Optimal depth and duration of mild hypothermia in a focal model of transient cerebral ischemia: effects on neurologic outcome, infarct size, apoptosis, and inflammation

BACKGROUND and PURPOSE: Mild hypothermia is possibly the single most effective method of cerebroprotection developed to date. However, many questions regarding mild hypothermia remain to be addressed before its potential implementation in the treatment of human stroke. Here we report the results of 2 studies designed to determine the optimal depth and duration of mild hypothermia in focal stroke and its effects on infarct size, neurological outcome, programmed cell death, and inflammation. METHODS: Rats underwent a 2-hour occlusion of the left middle cerebral artery. In the first study (I) animals were kept (intraischemically) at either 37 degreesC (n=8), 33 degreesC (n=8), or 30 degreesC (n=8). Study II consisted of 4 groups: (1) controls (37 degreesC, n=10), (2) 30 minutes of hypothermia started at ischemic onset (33 degreesC, n=9), (3)1 hour (33 degreesC, n=8), and (4) 2 hours (33 degreesC, n=8). Brain temperature was measured by a thermocouple probe placed in the contralateral cortex. After suture removal, all animals were rewarmed and reperfused for 22 hours (I) or 70 hours (II). RESULTS: Mild hypothermia to 33 degreesC or 30 degreesC was neuroprotective (17+/-7% and 27+/-6%, respectively) relative to controls (53+/-8%, P<0.02), but 33 degreesC was better tolerated and recovery from anesthesia was faster. The neurological score of hypothermic animals was significantly better than that of controls (I & II) at both 24 and 72 hours postischemia except for the 30-minute group (II), which showed no improvement. In Study II, 2 hours of hypothermia reduced injury by 59%, 1 hour reduced injury by 84% whereas 30 minutes did not reduce injury. Normalized for infarct size, 2 hours of mild hypothermia decreased neutrophil accumulation by 57% whereas both 1 hour and 30 minutes had no effect. At 72 hours, 1 and 2 hours of mild hypothermia decreased transferase dUTP nick-end labeling (TUNEL) staining by 78% and 99%, respectively, and 30 minutes of hypothermia had no effect. CONCLUSIONS: Intraischemic mild hypothermia must be maintained for 1 to 2 hours to obtain optimal neuroprotection against ischemic cell death due to necrosis and apoptosis.     

Effects of mild (33 degrees C) and moderate (29 degrees C) hypothermia on cerebral blood flow and metabolism, lactate, and extracellular glutamate in experimental head injury

The effects of mild (33 degrees C) and moderate (29 degrees C) hypothermia were investigated to determine which temperature was more effective against compression-induced cerebral ischemia. Eighteen cats were anesthetized. The animals were divided into three groups according to deep-brain temperature (control, 37 degrees C; mild hypothermia, 33 degrees C; and moderate hypothermia, 29 degrees C). Intracranial pressure (ICP) and cerebral blood flow (CBF) were monitored, the latter by hydrogen clearance. Arteriovenous oxygen difference (AVDO2) and cerebral venous oxygen saturation (ScvO2) were measured in blood samples from the superior sagittal sinus. The cerebral metabolic rate of oxygen (CMRO2) and the cerebral metabolic rate of lactate (CMR lactate) were calculated. Extracellular glutamate was measured by microdialysis. ICP was increased by inflation of an epidural balloon until CBF became zero, and this ischemia was maintained for 5 min, after which the balloon was quickly deflated. All parameters were recorded over 6 h. Evans blue was injected to examine vascular permeability changes. CBF was decreased by 56% by mild hypothermia and by 77% by moderate hypothermia. Mild hypothermia had a coupled metabolic suppression whereas moderate hypothermia significantly increased AVDO2 and decreased ScvO2, producing a low CBF/CMRO2 (relative ischemia). After balloon deflation, all three groups showed reactive hyperemia, which was significantly reduced by mild and moderate hypothermia. CBF then decreased to 50% of pre-inflation values and ScvO2 decreased (post-ischemic hypoperfusion). CBF/CMRO2, ScvO2, and AVDO2 did not differ significantly between the three groups. After balloon deflation, all three groups showed increased CMR lactate, which was significantly reduced by mild and moderate hypothermia. Extracellular glutamate increased in control animals (3.8 +/- 1.72 microM), an effect most effectively suppressed in the mild hypothermia group (1.0 +/- 0.46 microM). Damaged tissue volumes as indicated by Evans blue dye extravasation were 729 +/- 89 mm3 in control, 247 +/- 56 mm3 in mild hypothermia, and 267 +/- 35 mm3 in moderate hypothermia animals. These data suggest that mild hypothermia (33 degrees C) might be the optimal brain temperature to treat compression-related cerebral ischemia.     

