Euthanasia of mouse fetuses and neonates.

We sought to determine whether any of the common methods of euthanasia for adult rodents would lead to an acceptable death for fetuses or neonates. We wanted to identify a method that was rapid, free of signs of pain or distress, reliable, and minimally distressful to the person performing the procedure and that minimized the amount of handling required to perform the procedure. We evaluated six methods of euthanasia, with and without anesthesia, in three age groups of mice: gravid mice (E14-20) and neonatal pups (P1-P7 and P8-P14). Euthanasia methods included: halothane inhalation, carbon dioxide inhalation, intraperitoneal sodium pentobarbital, intravenous potassium chloride, and cervical dislocation with and without anesthesia. Noninvasive echocardiography was used to assess heartbeat during euthanasia. With cardiac arrest as the definition of death, no method of euthanasia killed fetal mice. Halothane inhalation (5% by vaporizer) was not an acceptable method of euthanasia for mice of the age groups tested. Intraperitoneal administration of sodium pentobarbital for euthanasia required a higher dose than the previously established dose, and there is a risk of reduced efficacy in pregnant animals due to potential intrauterine injection. Carbon dioxide asphyxiation was the most efficient method of euthanasia for neonatal mouse pups P1-14. For pregnant adult mice, intravenous potassium chloride under anesthesia, carbon dioxide asphyxiation, and cervical dislocation alone or under anesthesia were excellent methods of euthanasia.     

Euthanasia of Neonatal Rats with Carbon Dioxide

Exposure to CO₂ is a common method used to euthanize rodents in biomedical research and rodent production. The purpose of this study was to determine the length of CO₂ exposure required to euthanize neonatal rats (0 to 10 d old). Multiple groups of rats were exposed to 100% CO₂ for 5 to 60 min. After CO₂ exposure, rats were placed in room air for 20 min to allow for possible recovery. No difference was found in comparing 1 inbred strain and 1 outbred stock of rats. Time to death varied inversely with the age of the animals, requiring as long as 35 min on the day of birth. The time to death decreased steadily with increasing age, with 100% of the rats euthanized after 5 min of CO₂ exposure at 10 d of age. The time required for 100% mortality decreased by 3 min for every 1 d increase in age between days 0 and 10.Because laboratory rodents are the most common animals used in research today, optimizing the methods used for euthanasia of these animals is important to researchers, veterinarians, and care staff. Unlike many other laboratory animals, rodents often are bred in research facilities. Breeding may require euthanasia of very young animals, most frequently because of undesired phenotype or genotype or the production of surplus animals. Although many studies have evaluated the ethical and physiologic considerations of various euthanasia methods on adult laboratory rodents, the literature is relatively sparse on the euthanasia of neonatal rodents.Because most research institutions do not have to euthanize large numbers of neonates at one time, the limited research on the euthanasia of neonatal rodents is unsurprising. However, this paucity leaves rodent producers and others who must euthanize groups of neonatal rodents at a loss for best practice guidelines. The American Veterinary Medical Association (AVMA) Panel on Euthanasia defines “acceptable” methods of euthanasia for rodents and other small mammals as: barbiturates, inhalant anesthetics, carbon monoxide, potassium chloride in conjunction with general anesthesia, microwave irradiation, and CO₂.² For some of these methods, their use to ensure the humane death of large numbers of neonatal rats would present several challenges. Ideally, the method chosen would minimize distress to the animals, would have a minimal health impact on personnel performing the euthanasia, be easy to administer, lack complicated record-keeping or disposal requirements, not be distressing to personnel performing or witnessing euthanasia, and be cost-effective to employ for large numbers of animals. In addition, an ideal euthanasia method would leave the euthanized animals suitable for other uses, whether for research or as food for rodent-eating animals. None of the methods approved by the AVMA meet all these criteria, but CO₂ is the closest. European Union recommendations for the euthanasia of neonatal rodents include decapitation, concussion, rapid freezing, and hypothermia;⁷ of these methods, only rapid freezing could meet the criteria just listed, but hazards are associated with the use of liquid nitrogen. In addition, immersion in liquid nitrogen is not recommended as an euthanasia method in the United States, unless the animals are first rendered insensible.^2,15 In many cases, the method used prior to freezing or hypothermia is CO₂ inhalation. Carbon dioxide is widely accepted as an euthanasia agent due to the quick onset of effects, including unconsciousness,^4,5,16,29 minimal tissue artifact production,^12,15 and ease of use.¹⁴The use of CO₂ for euthanasia of adult rodents is the subject of much opinion and debate.^3,20,22 The current study was not intended to evaluate the merits of using CO₂ to euthanize laboratory animals, but rather to determine the parameters of its use in the euthanasia of neonatal rats. The aim was to determine the length of exposure to 100% CO₂ necessary to euthanize 100% of rats 0 to 10 d of age.     

