Hypoxic Preconditioning Increases Blood-Brain Barrier Disruption in the Early Stages of Cerebral Ischemia.

Even though hypoxic preconditioning has been reported to produce neuroprotection, its effect on blood-brain barrier (BBB) disruption in the early stages of cerebral ischemia within the therapeutic window is not clear. Since hypoxic preconditioning increases expression of vascular endothelial growth factor (VEGF) that modulates vascular permeability, the effects of hypoxic preconditioning and VEGF on BBB permeability were investigated after one hour of focal cerebral ischemia. Rats were exposed to 8% of oxygen for two hours or room air and then 24 hours later, permanent middle cerebral artery (MCA) occlusion was performed. In some of the hypoxic preconditioned rats, a VEGF-A antibody was applied to the ischemic cortex one hour before MCA occlusion. One hour after MCA occlusion, the transfer coefficient (Ki) of 14C-α-aminoisobutyric acid was determined to measure the degree of BBB disruption. MCA occlusion increased the Ki when compared with the contralateral cortex (14.1 ± 4.0 vs 4.2 ± 1.9 µL/g/min, p < 0.0001). Hypoxic preconditioning further increased the Ki in the ischemic cortex when compared with the control rats (25.1 ± 8.7 µL/g/min, p < 0.01). Application of VEGF antibody to the ischemic cortex of the hypoxic preconditioned animals reduced the Ki to the level of the control rats (13.6 ± 5.1 µL/g/min, p < 0.01). Our data demonstrated that hypoxic preconditioning increased BBB disruption through a VEGF related pathway and suggest the possibility of aggravation of brain edema by hypoxic preconditioning in the early stages of cerebral ischemia.

Effects of rapamycin pretreatment on blood-brain barrier disruption in cerebral ischemia-reperfusion.

The mammalian target of rapamycin (mTOR) pathway is essential in neuronal survival and repair in cerebral ischemia. Decreases in blood-brain barrier (BBB) disruption are associated with a decrease in neuronal damage in cerebral ischemia. This study was performed to investigate how pre-inhibition of the mTOR pathway with rapamycin would affect BBB disruption and the size of the infarcted cortical area in the early stage of focal cerebral ischemia-reperfusion using quantitative analysis of BBB disruption. Rats were treated with 20mg/kg of rapamycin i.p. once a day for 2days (Rapamycin Group) or vehicle (Control Group) before transient middle cerebral artery (MCA) occlusion. After one hour of MCA occlusion and two hours of reperfusion, the transfer coefficient (Ki) of (14)C-α-aminoisobutyric acid ((14)C-AIB) to measure the degree of BBB disruption and the size of the cortical infarct were determined. Ischemia-reperfusion increased the Ki in the Rapamycin treated (+15%) as well as in the untreated control group (+13%). However, rapamycin pretreatment moderately decreased Ki in the contralateral (-30%) as well as in the ischemic-reperfused (-29%) cortex when compared with the untreated control group. Rapamycin pretreatment substantially increased the percentage of cortical infarct compared with the control group (+56%). Our data suggest that activation of mTOR pathway is necessary for neuronal survival in the early stage of cerebral ischemia-perfusion and that the reason for the enlarged cortical infarct by rapamycin pretreatment may be related to its non-BBB effects on the mTOR pathway.

Inhibition of neuronal nitric oxide synthase improves microregional O2 balance in cerebral ischemia-reperfusion.

Objectives Return of regional cerebral blood flow (rCBF) in focal cerebral ischaemia may not ensure proper distribution of blood flow to meet metabolic demand. This study was performed to determine how inhibition of neuronal nitric oxide synthase (NOS) during ischaemia-reperfusion would affect microregional O2 supply/consumption balances and their variation. Methods Twenty minutes before middle cerebral artery (MCA) occlusion, a NOS inhibitor 7-nitroindazole (7-NI) 50 mg/kg ip (7-NI group) or vehicle (control group) was administered. At 1 hour of ischaemia and 2 hours of reperfusion, rCBF, the size of cortical infarct and arteriolar and venular O2 saturations (20-60 µm in diameter) using cryomicrospectrophotometry were determined. Results Ischaemia-reperfusion decreased the average venular O2 saturation and the ratio of O2 supply/consumption. But, 7-NI treatment improved the average O2 supply/consumption ratio and venular O2 saturation (57.6 ± 1.3 vs 52.0 ± 3.8%) in ischaemia-reperfusion. The heterogeneity of venular O2 saturations reported as coefficient of variation (CV = 100 × SD/mean) was much smaller in the 7-NI than the control group (8.8 vs 15.5). The number of veins with low O2 saturation ( < 50%) was also smaller with the 7-NI (4/70) than the control group (18/70). The size of cortical infarct was smaller with 7-NI treatment. Discussion Our data suggest that inhibition of neuronal NOS by 7-NI improved microregional O2 balance in the ischaemic-reperfused cortex (IR-C). The improvement in microregional O2 balance could be one of the contributing factors to the reduced size of cortical infarct.