Postischemic hyperthermia exacerbates neurologic injury after deep hypothermic circulatory arrest

BACKGROUND: Aggressive surface warming is a common practice in the pediatric intensive care unit. However, recent rodent data emphasize the protective effect of mild (2 degrees - 3 degrees C) hypothermia after cerebral ischemia. This study evaluates different temperature regulation strategies after deep hypothermic circulatory arrest with a survival piglet model. METHODS: Fifteen piglets were randomly assigned to 3 groups. All groups underwent 100 minutes of deep hypothermic circulatory arrest at 15 degrees C. Brain temperature was maintained at 34 degrees C for 24 hours after cardiopulmonary bypass in group I, 37 degrees C in group II, and 40 degrees C in group III. Neurobehavioral recovery was evaluated daily for 3 days after extubation by neurologic deficit score (0, normal; 500, brain death) and overall performance category (1, normal; 5, brain death). Histologic examination was assessed for hypoxic-ischemic injury (0, normal; 5, necrosis) in a blinded fashion. RESULTS: All results are expressed as mean +/- standard deviation. Recovery of neurologic deficit score (12.0 +/- 17.8, 47.0 +/- 49.95, 191.0 +/- 179.83; P = .05 for group I vs III), overall performance category (1.0 +/- 0.0, 1.4 +/- 0.6, 2.8 +/- 1.3; P < .05 for group I vs III), and histologic scores (0.0 +/- 0.0, 1.0 +/- 1.2, 2.8 +/- 1.8; P < .05 for group I vs III cortex) were significantly worse in hyperthermic group III. These findings were associated with a significantly lower cytochrome aa3 recovery determined by near-infrared spectroscopy in group III animals (P = .0041 for group I vs III). No animal recovered to baseline electroencephalographic value by 48 hours after deep hypothermic circulatory arrest. Recovery was significantly delayed in the hyperthermic group III animals, with a lower amplitude 14 hours after the operation, which gradually increased with time (P < .05 for group III vs groups I and II). CONCLUSIONS: Mild postischemic hyperthermia significantly exacerbates functional and structural neurologic injury after deep hypothermic circulatory arrest and should therefore be avoided.     

Effect of brain, body, and magnet bore temperatures on energy metabolism during global cerebral ischemia and reperfusion monitored by magnetic resonance spectroscopy in rats

To record brain temperature for comparison with rectal and temporalis muscle temperatures in preliminary studies before MR spectroscopy experiments, a thermistor was inserted into the basal ganglia in eight anesthetized, ventilated, and physiologically monitored rats. The rats were placed in an MR spectrometer and subjected to 60 min of global cerebral ischemia and 2 h of reperfusion without radiofrequency (RF) pulsing. Body temperature was maintained at 37.5-38.0 degrees C (normothermia) or 36.5-37.0 degrees C (mild hypothermia). Brain temperature during ischemia, which dropped to 31.9 +/- 0.3 (hypothermia) and 33.6 +/- 0.5 degrees C (normothermia), correlated with temporalis muscle temperature (r2 = 0.92) but not with body or magnet bore temperature measurements. Ischemia reduced brain temperature approximately 1.7 degrees C in rats subjected to mild hypothermia (1 degree reduction of body temperature). Parallel MR spectroscopy experiments showed no significant difference in energy metabolites between normothermic and hypothermic rats during ischemia. However, the metabolic recovery was more extensive 20-60 min after the onset of reperfusion in hypothermic rats, although not thereafter (P < 0.05). Mild hypothermia speeds metabolic recovery temporarily during reperfusion but does not retard energy failure during global ischemia in rats.     

Mild hypothermia as a protective therapy during intracranial aneurysm surgery: a randomized prospective pilot trial

OBJECTIVE: To conduct a pilot trial of mild intraoperative hypothermia during cerebral aneurysm surgery. METHODS: One hundred fourteen patients undergoing cerebral aneurysm clipping with (n = 52) (World Federation of Neurological Surgeons score < or =III) and without (n = 62) acute aneurysmal subarachnoid hemorrhage (SAH) were randomized to normothermic (target esophageal temperature at clip application of 36.5 degrees C) and hypothermic (target temperature of 33.5 degrees C) groups. Neurological status was prospectively evaluated before surgery, 24 and 72 hours postoperatively (National Institutes of Health Stroke Scale), and 3 to 6 months after surgery (Glasgow Outcome Scale). Secondary outcomes included postoperative critical care requirements, respiratory and cardiovascular complications, duration of hospitalization, and discharge disposition. RESULTS: Seven hypothermic patients (12%) could not be cooled to within 1 degrees C of target temperature; three of the seven were obese. Patients randomized to the hypothermic group more frequently required intubation and rewarming for the first 2 hours after surgery. Although not achieving statistical significance, patients with SAH randomized to the hypothermic group, when compared with patients in the normothermic group, had the following: 1) a lower frequency of neurological deterioration at 24 and 72 hours after surgery (21 versus 37-41%), 2) a greater frequency of discharge to home (75 versus 57%), and 3) a greater incidence of good long-term outcomes (71 versus 57%). For patients without acute SAH, there were no outcome differences between the temperature groups. There was no suggestion that hypothermia was associated with excess morbidity or mortality. CONCLUSION: Mild hypothermia during cerebral aneurysm surgery is feasible in nonobese patients and is well tolerated. Our results indicate that a multicenter trial enrolling 300 to 900 patients with acute aneurysmal SAH will be required to demonstrate a statistically significant benefit with mild intraoperative hypothermia.     

