Fig 1.
Schematic diagram illustrating the flowchart of the experimental protocol. After the left common carotid artery (CCA) was ligating for 60 min, the rats in the HI30, 40, 50, and 60 groups were exposed to PaO2 30–39 mmHg, 40–49 mmHg, 50–59 mmHg, and 60–69 mmHg, which was maintained at O2 concentrations of 11, 13, 15, or 18% of the gas mixture, respectively. In the HI+ hypercapnia groups, a humidified gas mixture of 8% CO2 containing the above O2 concentrations was used to maintain PaCO2 at 60–80 mmHg.
Table 1.
MAP and pH of rats exposed to hypercapnic cerebral hypoxia-ischemia.
Table 2.
PaO2 and PaCO2 of rats exposed to hypercapnic cerebral hypoxia-ischemia.
Fig 2.
The time-course of changes of cerebral blood flow (CBF).
A, B. Changes of CBF in the ipsilateral hemisphere during HI (A) and HI+ hypercapnia (B). C-F. CBF in the S, HI30, and HI30+ hypercapnia groups (C); S, HI40, and HI40+ hypercapnia groups (D); S, HI50, and HI50+ hypercapnia groups (E); and S, HI60, and HI60+ hypercapnia groups (F). CBF was measured before and after common carotid artery ligation. All CBF values are expressed as a percentage of pre-ligation (100%) in each animal. n = 5 for each group. *p < 0.05 compared to baseline; #p < 0.05 vs the S group.
Fig 3.
Changes in brain vascular permeability in the cortex and brain edema.
(A): The magnitude of blood–brain barrier disruption was quantified by measuring the extent of leakage of 10 kDa dextran. Values are expressed as a percentage of the values in the S group and presented as mean ± SD. (B): Brain water content (wet-to-dry [W/D] weight ratio) in the ipsilateral hemisphere. Clear bars are shams (S group), black bars are hypoxia (HI) and striped bars are hypoxia with hypercapnia (HI+hypercapnia). *p < 0.05 vs the S group; #p < 0.05 vs the HI group (n = 5).
Fig 4.
Effects of hypercapnia on the expression of aquaporin-4 (AQP4) in the ischemic cortex of rats in the S group.
(A) Western blot analyses of AQP4 expression and (B) densities of AQP4 expression relative to GAPDH in the sham and the HI30, HI40, HI50, and HI60 groups (B) alone or with hypercapnia. Clear bars are shams (S group), black bars are hypoxia (HI) and striped bars are hypoxia with hypercapnia (HI+hypercapnia). Data are expressed as mean ± SD. (n = 3). *p < 0.05 vs. Sham, #p < 0.05 vs. HI groups only. (C) Immunofluorescence labeling of AQP4 in the ischemic cerebral cortex. Magnification, ×20. Scale bar = 100 μm.
Fig 5.
Neuropathological examination.
(A) Nissl staining showed more neuronal damage in ipsilateral ischemic cortex of rats in the HI30 and HI40 groups with or without hypercapnia. Hypercapnia treatment afforded neuro-protection against hypoxic-ischemia (PaO2 >50) mmHg at 3h after hypoxic-ventilation. Sham animals had no damage. Scale bars = 1000 μm, and = 100 μm in the insets. (B) TUNEL staining revealed that more TUNEL-(+) cells were observed in the HI 30 and HI 40 with hypercapnia groups compared to the HI along groups, but fewer in the HI 50 and HI 60 with hypercapnia groups compared to the HI along groups. Scale bars = 100 μm. (C) Quantification of TUNEL-positive cells in the hippocampal CA1 region. Clear bars are hypoxia (HI), black bars are hypoxia with hypercapnia (HI+hypercapnia) and none bars are shams (S group). *p < 0.05 vs. Sham, #p < 0.05 vs. HI groups only, Фp < 0.05 vs. HI 40+ hypercapnia group (n = 5).