Rodent stereotaxic surgery and animal welfare outcome improvements for behavioral neuroscience

Stereotaxic surgery for the implantation of cannulae into specific brain regions has for many decades been a very successful experimental technique to investigate the effects of locally manipulated neurotransmitter and signaling pathways in awake, behaving animals. Moreover, the stereotaxic implantation of electrodes for electrophysiological stimulation and recording studies has been instrumental to our current understanding of neuroplasticity and brain networks in behaving animals. Ever-increasing knowledge about optimizing surgical techniques in rodents(1-4), public awareness concerning animal welfare issues and stringent legislation (e.g., the 2010 European Union Directive on the use of laboratory animals(5)) prompted us to refine these surgical procedures, particularly with respect to implementing new procedures for oxygen supplementation and the continuous monitoring of blood oxygenation and heart rate levels during the surgery as well as introducing a standardized protocol for post-surgical care. Our observations indicate that these modifications resulted in an increased survival rate and an improvement in the general condition of the animals after surgery (e.g. less weight loss and a more active animal). This video presentation will show the general procedures involved in this type of stereotaxic surgery with special attention to our several modifications. We will illustrate these surgical procedures in rats, but it is also possible to perform this type of surgery in mice or other small laboratory animals by using special adaptors for the stereotaxic apparatus(6).     

康复运动对冠脉支架植入术后患者相关因素的影响

目的：研究心脏康复运动对冠脉支架植入术后患者血脂、血糖、体重指数及生活质量的影响。方法：对实施冠脉支架植入术的 146例患者进行比较分析，根据随机原则分为试验组76 例及对照组70 例。对照组患者给予常规的健康教育及冠心病二级预防指 导，给予定期随访。试验组患者在此基础上给予规律的康复运动指导。经过6 个月随访，比较两组患者血脂、HbA1C、体重指数及 生活质量情况。结果：试验组患者通过为期6 个月的规律的心脏康复运动指导，其血脂、HbA1C等冠心病危险因素控制情况优于 对照组，差异有统计学意义(P<0.05)。同时，6个月后，试验组康复运动六月后sF 量表各项评分与对照组同期比较，差异均有统计 学意义(P<0.05)。结论：规范的心脏康复运动指导能够有效改善冠脉支架植入术后患者血脂、血糖情况，提高患者生活质量。     

Perception of Animals and Cardiovascular Responses During Verbalization With an Animal Present

Abstract

The presence of animals has been associated with decreased physiological responses to stressors. Not all individuals respond equally to the presence of friendly animals. The current study was designed to examine whether attitudes toward animals are related to individuals' physiological responses when an animal is present. The relationship of individuals' perceptions of animals to their blood pressure and heart rate responses during verbalization in the presence of a dog were examined among urban college students (n=218). Lockwood's projective Animal Thematic Apperception Test (ATAT) was used to assess subjects' attitudes toward animals and people in scenes containing animals and identical scenes without animals. The significant period by perception interactions in analyses of variance with repeated measures revealed that cardiovascular responses to verbalization with an animal present were significantly lower for individuals who perceived scenes with animals more positively than for individuals who perceived scenes with animals present less positively. Cardiovascular responses when the dog was present were not related to perceptions of scenes without animals present. The differences in cardiovascular responses depended upon the scenes used. This study supports the view that how people perceive animals moderates their physiological responses to stressors when an animal is present.     

Evidence of Yersinia pestis DNA from fleas in an endemic plague area of Zambia

Background

Study protocols involving experimental animals often require the monitoring of different parameters not only in anesthetized, but also in free moving animals. Most animal research involves small rodents, in which continuously monitoring parameters such as temperature and heart rate is very stressful for the awake animals or simply not possible. Aim of the underlying study was to monitor heart rate, temperature and activity and to assess inflammation in the heart, lungs, liver and kidney in the early postoperative phase after experimental cardiopulmonary bypass involving 45 min of deep hypothermic circulatory arrest in rats. Besides continuous monitoring of heart rate, temperature and behavioural activity, the main focus was on avoiding uncontrolled death of an animal in the early postoperative phase in order to harvest relevant organs before autolysis would render them unsuitable for the assessment of inflammation.