Mild hypothermia after cardiac arrest in dogs does not affect postarrest cerebral oxygen uptake/delivery mismatching

PURPOSE: To compare measurements of cerebral arteriovenous oxygen content differences (oxygen extraction ratios, oxygen utilization coefficients) in dogs after cardiac arrest, resuscitated under normothermia vs. mild hypothermia for 1-2 h or 12 h. METHODS: In 20 dogs, we used our model of ventricular fibrillation (no blood flow) of 12.5 min, reperfusion with brief cardiopulmonary bypass, and controlled ventilation, normotension, normoxemia, and mild hypocapnia to 24 h. We compared a normothermic control Group I (37.5 degrees C) (n = 8); with brief mild hypothermia in Group II (core and tympanic membrane temperature about 34 degrees C during the first hour after arrest) (n = 6); and with prolonged mild hypothermia in Group III (34 degrees C during the first 12 h after arrest) (n = 6). RESULTS: In Group I, the cerebral arteriovenous O2 content difference was 5.6 +/- 1.6 ml/dl before arrest; was low during reperfusion (transient hyperemia) and increased (worsened) significantly to 8.8 +/- 2.8 ml/dl at 1 h, remained increased until 18 h, and returned to baseline levels at 24 h after reperfusion. These values were not significantly different in hypothermic Groups II and III. The cerebral venous (saggital sinus) PO2 (PssO2) was about 40 mmHg (range 29-53) in all three groups before arrest and decreased significantly below baseline values, between 1 h and 18 h after arrest; the lowest mean values were 19 +/- 19 mmHg in Group I, 15 +/- 8 in Group II (NS), and 21 +/- 3 in Group III (NS). Postarrest PssO2 values of < or = 20 mmHg were found in 6/8 dogs in Group I, 5/6 in Group II and 4/6 in Group III. Among the 120 values of PssO2 measured between 1 h and 18 h after arrest, 32 were below the critical value of 20 mmHg. CONCLUSIONS: After prolonged cardiac arrest, critically low cerebral venous O2 values suggest inadequate cerebral O2 delivery. Brief or prolonged mild hypothermia after arrest does not mitigate the postarrest cerebral O2 uptake/delivery mismatching.     

Two proto-oncogenes that play dual roles in embryonal cell growth and differentiation

BACKGROUND AND PURPOSE: Clinical and experimental data indicate that hyperglycemia can aggravate the consequences of stroke and cerebral ischemia. The purpose of this study was to examine the effects of moderate hyperglycemia on the response of the blood-brain barrier to normothermic (37 degrees C) and hypothermic (30 degrees C) global forebrain ischemia. METHODS: Sixteen rats underwent 20 minutes of four-vessel occlusion followed by 30 minutes of postischemic recirculation. We used the protein tracer horseradish peroxidase as an indicator of increased vascular permeability, and rats were perfusion-fixed for microscopic analysis. To produce moderate hyperglycemia, we gave an intraperitoneal injection of 50% dextrose 15 minutes before the ischemic insult. RESULTS: After normothermic brain ischemia, normoglycemic rats (plasma glucose level, 115 +/- 3 mg/dl) demonstrated extravasated horseradish peroxidase mainly restricted to the cerebral cortex. In contrast, more severe and widespread protein extravasation was documented throughout the neuraxis of hyperglycemic (plasma glucose level, 342 +/- 27) rats. Sites of protein leakage included the cerebral cortex, striatum, hippocampus, thalamus, and cerebellum. Foci of protein extravasation were associated with pial and large penetrating vessels. Intraischemic hypothermia significantly attenuated the blood-brain barrier consequences of hyperglycemic brain ischemia. CONCLUSIONS: Under normothermic ischemic conditions, hyperglycemia significantly worsens the degree of acute blood-brain barrier breakdown compared with normoglycemia. Postischemic blood-brain barrier disruption may play an important role in the pathogenesis of increased brain damage associated with systemic hyperglycemia.     

Diabetics with coronary disease have a prevalence of asymptomatic ischemia during exercise treadmill testing and ambulatory ischemia monitoring similar to that of nondiabetic patients. An ACIP database study. ACIP Investigators. Asymptomatic Cardiac Ischemia Pilot Investigators