大鼠脑损伤对移植人胚神经干细胞存活和分化的影响

目的 探讨大鼠脑液压冲击伤(FPI)后对植入的人胚神经干细胞(HNSCs)存活和分化的影响。方法 取8周龄人胎儿大脑皮层细胞，体外培养获得神经干细胞(NSCs)，巢蛋白免疫荧光染色；SD大鼠制作FPI模型．于伤后24h在损伤区移植标有5-溴脱氧脲嘧啶(BrdU)的HNSCs，1周和4周后处死大鼠，邻片行BrdU／微管相关蛋白-2(MAP-2)和BrdU胶质纤维酸性蛋白(GFAP)免疫组织化学双染。结果 HNSCs移植后1周可见BrdU阳性细胞向周围迁移，且BrdU／MAP-2双阳性细胞多于BrdU GFAP双阳性细胞；移植后4周BrdU阳性细胞迁移的范围更广，但细胞数量明显减少，脉络丛和微血管中可见BrdU阳性细胞．BrdU／GFAP双阳性细胞多于BrdU／MAP-2双阳性细胞。结论 HNSCs能存活于损伤区域，移植后逐渐分化为星形胶质细胞．且易被内皮吞噬细胞吞噬。     

Euthanasia Method for Mice in Rapid Time-Course Pulmonary Pharmacokinetic Studies

