Transient lack of glucose but not O2 is involved in ischemic postconditioning-induced neuroprotection

Abstract

Aim: Cerebral ischemic postconditioning has emerged recently as a kind of endogenous strategy for neuroprotection. We set out to test whether hypoxia or glucose deprivation (GD) would substitute for ischemia in postconditioning.

Methods: Adult male C57BL/6J mice were treated with postconditioning evoked by ischemia (bilateral common carotid arteries occlusion) or hypoxia (8% O(2) ) after 45-min middle cerebral arterial occlusion. Corticostriatal slices from mice were subjected to 1-min oxygen-glucose deprivation (OGD), GD, or oxygen deprivation (OD) postconditioning at 5 min after 15-min OGD.

Results: Hypoxic postconditioning did not decrease infarct volume or improve neurologic function at 24 h after reperfusion, while ischemic postconditioning did. Similarly, OGD and GD but not OD postconditioning attenuated the OGD/reperfusion-induced injury in corticostriatal slices. The effective duration of low-glucose (1 mmol/L) postconditioning was longer than that of OGD postconditioning. Moreover, OGD and GD but not OD postconditioning reversed the changes of glutamate, GABA, glutamate transporter-1 protein expression, and glutamine synthetase activity induced by OGD/reperfusion.

Conclusions: These results suggest that the transient lack of glucose but not oxygen plays a key role in ischemic postconditioning-induced neuroprotection, at least partly by regulating glutamate metabolism. Low-glucose postconditioning might be a clinically safe and feasible therapeutic approach against cerebral ischemia/reperfusion injury.

Figure 1

Ischemic postconditioning model in vivo and in vitro. (A) In control group, adult male mice were subjected to 45‐min middle cerebral artery occlusion (MCAO). For ischemic postconditioning group, bilateral common carotid arteries (CCA) were occluded for 5 min at 10 min after reperfusion. Infarct volume was quantified by TTC staining at 24 h after reperfusion. The upper panel shows the representative slices from each group. Data are expressed as percentage of infarct volume (n = 5). (B) Time‐course of oxygen‐glucose deprivation (OGD) postconditioning (an in vitro ischemic postconditioning model). Corticostriatal slices from adult male mice were postconditioned with OGD for 30 s, 1 min or 2 min at 1 min after 15 min of OGD. (C) Time windows of OGD postconditioning. Corticostriatal slices were postconditioned with OGD for 1 min at 1, 5, or 10 min after 15 min of OGD. (D) Comparison of corticostriatal slice injury among the acute OGD group without reperfusion, OGD/reperfusion group and OGD postconditioning group (OGD postconditioning was applied for 1 min at 5 min after 15 min of OGD). Slice injury was quantified by TTC conversion at 1 h after reperfusion. Data are expressed as percentage of TTC conversion compared with control (n = 4–5). Bars represent mean ± SEM, *P < 0.05, **P < 0.01. OGD/R, OGD/reperfusion.

Figure 2

Effects of hypoxic postconditioning on infarct volume and neurologic deficit scores at 24 h after transient middle cerebral artery occlusion (MCAO). (A) Experimental groups and protocols for hypoxic postconditioning studies. The middle cerebral artery (MCA) was occluded for 45 min, and reperfusion lasted for 24 h; Post 30″/30″ indicates five cycles of 30‐s/30‐s reoxygenation/hypoxia; Post 1′/1′, five cycles of 1‐min/1‐min reoxygenation/hypoxia; Post 2′/2′, five cycles of 2‐min/2‐min reoxygenation/hypoxia; Post 10′/10′, a cycle of 10‐min/10‐min reoxygenation/hypoxia; Post 30′/30′, a cycle of 30‐min/30‐min reoxygenation/hypoxia. (B) Quantification of infarct volume by TTC staining after various permutations of ischemia and hypoxic postconditioning. Data are expressed as percentage of infarct volume. (C) Quantification of neurologic deficit scores. Bars represent mean ± SEM (n = 6–8). *P < 0.05 vs. control group.

Figure 3

Effects of postconditioning with oxygen‐glucose deprivation (OGD), glucose deprivation (GD), oxygen deprivation (OD) and low glucose on OGD/reperfusion‐induced injury. (A) Corticostriatal slices were postconditioned with OGD, GD or OD for 1 min at 5 min after 15 min of OGD, and 1 h later slice injury was assessed by TTC conversion. (B) The concentration response of low‐glucose postconditioning. Zero, 1, 2 or 5 mmol/L glucose postconditioning was applied for 1 min at 5 min after 15 min of OGD. (C) The time‐course of low‐glucose postconditioning. One, 5, or 30‐min of 1 mmol/L glucose postconditioning was applied at 5 min after 15 min of OGD, and 5 min of OGD postconditioning was applied at the same time point simultaneously. Data are expressed as percentage of TTC conversion compared with control. Bars represent the mean ± SEM (n = 4–5), **P < 0.01. OGD/R, OGD/reperfusion; Post‐OGD, OGD postconditioning; Post‐GD, GD postconditioning; Post‐LG, low‐glucose postconditioning; Post‐OD, OD postconditioning.

Figure 4

Effects of postconditioning with oxygen‐glucose deprivation (OGD), glucose deprivation (GD) and oxygen deprivation (OD) on glutamate metabolism and GABA level. Corticostriatal slices were postconditioned with OGD, GD or OD for 1 min at 5 min after 15 min of OGD, and glutamate (A) and GABA (B) levels, GLT‐1 (C) and glutamine synthetase (GS) (D) protein expression and GS activity (E) were assayed at 1 h after reperfusion. Bars represent mean ± SEM (n = 3–4), *P < 0.05, **P < 0.01. OGD/R, OGD/reperfusion; Post‐OGD, OGD postconditioning; Post‐GD, GD postconditioning; Post‐OD, OD postconditioning.