BACKGROUND: There are conflicting data as to whether diabetics have a higher prevalence of asymptomatic ST-segment depression during exercise treadmill testing (ETT) and ambulatory ECG (AECG) monitoring. This study was conducted to determine whether diabetic patients with coronary disease enrolled in the Asymptomatic Cardiac Ischemia Pilot (ACIP) have more episodes of asymptomatic ischemia during ETT and 48-hour AECG monitoring than nondiabetic patients and to compare differences in angiographic variables and the magnitude of ischemia as measured by standard ETT and AECG criteria. METHODS AND RESULTS: Angiographic variables and the prevalence and magnitude of ischemia during the qualifying ETT and 48-hour AECG were compared by the presence and absence of diabetes mellitus in 558 randomized ACIP patients. Seventy-seven patients had a history of diabetes and were taking oral hypoglycemics or insulin (diabetic group); 481 patients did not meet these criteria (nondiabetic group). Multivessel disease (87% versus 74%, P = .01) was more frequent in the diabetic group. The percentages of patients without angina during the ETT were similar in the diabetic and nondiabetic groups (36% and 39%, respectively). Time to onset of > or = 1-mm ST-segment depression and time to onset of angina were similar in both groups. The percentages of patients with only asymptomatic ST-segment depression during the 48-hour AECG were similar in the diabetic and nondiabetic groups (94% versus 88%, respectively). However, total ischemic time per 24 hours (15.0 +/- 21.4 versus 23.6 +/- 31.1 minutes, P = .02), ischemic time per episode (6.3 +/- 4.6 versus 9.0 +/- 8.7 minutes, P < .01), and the maximum depth of ST-segment depression tended to be less in the diabetic group. CONCLUSIONS: Patients enrolled in ACIP were selected on the basis of an abnormal ETT and 48-hour AECG and ability to undergo coronary revascularization. When patients with diabetes mellitus were compared with those without diabetes, there was a similar prevalence of asymptomatic ischemia during ETT and 48-hour AECG monitoring. Despite more extensive and diffuse coronary disease, diabetic ACIP patients tended to have less measurable ischemia during the 48-hour AECG.     

Long-term neuroprotection by benzodiazepine full versus partial agonists after transient cerebral ischemia in the gerbil [corrected

The ability of diazepam, a benzodiazepine full agonist, and imidazenil, a benzodiazepine partial agonist, to protect hippocampal area CA1 neurons from death for at least 35 days after cerebral ischemia was investigated. Diazepam (10 mg/kg) administered to gerbils 30 and 90 minutes after forebrain ischemia produced significant protection of hippocampal area CA1 pyramidal neurons 7 days later. In gerbils surviving for 35 days, diazepam produced the same degree of neuroprotection (70% +/- 30%) in the hippocampus compared with 7 days after ischemia. The therapeutic window for diazepam was short; there was no significant neuroprotection when the administration of diazepam was delayed to 4 hours after ischemia. The neuroprotective dose of diazepam also produced hypothermia (approximately 32 degrees C) for several hours after injection. To assess the role of hypothermia in neuroprotection by diazepam, hypothermia depth and duration was simulated using a cold-water spray in separate gerbils. Seven days after ischemia, neuroprotection by hypothermia was similar to that produced by diazepam. However, 35 days after ischemia, there was no significant protection by hypothermia, suggesting that hypothermia does not play a significant role in long-term diazepam neuroprotection. Imidazenil (3 mg/kg), which produced only minimal hypothermia, protected area CA1 of hippocampus to the same degree as that by diazepam 7 days after ischemia. At 35 days after ischemia, significant protection remained, but it was considerably reduced compared with 7 days. Like diazepam, the therapeutic window for imidazenil was short. Imidazenil neuroprotection was lost when the drug was administered as early as 2 hours after ischemia. The ability of ischemia to produce deficits in working memory and of benzodiazepines to prevent the deficits also was investigated. Gerbils trained on an eight-arm radial maze before ischemia demonstrated a significant increase in the number of working errors 1 month after ischemia. The ischemia-induced deficits in working memory were completely prevented by diazepam but not by imidazenil. There was a significant, but weak, negative correlation between the degree of CA1 pyramidal cell survival and the number of working errors in both the diazepam and imidazenil groups. Thus, if given early enough during reperfusion, both benzodiazepine full and partial agonists are neuroprotective for at least 35 days, but the lack of sedating side effects of imidazenil must be weighed against its reduced efficacy.     

Can acute stroke be treated with hypothermia?

In animal stroke models, treatment with mild hypothermia (30-34 degrees C) for 3-4 hours may reduce the size of cerebral infarction if started within three hours of the initiation of cerebral ischaemia. The mechanism by which hypothermia exerts its neuroprotective effect is unknown, but experimental studies have shown the release of neurotoxic excitatory amino acids and free oxygen radicals to be reduced during hypothermic ischaemia. In patients with acute stroke, body temperature above 37.5 degrees C are associated with poor outcome, and temperatures below 36.5 degrees C with improved outcome, compared to normothermic patients. Due to the unpleasantness of cooling and side effects as shivering, hypothermia may not be tolerated by stroke patients without sedation of light anaesthesia which may increase the risk of hypotension and respiratory complications. However, lowering body temperature by 1-2 degrees C may suffice to improve functional outcome in acute stroke patients, and such mild hypothermia should be tested in randomized controlled clinical trials.     