To develop a means of euthanasia to support rapid time-course pharmacokinetic studies in mice, we compared retroorbital and intravenous lateral tail vein injection of ketamine–xylazine with regard to preparation time, utility, tissue distribution, and time to onset of euthanasia. Tissue distribution and time to onset of euthanasia did not differ between administration methods. However, retroorbital injection could be performed more rapidly than intravenous injection and was considered to be a technically simple and superior alternative for mouse euthanasia. Retroorbital ketamine–xylazine, CO₂ gas, and intraperitoneal pentobarbital then were compared as euthanasia agents in a rapid time-point pharmacokinetic study. Retroorbital ketamine–xylazine was the most efficient and consistent of the 3 methods, with an average time to death of approximately 5 s after injection. In addition, euthanasia by retroorbital ketamine–xylazine enabled accurate sample collection at closely spaced time points and satisfied established criteria for acceptable euthanasia technique.Matching the attributes of the euthanasia method to different applications and study designs is an important consideration in selecting the euthanasia method for an in vivo study. Methods of euthanasia should adhere to the AVMA Guidelines on Euthanasia, ACLAM Task Force Guidelines on Euthanasia, and other references reiterating similar principles.^1,3,7,11,16, According to these guidelines, an acceptable euthanasia method is characterized by: (1) rapid loss of consciousness; (2) reliability; (3) safety of personnel; (4) irreversibility; (5) compatibility with study requirements; (6) minimal negative emotional effect on observers and personnel; and (7) compatibility with subsequent evaluation, examination, or use of tissue sample.^1,7 The purpose of the current set of studies was to compare commonly accepted means of euthanasia in mice with a novel method: retroorbital ketamine–xylazine euthanasia.Ketamine–xylazine is a commonly used combination for anesthesia and euthanasia in mice.^4,14,28 In our experience, ketamine–xylazine is most often given intraperitoneally as an anesthetic combination. When used for euthanasia purposes, typically an overdose of the anesthetic is administered intraperitoneally followed by a secondary means of euthanasia, such as exsanguination, thoracotomy, or cervical dislocation.For intravenous drug administration in mice, the retroorbital injection method is a technically simple, easily learned, reproducible, and rapid procedure, particularly as compared with intravenous tail vein dosing. Retroorbital injection has been shown to be interchangeable with the intravenous tail vein injection technique when parenteral access is desired in the mouse.^{9,10,12,18,20,22,29} The variability, technical demand, and other negative aspects of intravenous tail vein dosing in mice make the retroorbital method desirable.^{9,10,12,18,20,22,29} To minimize any potential associated pain or distress, retroorbital injections typically are given to anesthetized mice.¹⁵ This practice is feasible when mice are intended to recover after the injection; however, use of the retroorbital technique for euthanasia has not been documented. One goal of the current study was to demonstrate the adherence of retroorbital injection of ketamine–xylazine to the previously stated principles regarding euthanasia, with emphasis on the humaneness of the technique.We developed the retroorbital ketamine–xylazine euthanasia technique to support rapid time-course mouse pulmonary pharmakokinetics studies in drug development. For these types of studies, which involve direct delivery of compounds to the lungs, intratracheal dosing is often the preferred method because of its reproducibility, reliability, and translatability to the clinic setting.^{19,21,23,24,26,27} One key factor that affects the efficacy and potency of these drug candidates is their residence time in the lungs. Pharmacokinetics studies focus on the distribution, clearance, and metabolism of chemical or drug entities that are introduced into the body. Pulmonary pharmacokinetics parameters are often assessed in serum, lung tissue, and bronchoalveolar lavage fluid. Because of rapid local clearance of the compounds, the quality of these data relies on the precision of sample collection, particularly from early time points that often are within minutes of each other.When pulmonary pharmacokinetics studies are performed in mice, groups of animals are euthanized at specific time points after dosing to enable collection of tissue samples for concentration measurements over a time course. The method of euthanasia chosen for these studies must be nontraumatic and incorporate the attributes of rapid onset, ease of execution, reproducibility, and the ability to preserve tissues and samples. Currently, pulmonary pharmacokinetics studies use a variety of euthanasia techniques, including CO₂ exposure, intraperitioneal barbiturate overdose, cervical dislocation, and decapitation.^13,19,23,26 These methods, when used in rapid time-point pharmacokinetics studies, have attributes that can confound the results.^6,8,11,17 Cervical dislocation and decapitation result in rapid death but are traumatic in nature. These techniques damage the trachea and cervical region, making it difficult or impossible to lavage the lungs after the procedure. These methods also confound the results by causing hemorrhage into various tissues and contaminating the lung tissue. Other euthanasia methods, such as CO₂ exposure and intraperitoneal pentobarbital overdose are less traumatic, but the time to death is delayed and variable, thus preventing precise timing for tissue harvest.^2,5,8,11,17 Therefore, the goals of this study were to examine the utility of a novel method of euthanasia, retroorbital administration of ketamine–xylazine for euthanasia of mice. We here demonstrate its positive effect on the quality of in vivo data, and show that retroorbital administration of ketamine–xylazine meets the criteria for acceptable euthanasia. Retroorbital administration of ketamine–xylazine for euthanasia of mice is ideal for pulmonary pharmacokinetics studies and for any study needing a rapid, nontraumatic means of euthanasia.     

Assessing cervical dislocation as a humane euthanasia method in mice

Research investigators often choose to euthanize mice by cervical dislocation (CD) when other methods would interfere with the aims of a research project. Others choose CD to assure death in mice treated with injected or inhaled euthanasia agents. CD was first approved for mouse euthanasia in 1972 by the AVMA Panel on Euthanasia, although scientific assessment of its humaneness has been sparse. Here we compared 4 methods of spinal dislocation--3 targeting the cervical area (CD) and one the thoracic region--in regard to time to respiratory arrest in anesthetized mice. Of the 81 mice that underwent CD by 1 of the 3 methods tested, 17 (21%) continued to breathe, and euthanasia was scored as unsuccessful. Postmortem radiography revealed cervical spinal lesions in 5 of the 17 cases of unsuccessful CD euthanasia. In addition, 63 of the 64 successfully euthanized mice had radiographically visible lesions in the high cervical or atlantooccipital region. In addition, 50 of 64 (78%) mice euthanized successfully had radiographically visible thoracic or lumbar lesions or both. Intentionally creating a midthoracic dislocation in anesthetized mice failed to induce respiratory arrest and death in any of the 18 mice subjected to that procedure. We conclude that CD of mice holds the potential for unsuccessful euthanasia, that anesthesia could be valuable for CD skills training and assessment, and that postmortem radiography has minimal promise in quality-control assessments.     