Local O2 Balance in Cerebral Ischemia-Reperfusion Improved during Pentobarbital Compared with Isoflurane Anesthesia.

BACKGROUND: Most anesthetics affect cerebral blood flow and metabolism. We compared microregional O2 balance in cerebral ischemia-reperfusion during pentobarbital and isoflurane anesthesia. METHODS: After 1 hour of middle cerebral artery occlusion and a 2-hour reperfusion under isoflurane (1.4%, n = 14) or pentobarbital (50 mg/kg, n = 14) anesthesia in rats, regional cerebral blood flow using (14)C-iodoantipyrine autoradiography, microregional arterial and venous O2 saturation (20-60 µm in diameter) using cryomicrospectrophotometry, and the size of cortical infarct were determined. RESULTS: Ischemia-reperfusion decreased the average cortical venous O2 saturation in both pentobarbital and isoflurane groups (P < .0001), which was higher (P < .05) with pentobarbital despite a similar average regional cerebral blood flow and O2 consumption. The heterogeneity of venous O2 saturation reported as a coefficient of variation (100 × standard deviation/mean) was smaller (P < .005) with pentobarbital than that with isoflurane (7.5 versus 16.1). The number of veins with low venous O2 saturation (<50%) was smaller (P < .005) with pentobarbital (5 of 80 versus 24 of 80). The percentage of cortical infarct in total cortex was smaller with pentobarbital (5.2 ± 2.5% versus 12.3 ± 2.6%, P < .001). CONCLUSIONS: In the cerebral ischemic-reperfused cortex, the average venous O2 saturation was higher, and its heterogeneity and the number of veins with low O2 saturation were smaller under pentobarbital than isoflurane anesthesia. This improvement in microregional O2 balance with pentobarbital was accompanied by the reduced cortical infarct. Our data suggest that the neurologic outcome could vary during cerebral ischemia-reperfusion depending on the anesthetics used.

Lower incidence of hypo-magnesemia in surgical intensive care unit patients in 2011 versus 2001.

BACKGROUND: Hypo-magnesemia is described to occur in as many as 65% of intensive care unit (ICU) patients. Magnesium (Mg) is a cofactor in over 300 enzymatic reactions involving energy metabolism, protein, and nucleic acid synthesis. The membrane pump that creates the electrical gradient across the cell membrane is dependent on Mg, and it is important in the activity of electrically excitable tissues. Since Mg regulates the movement of calcium in smooth muscle cells, it is also important in peripheral vascular tone and blood pressure. Studies have linked hypo-magnesemia to multiple chronic diseases and to a higher mortality rate. METHODS: To explore trends within our own tertiary care surgical ICU, we sampled our patients' laboratory records in 2001 and in 2011. Hypo-magnesemia in our ICU is defined as an Mg less than 2.0 mg/dL. RESULTS: This retrospective review of all SICU patients from October to December revealed that there was a significant increase (P < 0.01) in the patients with their serum Mg level measured between 2001 (89%) and 2011 (95%). There was a significant decrease (P < 0.001) in patients with hypomagnesemia (< 2 mg/dL) between 2001 (47.5%) and 2011 (33.0%). On the other hand, there was a significant increase (P < 0.001) in patients with normal serum Mg level (> 2 mg/dL) between 2001 (52.5%) and 2011 (67.0%). CONCLUSIONS: There was not only more monitoring of Mg in 2011, but a lower incidence of hypo-Mg compared to 2001. Possible explanations include changing patterns of antibiotic and diuretic use, less amphotericin use, more frequent laboratory surveillance, and better trained ICU practitioners.