Proton and neutron excitations in superdeformed 150Tb

To determine which of two treatments for reducing ischemic injury after temporal focal ischemia is more effective, the effects of mild (33 degrees C) intraischemic hypothermia were compared with those of mannitol, the most commonly used neuroprotective agent. Four groups of Sprague-Dawley rats underwent 1 hour of endovascular middle cerebral artery occlusion followed by 23 hours of normothermic reperfusion. The four experimental groups were as follows: Group A, saline control; Group B, mannitol (25%, 1 g/kg); Group C, hypothermia; and Group D, hypothermia plus man-nitol. Laser-Doppler estimates of cortical blood flow showed that hypothermia did not affect blood flow during ischemia or reperfusion. Mannitol increased cortical blood flow during ischemia and reperfusion under both normothermic and hypothermic conditions (p < 0.05). Neurological deficit was significantly less severe in treated rats (Group B, p < 0.05; Group C or D, p < 0.01) than in controls (Group A). Infarct volume, measured on semiserial Nissl-stained sections, was significantly smaller in treated rats (p < 0.01) than in controls. Infarct volume was also significantly smaller in rats treated with hypothermia than in those treated with mannitol (Group C vs. Group B, p < 0.05); there was no difference between rats treated with mannitol and those treated with mannitol and hypothermia. All three treatments reduced infarct area in the ischemic penumbra; hypothermia with or without mannitol also reduced infarct area in the ischemic core. These results demonstrate that both mild intraischemic hypothermia and mannitol reduce infarct size and neurological deficit: hypothermia reduces infarct size more effectively than mannitol, and mannitol adds no significant protection to hypothermia, whereas hypothermia adds significant protection beyond that afforded by mannitol after brief focal ischemia followed by reperfusion in rats. The results suggest that mild intraischemic hypothermia alone, or in combination with mannitol, may be useful in avoiding ischemic injury from temporary vessel occlusion during cerebrovascular surgery.     

Effects of intermittent reperfusion on brain pHi, rCBF, and NADH during rabbit focal cerebral ischemia

BACKGROUND AND PURPOSE: The use of intermittent reperfusion versus straight occlusion during neurovascular procedures is controversial. This experiment studied the effects of intermittent reperfusion and single occlusion on intracellular brain pH (pHi), regional cerebral or cortical blood flow, and nicotinamide adenine dinucleotide (NADH) fluorescence during temporary focal ischemia. METHODS: Twenty fasted rabbits under 1.0% halothane anesthesia were divided into four groups: (1) nonischemic controls, (2) 60 minutes of uninterrupted focal ischemia, (3) 2 x 30-minute periods of focal ischemia separated by a 5-minute reperfusion, and (4) 4 x 15-minute periods of focal ischemia separated by three 5-minute reperfusion periods. Focal ischemia was produced by occlusion of both the middle cerebral and ipsilateral anterior cerebral arteries. After the final occlusion, there was a 3-hour reperfusion period in all groups. Regional cerebral and cortical blood flow, brain pHi, and NADH fluorescence were measured with in vivo panoramic fluorescence imaging. RESULTS: During occlusion, regional cerebral and cortical blood flows and NADH fluorescence values were not different among the groups. Brain pHi was significantly lower in the 4 x 15-minute group compared with the 1 x 60-minute group (6.57 +/- 0.02 versus 6.73 +/- 0.06; P < .03) but not significant when compared with the 2 x 30-minute group. During the short reperfusion periods, all parameters returned to normal except for NADH fluorescence levels, which remained elevated. During the postischemic final reperfusion period, there was a mild brain alkalosis of approximately 7.1 in all groups. There were no significant differences in NADH fluorescence among groups during the final reperfusion. Regional cerebral and cortical blood flow returned to near normal values in all groups. CONCLUSIONS: This study demonstrates that intermittent reperfusion during temporary focal ischemia has different effects on the intracytoplasmic and the intramitochondrial compartments: worsening of brain cytoplasmic pHi but no significant differences in the oxidation/reduction level of mitochondrial NADH.     

Cellular uncoupling during ischemia in hypertrophied and failing rabbit ventricular myocardium: effects of preconditioning

BACKGROUND: Patients with heart failure show a very high incidence of arrhythmias and sudden death that is often preceded by ischemia; however, data on electrophysiological changes during ischemia in failing myocardium are sparse. We studied electrical uncoupling during ischemia in normal and failing myocardium. METHODS AND RESULTS: Tissue resistance, intracellular Ca2+ concentration (Indo-1 fluorescence ratio), and mechanical activity were simultaneously determined in arterially perfused right ventricular papillary muscles from 11 normal and 15 failing rabbits. Heart failure was induced by combined volume and pressure overload. Before sustained ischemia, muscles were subjected to control perfusion (non-PC) or ischemic preconditioning (PC). The onset of uncoupling during ischemia was equal in non-PC normal (13.6+/-0.9 minutes of ischemia) and non-PC failing hearts (13.3+/-0.7 minutes of ischemia). PC postponed uncoupling in normal hearts by 10 minutes. In failing hearts, however, PC caused a large variability in the onset of uncoupling during ischemia (mean, 12.2+/-2.1; range, 5 to 22 minutes of ischemia). The duration of uncoupling process was prolonged in failing hearts (12.9+/-0.9 minutes) compared with normal hearts (7.8+/-0.4 minutes). The degree of heart failure and relative heart weight of the failing hearts significantly correlated with the earlier uncoupling after PC and the duration of uncoupling. In every experiment, the start of Ca2+ rise and contracture preceded uncoupling during ischemia. CONCLUSIONS: The duration of the process of ischemia-induced electrical uncoupling in failing hearts is prolonged compared with that in normal hearts. Ischemic PC has detrimental effects in severely failing papillary muscles because it advances the moment of irreversible ischemic damage.     