Evaluation of Rapid Cooling and Tricaine Methanesulfonate (MS222) as Methods of Euthanasia in Zebrafish (Danio rerio)

Despite the progressively increasing use of zebrafish (Danio rerio) in research, the most humane method of euthanasia for these fish has not been determined. Contemporary guidance documents state that hypothermia is an unacceptable method of euthanasia. The goal of this study was to compare rapid cooling and tricaine methanesulfonate (MS222) for zebrafish euthanasia. Zebrafish (n = 46) were euthanized by immersion in either an ice-water (4 °C or less) bath or unbuffered MS222 solution (250 mg/L; 25 to 30 °C). Another cohort (n = 10) was exposed to buffered MS222 to determine whether the acidity of the water alone caused distress. The times from exposure until the animals became unable to swim, right themselves, and death were measured, and signs of distress were recorded. Fish then were placed in a ‘recovery tank’ of system water to verify that recovery did not occur. Tissues were examined histologically. The mean time for euthanasia was longer and the number of fish exhibiting signs of distress was greater for fish exposed to MS222 than those exposed to chilled water. In addition, 4 of the 23 fish exposed to MS222 regained consciousness in the recovery tank, whereas none of 23 fish exposed to chilled water recovered. No histopathologic changes or evidence of ice crystal formation were seen in either group. In light of the faster time to death and fewer signs of distress in zebrafish euthanized by rapid cooling, we advocate this method as a humane veterinary practice.Abbreviation: MS222, tricaine methanesulfonateIn contemporary biomedical research facilities, the use of zebrafish (Danio rerio) has progressively increased in recent years.³ With this increase has come an intensified effort to identify the ideal husbandry, care, and management parameters for this species. One important, yet controversial, issue has involved the selection of humane euthanasia methods for zebrafish. Zebrafish are considered eurythermic animals as they are adaptable to a wide range of temperatures.⁵ The most commonly used maintenance temperature for zebrafish is 28.5 °C (83 °F), although temperatures between 24 and 30 °C (75 and 86 °F) have been recommended.^7,13 Following periods of acclimation, zebrafish can tolerate a much broader temperature range.⁵ However, acute exposure to temperatures below their thermal neutral zone can cause death in zebrafish due to their inability to quickly acclimate. This natural phenomenon has been used as a method of euthanasia in zebrafish, but the AVMA Guidelines on Euthanasia¹ and the report Recognition and Alleviation of Pain and Distress in Laboratory Animals⁸ from the Institute for Laboratory Animal Research both state that hypothermia (also referred to as rapid cooling) is unacceptable as a method of euthanasia for fish. Although these reports provide no scientific explanation regarding why rapid cooling is considered unacceptable, some speculate that ice crystal formation occurs in tissues during rapid cooling. Currently, the AVMA recommends the use of tricaine methanesulfonate (MS222), decapitation, or injectable methods such as sodium pentobarbital for fish euthanasia.¹ These recommendations appear more applicable to larger, nontropical species of fish, because decapitation and sodium pentobarbital injection are impractical for euthanasia of small fish, such as zebrafish. MS222 may be a favored method of euthanasia because, according to 1 set of guidelines, it “causes no signs of stress, such as elevation of blood glucose, cortisol, or catecholamines.”⁴ However, this statement was not supported by published scientific data.In mammals, exposure to cold temperatures may result in anesthesia. In humans, exposure to 9 °C (48 °F) is predicted to result in complete nerve conduction blockade. In cats, 20 °C (68 °F) results in the inability of evoked potentials in the central nervous system, and goats at the same temperature do not react to painful peripheral stimuli in the absence of general anesthesia.⁶ Physiologically, peripheral nerve conduction velocity is highly correlated with temperature, that is, as temperature decreases, nerve conduction velocity also decreases.⁶ In reptiles and amphibians also, peripheral nerve conduction velocity decreased with decreasing temperature, suggesting that exposure of these species to cold temperatures is anesthetic.⁶ Other teleost fish actually lack receptors that respond to cold and likely do not experience pain associated with cold.² Although these studies provide valuable insights into cold exposure in other poikilotherms, one cannot draw direct correlations specific to fish, especially tropical species, from these data.Rapid cooling affords several advantages as a method of zebrafish euthanasia. These include the ability to euthanize many animals simultaneously, minimization of handling of individual animals (which is necessary when using injectable agents or decapitation), minimal risk of operator error when preparing the euthanasia bath, and reduction in occupational health and safety risk to personnel associated with chemical and physical methods of euthanasia. The goal of our study was to compare the effects of rapid cooling and MS222 (as described by the AVMA Guidelines on Euthanasia) as methods of zebrafish euthanasia.¹ We hypothesized that rapid cooling in zebrafish would result in a shorter time between exposure and death than with MS222, would not cause ice crystallization in tissue, and would result in minimal signs of distress. This study is the first to compare these 2 euthanasia methods for zebrafish, and the outcome will be important in assisting institutional animal care and use committees and researchers in the determination of the most appropriate method of euthanasia for zebrafish.     