Findings

We therefore set up a telemetry-based system (Data Science International, DSI?) that continuously monitored the rat's temperature, heart rate and activity in their cages. The data collection using telemetry was combined with an analysis software (Microsoft excel?), a webmail application (GMX) and a text message-service. Whenever an animal's heart rate dropped below the pre-defined threshold of 150 beats per minute (bpm), a notification in the form of a text message was automatically sent to the experimenter's mobile phone. With a positive predictive value of 93.1% and a negative predictive value of 90.5%, the designed surveillance and alarm system proved a reliable and inexpensive tool to avoid uncontrolled death in order to minimize suffering and harvest relevant organs before autolysis would set in.

Conclusions

This combination of a telemetry-based system and software tools provided us with a reliable notification system of imminent death. The system's high positive predictive value helped to avoid uncontrolled death and facilitated timely organ harvesting. Additionally we were able to markedly reduce the drop out rate of experimental animals, and therefore the total number of animals used in our study. This system can be easily adapted to different study designs and prove a helpful tool to relieve stress and more importantly help to reduce animal numbers.     

Norepinephrine turnover in the heart and blood vessels of rats subjected to bilateral renal infarction

The total norepinephrine (NE) content, the uptake of [3H]NE, the turnover rate and the synthesis rate of the neurotransmitter at the heart and blood vessels have been studied during the development of hypertension in rats subjected to bilateral renal infarction. Normal and sham-operated rats were used as controls. Fifty percent of the rats with renal infarction became hypertensive. The weight of the hearts and blood vessels of the experimental animals was significantly increased 15 days after renal infarction. Changes were greater in hypertensive animals. NE concentration in the heart was slightly decreased without achieving statistical significance, while total NE content was unchanged. In the artery wall NE concentration was significantly decreased in normotensive and hypertensive operated rats. [3H]NE uptake in the heart and blood vessels was similar in experimental and control animals. In relation to NE turnover, in both the heart and blood vessels, normal and sham-operated animals behaved as one population while normotensive and hypertensive rats behaved as another population. The rate constant of NE turnover was increased in both tissues of operated experimental animals without achieving statistical significance in the case of the heart. NE synthesis rate was unchanged in the cardiac muscle but was significantly increased in the blood vessels of operated animals. Present data indicate that results describing NE dynamics in the heart cannot be extrapolated for the blood vessels level; on the other hand changes in the neurotransmitter do not seem to be related to the development of high blood pressure after renal infarction in the rat.     

Studies on a peptidase acting on a synthetic collagenase substrate Developmental pattern in rat granuloma tissue and distribution of enzyme in the tissues of various animals

The developmental pattern in experimental rat granuloma tissue and the distribution in the tissues of a few animals (monkey, rabbit, guinea pig anrat) of a peptidase acting on a synthetic collagenase substrate, 4-phenylazobenzyloxycarbonyl-L-Pro-L-Leu-Gly-L-Pro-D-Arg (Pz-peptide) has been studied. Maximum enzyme activity was found in 4-month-old rats and on the fourth day of implantation of the cotton wick. Pz-peptidase appears to have a ubiquitous distribution in animal tissues; the highest enzyme activity was generally found in liver, intestine and kidney of the animals. The total activity in other organs (spleen, heart, lungs and brain) was much less compared to that of liver, intestine or kidney.     

Imaging of inflammatory foci with 99m technecium labeled antimicrobial oligopeptide

A direct method using 99mtechnecium to label the synthetic antimicrobial oligopeptide ubiquicidine with a molecular mass of 1.7 kDa has been developed. The radiochemical purity of the resultant labeled compound was more than 98%. A combination of ubiquicidine and 99mtechnecium showed stability in blood plasma. The biological study indicated that the complex was renally excreted from blood flow rapidly (the half-life is 30 min). It accumulated in the experimental inflammatory focus of laboratory animals, peaking 1-2 hours after intravenous injection when the inflammatory focus/intact tissue ratio amounted up to 2.6-3.4, which could yield satisfactory abscess scintigrams. The findings make it possible to consider ubiquicidine as a promising compound for the design of radiopharmaceutical to image inflammatory foci.     