Cerebral protection by intermittent reperfusion during temporary focal ischemia in the rat

Temporary arterial occlusion has been routinely used as an adjunct in intracranial aneurysm surgery. This has commonly been performed using a protocol of multiple short periods of occlusion alternating with periods of restoration of normal circulation. Recently, the logical basis of this method has come under scrutiny. There is extensive experimental evidence to suggest that repetitive, brief periods of global ischemia may cause more severe cerebral injury than an equivalent single period of global ischemia. Only recently has this issue begun to be addressed with regard to focal ischemia. Hence, despite the common use of temporary clipping, little experimental data are available regarding the ischemic consequences of temporary arterial occlusion with periods of reperfusion versus uninterrupted temporary occlusion. To investigate this issue, a protocol of occlusion/reperfusion that simulates the temporal profile that occurs during surgery was performed in a rat model of focal ischemia. Sixteen anesthetized Sprague-Dawley rats were divided into two groups. The animals in Group I underwent 60 minutes of uninterrupted middle cerebral artery occlusion and the animals in Group II were subjected to six separate 10-minute occlusion periods with 5 minutes of reperfusion between occlusions. Histopathological analysis was performed 72 hours postischemia. Group I had significantly increased mean infarction volumes (50.0 +/- 12.1 mm3) compared to Group II (8.7 +/- 3.1 mm3) (p = 0.008). Injuries in Group I occurred in both the cortex and striatum, whereas Group II showed only striatal injuries. Furthermore, the extent of the injuries in Group II was less severe, characterized by ischemic neuronal injury rather than frank infarction. The results indicate that intermittent reperfusion is neuroprotective during temporary focal ischemia and support the hypothesis that intermittent reperfusion is beneficial if temporary clipping is required during aneurysm repair.     

Interactions between hypothermia and the latency to ischemic depolarization: implications for neuroprotection

BACKGROUND: The authors postulated that hypothermic neuroprotection can be attributed to a delayed onset of ischemic depolarization. METHODS: Halothane-anesthetized rats were prepared for near-complete forebrain ischemia. Direct current (DC) potential microelectrodes were placed in hippocampal CA1. The pericranial temperature was maintained at 31 degrees C, 33 degrees C, 35 degrees C, or 37 degrees C (n = 6 per group). Bilateral carotid occlusion with systemic hypotension was initiated for 10 min. The time to onset of the DC shift was recorded. In a second experiment, rats were assigned to 37 degrees C or 31 degrees C for 10 min of ischemia, or to 31 degrees C for 14 min of ischemia (n = 8 per group). These durations of ischemia were defined to allow 9 min of ischemic depolarization in the 37 degrees C-10 min and 31 degrees C-14 min groups. Neurologic and histologic outcomes were examined 7 days later. RESULTS: Hippocampal CA1 time to depolarization increased with decreasing temperature (P < 0.0001). Time to depolarization was increased by approximately 4 min in the rats maintained at 31 degrees C compared with those at 37 degrees C. Time to repolarization during reperfusion was not affected by temperature. Increasing the duration of ischemia from 10 min to 14 min with the pericranial temperature maintained at 31 degrees C resulted in a duration of depolarization that was equivalent in the 37 degrees C-10 min and 31 degrees C-14 min groups. However, hippocampal CA1 damage was not increased (31 degrees C-10 min = 4 +/- 1% dead neurons; 31 degrees C-14 min = 6 +/- 1% dead neurons, 95% CI, -1% to 3%; 37 degrees C-10 min = 90 +/- 17% dead neurons). CONCLUSIONS: Despite similar durations of DC depolarization, outcome in hypothermic rats was markedly improved compared with normothermic rats. This indicates that hypothermic neuroprotection can be attributed to mechanisms other than the delay in time to onset of ischemic depolarization.     