实验性大鼠脑损伤后介质白三烯C_4的变化在继发性脑损害中的作用

目的探讨实验性大鼠脑损伤后,炎性介质白三烯C4(LTC4)的变化及在继发性脑损伤中的作用。方法应用流体冲击装置致大鼠脑损伤模型,应用放射免疫分析法检测大脑皮层LTC4的含量变化,采用氢清除法定量监测皮层血流量的相应改变,并用木条行走作业试验行大鼠神经功能观察,并用5-脂加氧酶抑制剂菲尼酮阻止LTC4产生。结果流体冲击致大鼠脑损伤后,冲击侧皮层LTC4含量在伤后30min、1h显著增加,同时皮层血流量均显著下降,伤后5d内大鼠完成木条行走作业的能力明显受损。给予菲尼酮预处理后,皮层中LTC4明显减少,并伴有局部血流量增加,且大鼠完成木条行走作业恢复正常的时间缩短。结论流体冲击致大鼠脑损伤后冲击侧皮层中LTC4的产生明显增加,不仅加重了皮层血循环障碍,同时也直接或间接参与了继发性脑损伤过程。     

Fos蛋白和Jun蛋白在侧位液压冲击脑损伤大鼠大脑皮质的表达

目的：研究大鼠中度（０．２ＭＰａ）侧位液压冲击脑损伤时大脑皮质立即早期基因ｃ－ｆｏｓ和ｃ－ｊｕｎ表达产物Ｆｏｓ蛋白和Ｊｕｎ蛋白的变化规律。方法：雄性ＳＤ大鼠,随机分为正常对照物、手术对照组和损伤组。损伤组动物均给以０．２ＭＰａ液压冲击脑损伤,按冲击后处死时间不同再分为５、１５、３０、６０、１２０、２４０、４８０和７２０分钟组。应用免疫组织化学方法观察Ｆｏｓ和Ｊｕｎ蛋白在大脑皮质的表达特点。结果：冲击后３０分钟,双侧大脑皮质Ｆｏｓ阳性细胞数逐渐增多,冲击后７２０分钟达高峰。Ｆｏｓ阳性细胞面积在冲击后１２０分钟逐渐增大,７２０分钟达高峰;冲击后６０分钟双侧大脑皮质Ｊｕｎ阳性细胞数逐渐增多。冲击后１２０分钟阳性细胞面积逐渐增大,冲击后７２０分钟阳性细胞数和面积均达高峰。结论：中度侧位液压冲击脑损伤后Ｆｏｓ蛋白和Ｊｕｎ     

