Effect Of Anticoagulation With Heparin In Advance On Cerebral Resuscitation In The Gerbil Suffered Bilateral Common Carotid Artery Occlusion And The Rat Post Cardiopulmonary Resuscitation

BackgroundAfter cardiac arrest(CA) of 5 to 10 minutes, a marked activation of blood coagulation occurs and the presence of microthrombiare found in cerebral vessels.These micro-circulatory disturbances directly affect the outcome on cardiopulmonary resuscitation(CPR). Therefore, this study involved the comparison of the effect of anticoagulation and non-anticoagulation to prove the beneficial effect of thrombolysis treatment by using two types animal experiment. Thus, the study was carried out on the experiment one: comparison of the effect of anticoagulation on cerebral resuscitation after brain ischemia induced by occlusion of bilateral common carotid artery between two groups in gerbilles, and the experiment two: comparison of the effect of anticoagulation on cardiopulmonary-cerebral resuscitation(CPCR) after asphyxial CA in rats between two groups.Materials and MethodsThere were two experiments in this study, including experiment one(whole brain ischemia by clamping bilateral common carotid artery in gerbils) and experiment two(CPCR after asphyxial CA in rats).Experiment one: Matured gerbils were randomly divided into normal control group(groupâ… , had 10 gerbiles without anesthesia and operation), a sham group(groupâ…¡, had 10 gerbiles with anesthesia and operation), non-anticoagulation group(group â…¢, had 30 gerbiles with anesthesia and operation and whole brain ischemia induced by clamping of bilateral common carotid artery), and anticoagulation group(groupâ…£, had 30 rats with anesthesia, operation and heparinization before whole brain ischemia). The animal in groupsâ…¡,â…¢andâ…£were anesthetized with pentobarbital sodium 40mg/kg through intraperitoneal injection. After the anesthesia took full effect, a surgical incision was made at the middle line in the neck at supine position and the bilateral common carotid arteries were carefully isolated without damage of the vessels and nerve fibre and an intravenous cannula was put in at one side of the neck venous in all the animals of these three groups. The gerbilles which were assigned to groupâ… were assessed for the neurological deficit(ND) scores after selected for the test and before the fixation of brain for histological exams. The animal in groupâ…¡was given normal saline 0.5ml after awakening from anesthesia then removed the cannula, ligated the venous and sutured the incision. And the ND scores were evaluated before experiment and the fixation of brain for histological exams. After awakening from anesthesia, the bilateral common carotid arteries were blocked with bulldog clamp for 10 minutes in the animals of groupâ…¢. After clamping for 10 minutes, epinephrine 0.01mg/kg was administrated and the arteries re-opened to supply blood to brain by unclamping the bulldog clamp. In groupâ…£, except for administrating 50IU/100g after the animal awakening from anesthesia and before clamping the arteries, all the other procedures were same as groupâ…¢.In this experiment the time intervals from whole brain ischemia to coma and dyspnea, and from unclamping of common carotid artery to regaining of consciousness, righting reflex, movement on limbs, foraging, and stable movement on limbs were studied during the experiment. Neurological deficit(ND) scores of four groups before experiment, and at 20 min,60 min,24h,48h between groupsâ…¢andâ…£after unclamping of common carotid artery and at 72h of four groups were compared(at 72h,the ND scores were compared between groupsâ… andâ…¡, and groupsâ…¢andâ…£,(ND scores 0%=best; 100%=worst). Mortality of the animals between groupsâ…¢andâ…£ during the experiment was compared. Positive expression rate of TNF-aand IL-1βin hippocampus region(CA1～3)and brain histopathological damage (HD) scores of ischemic neurons in five regions between groupâ…¢andâ…£were compared after 72 hours of unclamping of common carotid artery.Experiment two: Male Sprague-Dawley rats were randomly divided into four groups. Based on study protocol, normal control group (groupâ… , had 10 rats), a sham group (groupâ…¡, had 10 rats), non-anticoagulation arrest group (groupâ…¢, had 25 rats), and anticoagulation arrest group (groupâ…£, had 25 rats) were studied. Groupâ… had no anesthesia and surgery, groupâ…¡had anesthesia and surgery but no asphyxia. Groupâ…¢had apneic asphyxia of 8 min. Groupâ…£was the same as groupâ…¢except that heparin 50IU/100g was administered before CA. All rats in groupâ…¢andâ…£were given epinephrine and standard external cardiopulmonary resuscitation to restore spontaneous circulation (ROSC).For the animals in groupâ…¡,â…¢andâ…£, anesthesia was induced with pentobarbital