Influence of preconditioning on rat heart subjected to prolonged cardioplegic arrest

BACKGROUND: Ischemic preconditioning (IP) can reduce lethal injury to the myocardium induced by prolonged ischemia. However, little is known about the effect of preconditioning on the heart subjected to cardioplegic arrest and hypothermic preservation. We evaluated the effect of IP on myocardial metabolism, mechanical performance, and coronary endothelial function after cardioplegic arrest and prolonged hypothermic preservation. METHODS: An isovolumic Langendorff perfused rat heart model was used, and hearts were divided into two groups. The first group (IP, n = 14) was preconditioned by 5 minutes of global normothermic (37 degrees C) ischemia followed by 10 minutes of normothermic reperfusion before 6 hours of cold (4 degrees C) preservation, followed by 60 minutes of reperfusion. The second group (control, n = 15) was subjected to 6 hours of cold preservation followed by 60 minutes of reperfusion without preconditioning. Mechanical function was assessed using left ventricular balloon by constructing pressure-volume curves in two ways: at defined left ventricular volumes or at defined left ventricular end-diastolic pressures. Initially, the volume of the balloon was increased incrementally from 0 to 150 microL in 25-microL steps. Measurements were then repeated with loading balloon to achieve left ventricular end-diastolic pressure of 5, 10, 15, or 20 mm Hg. Myocardial function was assessed before ischemia and at 15 or 60 minutes of reperfusion. Metabolic status of the heart was evaluated by measuring the release of purine catabolites during the initial 15 minutes of reperfusion and concentrations of myocardial nucleotides at the end of reperfusion. Endothelium-mediated vasodilatation was evaluated using 10 mumol/L 5-hydroxytryptamine before and after ischemia. RESULTS: Left ventricular end-diastolic pressure values were significantly lower in the IP group, by 20% to 40%, during the reperfusion phase at each volume of the balloon compared with the control group. The rate-pressure product was more favorable during reperfusion in the IP than in the control group because of a 15% increased heart rate in the IP group. The release of purine catabolites from the heart during the reperfusion phase was reduced (p < 0.01) in the IP group (0.66 +/- 0.04 mumol) relative to the control group (0.92 +/- 0.06 mumol). No difference in the recovery of systolic function, myocardial adenosine triphosphate concentration, or endothelial function was observed between the groups. CONCLUSIONS: Under conditions of cardioplegic arrest and hypothermic preservation, IP can offer additional protection for the heart by preventing an increase in diastolic stiffness. However, metabolic improvement or better preservation of the systolic or endothelial function was not observed in this model.     

Effects of cold and warm blood cardioplegia assessed by myocardial pH and release of metabolic markers

The optimal temperature of blood cardioplegia remains controversial. Interstitial myocardial pH was monitored online with a probe that was inserted in the anterior wall of the left ventricle. Venous pH, lactate production, and creatine kinase and troponin T release were measured in coronary sinus blood obtained in 14 dogs after ischemic arrest periods of 5, 10, 20, and 40 minutes with warm (n = 7; mean myocardial temperature, 35 degrees +/- 2 degrees C) and cold (n = 7; mean myocardial temperature, 12 degrees +/- 1 degree C) blood cardioplegic protection. Blood cardioplegic solution was delivered at a rate of 100 mL/min during the 10 minutes between each ischemic arrest. The interstitial myocardial pH decreased significantly (p < 0.05) from 7.1 +/- 0.3 to 6.53 +/- 0.3 after ischemia in animals perfused with warm blood cardioplegia and from 7.04 +/- 0.3 to 6.64 +/- 0.1 in those receiving cold blood cardioplegic protection; however, the difference between the groups was not significant (p > 0.05). Lactate production and creatine kinase and troponin T release increased significantly after ischemia, but there was no difference in the changes between the warm and cold blood cardioplegia groups. In conclusion, ischemia caused significant changes in all variables measured, and these changes were directly proportional to the duration of ischemia. However, there was no significant difference (p > 0.05) in the myocardial metabolic changes between the warm and cold blood cardioplegia groups in terms of the duration of ischemic arrest studied.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of the no-flow interval and hypothermia on cerebral blood flow and metabolism during cardiopulmonary resuscitation in dogs

BACKGROUND and PURPOSE: We sought (1) to determine the effect of brief periods of no flow on the subsequent forebrain blood flow during cardiopulmonary resuscitation (CPR) and (2) to test the hypothesis that hypothermia prevents the impact of the no-flow duration on cerebral blood flow (CBF) during CPR. METHODS: No-flow intervals of 1.5, 3, and 6 minutes before CPR at brain temperatures of 28 degreesC and 38 degreesC were compared in 6 groups of anesthetized dogs. Microsphere-determined CBF and metabolism were measured before and during vest CPR adjusted to maintain cerebral perfusion pressure at 25 mm Hg. RESULTS: Increasing the no-flow interval from 1.5 to 6 minutes at 38 degreesC decreased the CBF (18. 6+/-3.6 to 6.1+/-1.7 mL/100 g per minute) and the cerebral metabolic rate (2.1+/-0.3 to 0.7+/-0.2 mL/100 g per minute) during CPR. Cooling to 28 degreesC before and during the arrest eliminated the detrimental effects of increasing the no-flow interval on CBF (16. 8+/-1.0 to 14.8+/-1.9 mL/100 g per minute) and cerebral metabolic rate (1.1+/-0.1 to 1.3+/-0.1 mL/100 g per minute). Unlike the forebrain, 6 minutes of preceding cardiac arrest did not affect brain stem blood flow during CPR. CONCLUSIONS: Increasing the no-flow interval to 6 minutes in normothermic animals decreases the supratentorial blood flow and cerebral metabolic rate during CPR at a cerebral perfusion pressure of 25 mm Hg. Cooling to 28 degreesC eliminates the detrimental impact of the 6-minute no-flow interval on the reflow produced during CPR. The brain-protective effects of hypothermia include improving reflow during CPR after cardiac arrest. The effect of hypothermia and the impact of short durations of no flow on reperfusion indicate that increasing viscosity and reflex vasoconstriction are unlikely causes of the "no-reflow" phenomenon.     