液压冲击脑损伤大鼠海马BDNF mRNA表达实验研究

目的：研究脑损伤后脑脑源性神经营养因子（BDNF）mRNA表达及其时序性变化,进一步研究脑损伤的分子机制。方法：用免疫组织化学及原位杂交方法,观察液压冲击脑损伤大鼠海马BDNF mRNA受伤后不同时间（5、15、30分钟和1、3、6、12小时及1、3、7日）的表达情况,以正常大鼠及手术大鼠作为对照。结果：正常及手术对照组大鼠脑组织海马表达少量的BDNF;脑损伤后1小时海马齿状回CA2及CA3细胞表达增强,3－6小时达高峰,12小时开始下降,24小时后降至对照水平。1、3、6和12小时与正常及手术对照组比较均有显著性差异。结论：液压冲击脑损伤可诱导BDNF基因的表达,BDNF mRNA表达存在时间差异,提示机体对脑损伤存在自身保护机制。     

大鼠颅脑外伤后突触素P38在中枢神经系统的表达变化

目的观察大鼠颅脑外伤后突触素p38在中枢神经系统表达的变化。方法将SD大鼠分为正常对照组、假手术对照组和损伤组,损伤组用侧位液压冲击法应用不同压力制成颅脑损伤动物模型,于损伤后不同时间点采集标本,采用免疫组织化学技术,检测大鼠损伤灶周边皮层及海马CA1区p38的表达变化,结果采用方差分析和q检验进行统计学分析。结果创伤性颅脑外伤后,损伤灶周边皮层及海马CA1区p38免疫阳性表达增强。结论中枢神经系统损伤后,突触素p38表达增强,且损伤越重,表达增强越明显。     

Characterization of the alteration of nutritional state in brain injury induced by fluid percussion in rats

Objective Patients suffering from traumatic brain injury (TBI) undergo rapid weight loss with negative nitrogen balance and enhanced whole-body protein breakdown, with protein wasting causing morbidity and increased mortality. Many experimental models of TBI have been used to evaluate strategies to improve the outcome of these patients, but nutritional status has not been considered in experiments published to date, although this may have great importance and influence the results obtained with TBI models. This study characterized the hypercatabolism level and nutritional status of TBI rats.Design Twenty-four male Wistar rats were randomized into three groups. Rats from the TBI group were anesthetized and fluid percussion was applied. The pair-fed (PF) group was healthy but was pair-fed to the TBI group. The ad libitum (AL) group was healthy and fed ad libitum. The study was performed over 10 days post-TBI.Measurements and results TBI in rats was characterized by remarkable long-lasting anorexia, renal failure (creatinine clearance: AL 1.8±0.2 and PF 1.5±0.1 vs. TBI 0.9±0.1 l/24 hour), anorexia (appetite depressed throughout the study), increased myofibrillar proteolysis (3-methylhistidine/creatinine ratio (day 2: AL 36±1 and PF 38±2 vs. TBI 54±5 µmol/mmol), and intestinal atrophy (ileum: AL 29.3±2.5 and PF 28.7±1.1 vs. TBI 22.5±1.4 mg/cm). In addition, anorexia led to muscular atrophy and decreased nitrogen balance. The metabolic alterations described above can increase morbidity and mortality.Conclusions TBI by fluid percussion in rats is a model reproducing the metabolic and nutritional alterations observed in clinical practice and is suitable for further studies exploring the efficacy of optimized nutritional support.     

自然衰老大鼠脑线粒体和血小板膜外周型苯二氮 受体结合活性的变化

目的以3月龄青年大鼠为对照,观察24月龄自然衰老大鼠大脑皮层线粒体和血小板膜外周型苯二氮 受体(PBRs)结合数量及亲和力的改变。方法雄性SD大鼠断头取脑,以差速离心法提取大脑皮层线粒体,低渗溶血法制备外周血小板膜。采用放射配基［³H］PK11195单点结合实验测定PBRs特异结合活性,通过受体饱和实验和Scatchard作图得到最大结合容量Bmax值和平衡解离常数Kd值。结果与3月龄组大鼠相比,24月龄组的大脑皮层线粒体及外周血小板膜PBRs特异结合活性均显著上升(P<0-001)。饱和实验结果表明,24月龄组大鼠大脑皮层线粒体PBRs的受体容量(Bmax)升高而受体亲和力明显下降,Kd值显著增高(P<0-001)。结论老龄鼠PBRs容量上升而亲和力下降,可能参与脑老化进程。外周血小板膜［³H］PK11195结合活性与脑内该指标的变化一致。     

Management of lacerations in the emergency department

The goals of wound management are simple: avoid infection and achieve a functional and aesthetically pleasing scar. This is achieved by reducing tissue contamination, debriding devitalized tissue, and restoring perfusion in poorly perfused wounds, in conjunction with a well-approximated skin closure.     