Trial Implantation of Heart Rate Data Loggers in Pinnipeds

ABSTRACT Pinnipeds are major consumers in marine ecosystems, and understanding their energy budgets is essential to determining their role in food webs, particularly where there is competition with fisheries. Food consumption and energy expenditure have been evaluated in pinnipeds using different methods, but the use of heart rate to estimate energy expenditure is potentially a very powerful tool suited to the life history of these animals. We tested a procedure for the subcutaneous implantation of heart rate data loggers to determine whether heart rate could be recorded for ≥1 year in free-ranging pinnipeds, as it has been in birds. We implanted 3 captive California sea lions (Zalophus californianus) and 3 captive northern elephant seals (Mirounga angustirostris) with heart rate data loggers and monitored their recovery and behavior in a controlled environment. In both species, the implantation site allowed for excellent detection of the electrocardiogram, and we observed heart rate signatures characteristic of behaviors such as resting and diving. Although all 3 sea lions recovered well from the implantation surgery, all 3 elephant seals showed a substantial inflammatory response for unknown reasons, and we removed the implanted data loggers. Subcutaneous implantation of data loggers is a powerful technique to study physiology, energetics, and behavior in California sea lions, but more work is required to realize the potential of this technique in northern elephant seals.     

Different Dosage Effects of Ozone on Oxygen Transport in Blood during in vitro Experiments

Biophysics - The effects of ozone on oxygen transport in the blood of experimental animals have been studied in vitro. Incubation of the blood with ozonated water at О3 concentrations of 2,...     

Controlling gene loss of function in newts with emphasis on lens regeneration

Here we describe a protocol for gene loss of function during regeneration in newts, specifically applied to lens regeneration. Knockdown with the use of morpholinos can be achieved both in vitro and in vivo, depending on the experimental design. These methods achieve desirable levels of gene knockdown, and thus can be compared with methods developed for use in other animals, such as zebrafish. The technology has been applied to study molecular mechanisms during the process of lens regeneration by knocking down genes at specific stages and examining their effects on other genes and lens differentiation. The protocol can take a few days or up to 20 d to complete, depending on the duration of the experiment.     

Role of the hypothalamus in the reflex regulation of cardiac activity

A review of literary and own experimental data is given. More adequate method in the study of the hypothalamic heart regulation is the investigation of the hypothalamic influences on the cardiac reflexes. The necessity of the experiments on the unanesthetized unrestrained animals is underlined. The dual modulating hypothalamic influences existing already in fishes having been revealed on the unanesthetized animals. The character of the modulating influence is determinated to a certain extent by the cardiac reflex intensity: small responses are increased, larger ones are reduced. The extension of system regions involved into the reflex response pattern at the increase of the degree of temperature effect of venous blood on heart has been discovered in unanesthetized unrestrained cats.     

Techniques for long-term multisite neuronal ensemble recordings in behaving animals.

Advances in our understanding of neural systems will go hand in hand with improvements in the experimental techniques used to study these systems. This article describes a series of methodological developments aimed at enhancing the power of the methods needed to record simultaneously from populations of neurons over broad regions of the brain in awake, behaving animals. First, our laboratory has made many advances in electrode design, including movable bundle and array electrodes and smaller electrode assemblies. Second, to perform longer and more complex multielectrode implantation surgeries in primates, we have modified our surgical procedures by employing comprehensive physiological monitoring akin to human neuroanesthesia. We have also developed surgical implantation techniques aimed at minimizing brain tissue damage and facilitating penetration of the cortical surface. Third, we have integrated new technologies into our neural ensemble, stimulus and behavioral recording experiments to provide more detailed measurements of experimental variables. Finally, new data analytical techniques are being used in the laboratory to analyze increasingly large quantities of data.     