Protective efficacy of a hypothermic pharmacological agent in gerbil forebrain ischemia

BACKGROUND AND PURPOSE: The novel muscarinic cholinergic partial agonist U-80816E was tested in the gerbil brief bilateral carotid occlusion ischemia model based on the rationale that the compound's hypothermic properties might afford effective protection of the selectively vulnerable hippocampal CA1 region. METHODS: Male gerbils were subjected to either 10 or 15 minutes of bilateral carotid occlusion, followed by histopathological assessment of the CA1 neuronal survival 7 days later. RESULTS: In saline-treated animals, 10 minutes of bilateral carotid occlusion resulted in a 30.5% loss of CA1 neurons, whereas a 15-minute insult resulted in a 49.6% loss. Administration of U-80816E (6 mg/kg i.p. 30 minutes before bilateral carotid occlusion and again 2 hours after reperfusion) resulted in a significant protective effect of the CA1 neuronal population with either duration of ischemia; neuronal loss was reduced to 12.6% in the milder model (p < 0.05 versus saline-treated) and 24.9% in the more severe model (p < 0.04 versus saline). However, the 6 mg/kg i.p. dose of U-80816E was found to produce a 1.0 degree C decrease in brain temperature (measured with a tympanic temperature probe) at 10 minutes of ischemia compared with that of saline-treated gerbils. At 10 minutes of reperfusion, after the 10-minute episode of ischemia, the brain temperature of the U-80816E-treated gerbils was 2.2 degrees C lower than that of saline-treated animals. When the U-80816E-treated gerbils were subjected to either 10 or 15 minutes of ischemia but placed in a heated chamber that prevented the hypothermic effects, no cerebroprotection was observed. CONCLUSIONS: These results show that the anti-ischemic efficacy of U-80816E is mediated through its hypothermic properties, thus suggesting the feasibility of pharmacologically induced hypothermia as a cerebroprotective approach.     

Time window for the contribution of the delta-opioid receptor to cardioprotection by ischemic preconditioning in the rat heart

The present study aimed to examine (1) whether the role of the opioid receptor in ischemic preconditioning (PC) is consistent regardless of the duration of ischemic insult and (2) which opioid receptor subtype contributes to PC. In the first series of experiments, the effects of PC, a nonselective opioid receptor antagonist (naloxone), and their combination on the infarct size after various durations of ischemia were assessed. In anesthetized, open-chest rats, the coronary artery was occluded for 20, 30, or 40 minutes to induce infarction and was reperfused for 3 hours, PC was performed with two cycles of 5-minute ischemia followed by 5-minute reperfusion before the sustained ischemia. At 25 minutes before the ischemia, naloxone was injected alone or in combination with subsequent PC. Infarct size was determined by tetrazolium staining and was expressed as a percentage of the risk area size (%IS/RA). In the second series of experiments, the effects of a delta-receptor-selective antagonist, naltrindole (NTI), and a kappa-receptor selective antagonist, nor-binaltrophimine (nor-BNI), on PC before 30-minute coronary occlusion were assessed. In untreated controls, %IS/RA was 53.1 +/- 3.2 after 20 minutes, 67.9 +/- 3.9 after 30 minutes, and 87.8 +/- 2.0 after 40 minutes of ischemia, respectively. PC significantly reduced %IS/RA after 20, 30, and 40 minutes of ischemia to 3.1 +/- 0.8, 12.8 +/- 1.1, and 42.1 +/- 4.3, respectively (P < 0.05 vs. each control). Naloxone (6 mg/kg) partially attenuated the protection afforded by PC when the sustained ischemia was 30 minutes (%IS/RA = 27.4 +/- 4.5; P < 0.05 vs. PC), but this inhibitory effect of naloxone was not detected when the duration of the ischemia was 20 or 40 minutes. NTI (10 mg/kg) also attenuated infarct size limitation by PC after 30 minutes of ischemia (%IS/RA = 25.6 +/- 3.7), but nor-BNI (10 mg/kg) failed to modify infarct size limitation by PC (%IS/RA = 13.3 +/- 3.2). The present results suggest that activation of the opioid delta-receptor partly contributes to preconditioning against infarction in the rat and that there may be a time window (at around 30 minutes after the onset of ischemia) for this opioid receptor-mediated protective mechanism.