Attitudes of Tennessee physicians toward euthanasia and assisted death

BACKGROUND: Although many studies of euthanasia and physician-assisted death (PAD) have been performed in the United States, none have specifically addressed attitudes among physicians practicing in Tennessee. METHODS: In January 2001, we mailed a 30-item survey instrument to a stratified random sample of 1,117 physicians drawn from the Tennessee Licensing Bureau. RESULTS: Tennessee physicians are highly polarized over the issues of euthanasia and assisted death. A slight majority (47%) did not favor euthanasia or PAD and would oppose the legalization of such procedures. Of the physicians supporting euthanasia or PAD (43%), only 25% would administer a lethal overdose and less than a third would counsel/prescribe medication for an overdose. Attitudes were influenced by three primary factors: ethics, religion, and the role of the physician to relieve pain and suffering. CONCLUSION: Regardless of their overall position, the majority of physicians agreed on basic restrictions and safeguards to prevent abuses and to protect vulnerable patients.     

大鼠创伤性脑损伤早期基因差异表达的研究

目的研究创伤性脑损伤早期基因表达谱与正常脑组织基因表达谱的差异,以期阐明脑损伤后早期基因表达的改变规律,阐明脑损伤发生发展的分子机制,从而为临床治疗提供帮助,同时为法医损伤时间推断研究寻找标志物提供帮助。方法以大鼠自由落体损伤模型为对象,从损伤区脑组织和假手术对照组脑组织分别提取mRNA,经反转录成cDNA后与含有4096个随机基因的基因表达谱芯片杂交,杂交后的芯片经扫描仪扫描,并用GenePix3.0软件分析结果。结果发现有124个差异表达基因或表达序列标签（expression sequencetags,ESTs）;其中有46个基因和26个EST表达下调;28个基因和24个EST表达上调;在这些表达有差异的基因中,有涉及细胞内信号传导、神经递质释放、参与炎症的蛋白、离子通道及其受体蛋白和参与炎症反应的蛋白等被发现。结论创伤性脑损伤的发生发展涉及多个基因的改变;研究一个或少数几个基因很难解释其损伤后分子变化机制;基因芯片是研究颅脑损伤这种多基因改变、多因素作用的理想工具。     

Somatostatin causes vasoconstriction, reduces blood flow and increases vascular permeability in the rat central nervous system.

Using radiolabeled microspheres, spinal cord blood flow was measured after spinal subarachnoid injections of 3.1- to 12.5-nmol doses of somatostatin through either indwelling i.t. catheters or acutely inserted intervertebral needles. With either injection technique, somatostatin caused significant dose-dependent reductions in thoracic and lumbosacral blood flow that could be partially blocked by a 5-min preinjection of the somatostatin receptor antagonist cyclo[7-aminoheptanoyl-Phe-D-Trp-Lys-Thr(Bzl)], which has previously been shown to block the hindlimb flaccidity produced by these doses of somatostatin in conscious rats. The duration of these blood flow changes were appreciably less in the rats injected through indwelling i.t. catheters. Somatostatin-induced reductions in spinal cord perfusion were accompanied by transient pressor responses, reduced cardiac output, 3-fold increases in spinal cord cerebrospinal fluid lactic acid concentrations and breakdown of the blood-spinal cord barrier, as reflected by significantly increased extravasation of [125I]bovine serum albumin. By 24 hr postinjection, a 12.5-nmol dose of somatostatin caused appreciable spinal cord cellular injury, as evidenced by significant elevations in cerebrospinal fluid concentrations of lactate dehydrogenase. After topical application to exposed pial vessels of the parietal cortex, comparable doses of somatostatin caused immediate intense dose-related arteriolar vasospasm and subsequent extravasation of the visible macromolecular tracer Evans blue dye. We conclude that somatostatin has significant vasoconstrictory effects on the blood vessels of the brain and spinal cord of the rat that must be recognized and appreciated when studying its neuropharmacological actions in vivo.     