sodium 45mg/kg injected intraperitoneally. At the onset of unconsciousness and relaxation without lash reflex, the rats were immediately intubated with a 16-G plastic catheter by direct laryngoscope. The tracheal cannula was connected to rodent piston ventilator; end-tidal partial pressure of carbon dioxide (P_EtCO₂) and inhaled oxygen concentration were monitored The ventilator was initially preset to deliver tidal volumes of 1ml/100g, a positive end-expiratory pressure of 3 cm H₂O, and at a ventilation rate of 40-50/min. Mechanical ventilation was later adjusted to control P_EtCO₂ between 30 to 40mmHg. The fraction of inspired oxygen (FiO₂) was maintained at 0.70 to 0.85. Electrocardiograph (ECG) electrodes were attached to monitor leadâ…¡ECG waves, and heart rate (HR). A temperature (RT) probe was inserted into the rectum of the rats up to a depth of 1 to 1.5cm to monitor temperature changes. The rats were placed on a heating pad and secured to a surgical board with adhesive tape. The RT was controlled throughout the experiment at a normal range of between 37.8 to 38.7℃by using the heating system inside the operation table and the aid of a lamp. Under aseptic technique an incision was made in the right groin. 24-G catheters (length, 5.5 cm) were advanced into the abdominal aorta and inferior vena cava through femoral artery and vein, and connected to an arterial pressure transducer and intravenous infusion pump respectively. An intravenous dose of vecuronium 2mg/kg was then administered. The mean arterial blood pressure (MABP), RT, P_EtCO₂ and HR were recorded at 10-15 min before the ischemic insult and after ROSC at 10, 30 and 60 minThe differences of MABP, HR, RT and end-tidal partial pressure of carbon dioxide (P_EtCO₂) of baseline values among groupâ…¡,â…¢andâ…£, and at time points: immediately after ROSC, 10 min after ROSC, 30 min after ROSC, 60 min after ROSC between groupâ…¢andâ…£, were compared. The time intervals from asphyxia to CA, CPR to ROSC, and after stopping mechanical ventilation to extubation between groupâ…¢andâ…£were compared. Achievement ratio of CPR and 72 hours survival rate after CPR were compared between groupâ…¢andâ…£. Neurological deficit (ND) was compared between groupsâ…¢andâ…£before CA and after CPR at 2h,24h,48h,and 72h (0%=best; 100%=worst). Positive expression rate of TNF-aand IL-1βin hippocampus region(CA1～3)and brain histopathological damage (HD) of ischemic neurons in five regions between groupâ…¢andâ…£were compared after 72 hours of CPR.ResultsThe results of experiment one: The time intervals from whole brain ischemia to coma and dyspnea,and from unclamping of common carotid artery to regaining of consciousness, righting reflex, movement on limbs, foraging, and stable movement on limbs between groupsâ…¢andâ…£were significantly different, P＜0.01. The difference of the ND scores between groupâ…¢andâ…£was detected, P＜0.01.There were significant differences of the mortality, HD scores and the positive expression rate of TNF-aand IL-1βbetween groupsâ…¢andâ…£, P＜0.05 or 0.01.The results of experiment two: After asphyxiation of 8 min of the rats in groupâ…¢andâ…£, cardiac arrest of 4 to 5 min was obtained. The differences of MABP, HR, RT and P_EtCO₂ of baseline values among groupâ…¡,â…¢andâ…£were not significant. Only the difference of MABP between groupâ…¢andâ…£was found at immediately after ROSC,P＜0.01. The rest parameters had no significant difference between groupâ…¢andâ…£at the time points: immediately after ROSC, 10 min after ROSC, 30 min after ROSC, 60 min after ROSC. The time intervals from asphyxia to CA, CPR to ROSC, and from weaning off mechanical ventilation to extubation between groupâ…¢andâ…£, the achievement ratio of CPR and the survival rate at 72 hours, and the ND and HD between groupâ…¢andâ…£were significantly different, P＜0.05 or 0.01. The positive expression rates of TNF-a and IL-1βbetween groupsâ…¢andâ…£were statistically different, P＜0.05 or 0.01.ConclusionIt appears that anticoagulation with heparin may obviously improve the outcome and prognosis of brain ischemia induced by clamping of bilateral common carotid artery in for 10 minutes, including improvement of neurological function recovery, prevention of morphological damage of brain and immuno-histochemistry injury. And also it shows that anticoagulation with heparin may increase the achievement ratio of CPR after cardiac arrest and survival rate of 72 hours after CPR in rats. At the same time the recovery of neurological function is improved, and the morphological damage of brain and immuno-histochemistry injury are prevented. Therefore the future clinical studies may be carried out to evaluate the relevance of administration of heparin as early as possible during CA and even before administration adrenaline for CPR.