Effects of dexmedetomidine on microregional O2 balance during reperfusion after focal cerebral ischemia.

BACKGROUND: This study was performed to determine whether there is an association between microregional O2 balance and neuronal survival in cerebral ischemia-reperfusion using dexmedetomidine, an α2-adrenoreceptor agonist and a sedative. METHODS: Rats were subjected to 1 hour middle cerebral artery occlusion and a 2-hour reperfusion. During reperfusion, normal saline (n = 14) or dexmedetomidine 1 µg/kg/minute (n = 14) was infused intravenously. At 2 hours of reperfusion, regional cerebral blood flow using (14)C-iodoantipyrine autoradiography, microregional arterial and venous (20-60 µm in diameter) O2 saturation (SvO2) using cryomicrospectrophotometry, and the size of cortical infarction were determined. RESULTS: Ischemia-reperfusion decreased microregional SvO2 (52.9 ± 3.7% vs. 61.1 ± .6%, P < .005) with increased variation or heterogeneity (P < .0001) with similar regional cerebral blood flow and O2 consumption. Dexmedetomidine during reperfusion decreased the heterogeneity of SvO2 that was analyzed with an analysis of variance (P < .01) and reported as coefficient of variation (100 × standard deviation/Mean) (11.8 vs. 16.4). The number of veins with O2 saturation less than 50% decreased with dexmedetomidine (13/80 vs. 27/81, P < .01). The percentage of cortical infarct in total cortex was smaller with dexmedetomidine (8.3 ± 2.2% vs. 12.6 ± 1.5%, P < .005). CONCLUSIONS: In the cerebral ischemic reperfused cortex, dexmedetomidine decreased the heterogeneity of SvO2 and the number of small veins with low O2 saturation suggesting improved microregional O2 supply/consumption balance. The improvement was accompanied by the reduced size of cortical infarction.

Effects of the thioredoxin-1 inhibitor PX-12 on blood-brain barrier permeability in the early stage of focal cerebral ischemia.

BACKGROUND/AIMS: Since a thioredoxin-1 (Trx-1) inhibitor, 1-methylpropyl-2-imidazolyl disulfide (PX-12) which is an antitumor agent, significantly decreased vascular permeability in tumor xenografts within a few hours of treatment, we tested whether PX-12 would attenuate blood-brain barrier (BBB) disruption in the early stage of focal cerebral ischemia and whether its action could be affected by vascular endothelial growth factor (VEGF) which interacts with the Trx-1 system. METHODS: In rats, 40 min after intravenous infusion of either 25 mg/kg of PX-12 (PX-12 group) or normal saline (control group), a middle cerebral artery (MCA) was occluded. In half of each group, VEGF (10(-10) mol/l) was applied topically in the ischemic cortex (IC). Ninety minutes after MCA occlusion, the transfer coefficient (Ki) of (14)C-α-aminoisobutyric acid and the volume of (3)H-dextran distribution were determined to measure the degree of BBB disruption. VEGF protein levels were determined using Western blot analysis. RESULTS: MCA occlusion increased the Ki in the control (+196%) as well as in the PX-12-treated rats (+90%), but the Ki of the IC of the PX-12 group was lower (-42%) than that of the control rats. VEGF protein levels were decreased in both the IC (-9.5%) and the contralateral cortex (CC; -10.2%) with PX-12 treatment. In the VEGF-treated rats, PX-12 also attenuated (-41%) the Ki of the IC. The difference in the volume of dextran distribution between the IC and the CC became insignificant with PX-12 treatment with or without VEGF application. CONCLUSION: Our data demonstrated that PX-12 was effective in decreasing BBB disruption in the early stage of focal cerebral ischemia and that VEGF is not an important factor involved in the action of PX-12 on BBB permeability.

Cerebral ischemia and reperfusion increases the heterogeneity of local oxygen supply/consumption balance.

