genotypes during occlusion, but during reperfusion, cerebral blood flow was 45% less in sGCalpha(1)(-/-) mice. Infarct volumes and neurological deficits were similar after 24 hours of occlusion in both genotypes. After 1 hour of ischemia and 23 hours of reperfusion, infarct volumes were 2-fold larger and neurological deficits were worse in sGCalpha(1)(-/-) than in the wild-type mice.

Conclusions: sGCalpha(1) deficiency impairs vascular reactivity to nitric oxide and is associated with incomplete reperfusion, larger infarct size, and worse neurological damage, suggesting that cGMP generated by sGCalpha(1)beta(1) is protective in ischemic stroke.

Figure 1

Infarct size in wild-type (WT) and sGCα₁-deficient mice (sGCα₁^−/−) subjected to the filament model of MCA occlusion for 1 hour, followed by 23 hours of reperfusion (Transient, n=9 and 6 for WT and sGCα₁^−/− mice, respectively) or to MCA occlusion for 24 hours (Permanent, n=10 and 12 for WT and sGCα₁^−/− mice, respectively). *P<0.01 and ** P<0.001 vs WT transient by two-way ANOVA with Bonferroni post-hoc testing.

Figure 2

Neurologic scores in wild-type (WT, ●) and sGCα₁-deficient mice (sGCα₁^−/−, ○) subjected to MCA occlusion for 1 hour, followed by 23 hours of reperfusion (Transient) or to MCA occlusion for 24 hours (Permanent). N=10 and 10 for WT, and 6 and 12 for sGCα₁^−/− mice in the transient and permanent groups, respectively. *P=0.003 vs WT transient by the Kruskal-Wallis test.

Figure 3

Effect of sGCα₁-deficiency on CBF deficit imaged by laser speckle flowmetry. (A) Representative images in wild-type (WT) and sGCα₁-deficient mice (sGCα₁^−/−) 10 min after distal MCAO. Superimposed colored pixels indicate regions with ≤20% (light blue) or 21–30% (dark blue) residual CBF. Imaging field dimensions are 6 × 8 mm. (B) Composite graph showing the area of cortex (mm²) with ≤20% (black) or 21–30% (gray) residual CBF 10 min after distal MCAO, compared with preischemic baseline in WT and sGCα₁^−/− mice. N=9 and 10 for WT and sGCα₁^−/− mice, respectively. P=NS between WT and sGCα₁^−/− mice.

Figure 4

Effect of sGCα₁-deficiency on CBF, measured by LDF, during 1 hour of middle cerebral artery (MCA) occlusion, 30 minutes of MCA reperfusion and 30 minutes of carotid artery reperfusion. N=10 and 9 for wild-type (WT) and sGCα₁-deficient mice (sGCα₁^−/−), respectively. Data are shown as mean ± SEM. *P<0.05 between WT and sGCα₁^−/− mice.

Figure 5

Relaxation effect of SNP (A) and ACh (B) on phenylephrine precontracted carotid arteries from wild-type (WT, n=12) and sGCα₁-deficient mice (sGCα₁^−/−, n=6). *P<0.001 for the effect of genotype on vasodilation.