Surgeon at work: hand-assisted laparoscopic construction of gastric conduit for thoracic esophageal cancer

A method for hand-assisted laparoscopic construction of gastric conduit for thoracic esophageal cancer was developed. Since this endoscopic surgical procedure is less invasive than open surgery, it contributes to improvement of post-operative respiratory functions and reduces respiratory complications. What distinguishes our surgical procedure is that unlike methods described in previous reports, it begins with treatment of the left gastroepiploic vessels at the height of the inferior edge of the spleen, followed by dissection from the esophageal hiatus to the lesser curvature and then dissection and excision of left gastric arteries and veins. Finally, the exposed esophagus and stomach are drawn outside the body and the right gastroepiploic blood vessels are preserved, followed by dissection of the greater omentum. This approach to gastric conduit construction was undertaken in 6 patients and the mean operating time was 123 minutes. Although in the first 3 of these patients the operating time was 150 minutes or more, the time required shortened to around 90 minutes for each of the last 3 cases, as the procedure was mastered. In each case, the volume of intraoperative hemorrhage did not exceed 50 ml.     

Blunt and penetrating chest trauma

Blunt or penetrating trauma to the chest can cause several life-threatening conditions: open or closed pneumothorax, each with or without hemothorax; flail chest; pericardial tamponade, and injury to other structures in the chest--the esophagus, trachea or great vessels. Any trauma sufficient to compromise function of thoracic organs must also be suspect for extrathoracic injuries, especially to the spleen and other abdominal viscera.     

高压氧对外源性人神经干细胞移植治疗损伤脑组织神经元病理状态的改善效应

目的：研究证明高压氧治疗能明显减轻缺氧后宿主脑区深部水肿，减少内源性神经元变性与坏死，并增加围产期鼠新生神经元数量与碱性成纤维生长因子活性。实验拟进一步观察缺氧缺血性脑损伤大鼠移植人神经干细胞后予高压氧治疗对脑组织神经元病理改变的影响。 方法：实验于2007-04在解放军海军总医院完成。①细胞来源及动物：经医院伦理委员会授权，孕妇知情同意下留取孕12周流产的人胎儿脑组织。新生7d龄SD大鼠20只，购自北京维通利华实验动物技术有限公司，实验过程中对动物的处置符合动物伦理学标准。②实验方法：取胎儿脑组织，在无菌条件下培养扩增并制备成人神经干细胞单细胞悬液。20只大鼠均建立缺氧缺血性脑损伤模型，3只死亡，剩余鼠随机数字表法分为2组，细胞移植＋高压氧组9只，单纯细胞移植组8只。造模后3d，两组大鼠均于左侧侧脑室进行细胞移植，注射位点以前囟为参照点，AP=-1mm，ML=-1．5mm，DV=-4．0mm，缓慢注入2×10^6L^-1细胞悬液5μL。移植后1h，将细胞移植＋高压氧组大鼠放入高压氧舱内，给予高压氧通气，升压及降压过程各15min，最终达压力为1．8个绝对大气压，稳压60min，1次/d，连续10d。两组大鼠麻醉后断头取脑，制备组织切片。③实验评估：免疫荧光法检测神经干细胞巢蛋白的表达、人神经干细胞植入后神经丝蛋白表达及其向神经元分化情况。尼氏染色检测移植后脑组织皮质、海马神经元形态、结构的变化。 结果：移植过程中细胞移植＋高压氧组有1只鼠因麻醉死亡，高压氧通气过程中没有动物死亡。①人神经干细胞的鉴定：人胎儿脑组织体外培养12d获取生长旺盛的人神经干细胞小集落，即神经球。倒置显微镜下观察可见每个小集落为数十个细胞构成，细胞折光性强，周边有清晰的光晕。免