BACKGROUND AND PURPOSE: After cerebral vessel blockage, local blood flow and O2 consumption becomes lower and oxygen extraction increases. With reperfusion, blood flow is partially restored. We examined the effects of ischemia-reperfusion on the heterogeneity of local venous oxygen saturation in rats in order to determine the pattern of microregional O2 supply/consumption balance in reperfusion. METHODS: The middle cerebral artery was blocked for 1 hour using the internal carotid approach in 1 group (n=9) and was then reperfused for 2 hours in another group (n=9) of isoflurane-anesthetized rats. Regional cerebral blood flow was determined using a C(14)-iodoantipyrine autoradiographic technique. Regional small vessel arterial and venous oxygen saturations were determined microspectrophotometrically. RESULTS: After 1 hour of ischemia, local cerebral blood flow (92±10 versus 50±10 mL/min per 100 g) and O2 consumption (4.5±0.6 versus 2.7±0.5 mL O2/min per 100 g) decreased compared with the contralateral cortex. Oxygen extraction increased (4.7±0.2 versus 5.4±0.3 mL O2/100 mL) and the variation in small vein (20-60 µm) O2 saturation as determined by its coefficient of variation (=100×SD/mean) increased (5.5 versus 10.5). With 2 hours of reperfusion, the blood flow decrement was reduced and O2 consumption returned to the value in the contralateral cortex. Oxygen extraction remained elevated in the ischemic-reperfused area and the coefficient of variation of small vein O2 saturation increased further (17.3). CONCLUSIONS: These data indicated continued reduction of O2 supply/consumption balance with reperfusion. They also demonstrated many small regions of low oxygenation within the reperfused cortical region.

Effects of blockade of NMDA receptors on cerebral oxygen consumption during hyperosmolar BBB disruption in rats.

Hyperosmolar blood-brain barrier (BBB) disruption has been reported to increase cerebral O2 consumption. This study was performed to test whether blockade of N-methyl-d-aspartate (NMDA) receptor would affect cerebral O2 consumption during hyperosmolar BBB disruption. A competitive NMDA receptor antagonist CGS-19755 10mg/kg was injected iv 15min before intracarotid infusion of 25% mannitol. Twelve min after BBB disruption, the BBB transfer coefficient (Ki) of (14)C-α-aminoisobutyric acid ((14)C-AIB) was measured. Regional cerebral blood flow (rCBF), regional arteriolar and venular O2 saturation (SaO2 and SvO2 respectively), and O2 consumption were determined using (14)C-iodoantipyrine autoradiography and cryomicrospectrophotometry in alternate slices of the brain tissue. The Ki of (14)C-AIB was markedly increased with hyperosmolar mannitol in both the control (5.8×) and the CGS treated rats (5.2×). With BBB disruption, the O2 consumption was significantly increased (+39%) only in the control but not in the CGS treated rats and was significantly lower (-29%) in the CGS treated than the control rats. The distribution of SvO2 was significantly shifted to the higher concentrations with CGS treatment. Our data demonstrated an increase of O2 consumption by hyperosmolar BBB disruption and attenuation of the increase with NMDA blockade without affecting the degree of BBB disruption.

Effects of cannabinoid receptor agonist WIN 55,212-2 on blood-brain barrier disruption in focal cerebral ischemia in rats.

This study was performed to investigate whether WIN 55,212-2 (WIN), a cannabinoid receptor agonist, could attenuate blood-brain barrier (BBB) disruption in focal cerebral ischemia in rats and whether the CB 1 receptor antagonist rimonabant could prevent this attenuation. A total of 0.3 or 1 mg/kg of WIN was injected intravenously before and after permanent middle cerebral artery (MCA) occlusion. Some animals were pretreated with rimonabant 2 mg/kg i.p. before receiving 0.3 mg/kg of WIN. At 1 h after MCA occlusion, BBB permeability was determined by measuring the transfer coefficient (K(i)) of (14)C-α-aminoisobutyric acid and the volume of dextran distribution. With MCA occlusion, K(i) increased in the ischemic cortex (IC) in all of the experimental groups. However, the K(i) of the IC of the WIN 0.3 and 1 mg/kg groups was lower (­46 and ­42%, respectively, p < 0.05) than that of the control group. With rimonabant pretreatment, the K(i) of the IC became higher ((+)88%, p < 0.05) than with WIN 0.3 mg/kg alone and similar to that of the control rats. The difference in the volume of dextran distribution between the IC and the contralateral cortex was significant in the control but not in the WIN-treated rats. With rimonabant pretreatment, however, the difference became significant. Our data demonstrated that WIN could attenuate BBB disruption in focal cerebral ischemia and this attenuation could be prevented with rimonabant. Our data suggest an involvement of CB(1) receptors in the regulation of BBB disruption in the early stage of stroke.