Drawing blood from rats through the saphenous vein and by cardiac puncture

Drawing blood from rodents is necessary for a large number of both in vitro and in vivo studies. Sites of blood draws are numerous in rodents: retro-orbital sinus, jugular vein, maxillary vein, saphenous vein, heart. Each technique has its advantages and disadvantages, and some are not approved any more in some countries (e.g., retro-orbital draws in Holland). A discussion of different techniques for drawing blood are available (1-3). Here, we present two techniques for drawing blood from rats, each with its specific applications. Blood draw from the saphenous vein, provided it is done properly, induces minimal distress in animals and does not require anesthesia. This technique allows repeated draws of small amounts of blood, such as needed for pharmacokinetic studies (4,5), determining plasma chemistry, or blood counts (6). Cardiac puncture allows the collection of large amounts of blood from a single animal (up to 10 ml of blood can be drawn from a 150 g rat). This technique is therefore very useful as a terminal procedure when drawing blood from the saphenous would not provide a large enough sample. We use cardiac puncture when we need sufficient amounts of serum from a specific strain of rats to grow T lymphocyte lines in vitro (4-9).     

Reduction in animal numbers by long-term implantation of intravenous and intra-arterial catheters in thyroparathyroidectomized rats

The thyroparathyroidectomized (TPTx) rat has been extensively used to study parathyroid hormone (PTH)-mediated bone resorption by measuring systemic Ca2+ concentrations. Animals have been traditionally used acutely; that is, they are often infused immediately after surgery and are sacrificed after a single use. To perform multiple experiments using a single group of animals we developed a system of long-term implanted intravenous/arterial catheters. Using calcitonin (CT) as a positive control, we successfully completed 12 separate controlled subexperiments documenting significant reductions in PTH-induced hypercalcemia in rats of the CT group. We then successfully completed two separate TPTx subexperiments, using a 3 x 3 Latin square experimental design. In both subexperiments, CT significantly inhibited the increase of blood Ca2+ concentration resulting from continuous PTH infusion. Our results indicate that, by combining the long-term use of catheters with the Latin square design, we can successfully reduce the number of animals used, increase the number of compounds screened, and improve the quality of the data. Although results of this study confirmed the acceptability of multiple infusions in anti-resorptive studies, investigations into the applicability of this set up to other areas of study requiring infusions and frequent blood sample collections seem appropriate.     

Revealing the hidden mechanisms of smoke-induced fallopian tubal implantation

Ectopic pregnancy (EP) is an enigmatic reproductive disorder. Although tubal EP is difficult to predict, several hypotheses about its etiology have been proposed. In retrospective case-control studies, smoking is associated with an increased rate of EPs in the fallopian tube. Studies of experimental animals in vivo and human fallopian tubal tissues in vitro have suggested mechanisms of fallopian tubal damage and dysfunction induced by nicotine and other smoking-related chemicals that may explain this association. However, the pathogenesis of smoking-induced modulation of implantation leading to tubal EP is largely unknown. Because cigarette/tobacco smoke adversely affects the success of intrauterine implantation, there is a great need to determine how embryo implantation occurs in the fallopian tube in female smokers of reproductive age.     

Trypanosoma cruzi: A Specific Surface Marker for the Amastigote Form1

ABSTRACT. Among the known life cycle stages of Trypanosoma cruzi only the amastigote form bound lactoferrin (LF), a glycoprotein produced by neutrophils. This capacity was readily demonstrable by indirect immunofluorescence in amastigotes derived from mice, a mammalian cell culture, or grown in an axenic medium. No LF binding was detectable on trypomastigotes from blood or mammalian cells, insect-derived metacyclics or epimastigotes, or on epimastigotes grown in Warren's medium. Serum levels of LF were increased in mice acutely infected with T. cruzi, and amastigotes from the spleens of these animals were found to have the glycoprotein on their surface. The amastigote LF receptor may have biological significance in parasite-host interaction since mononuclear phagocytes also express a LF receptor, and treatment of these cells with LF has been shown to increase their capacities to take up and kill T. cruzi amastigotes in vitro. The LF receptor is the first marker for T. cruzi amastigotes for which a naturally occurring ligand has been described.     

Implantation of embryonic pancreas into the anterior chamber of healthy and diabetic rats

The possibility of pancreatic-islet embryonic implantation into the frontal chamber of the eye in Wistar rats has been demonstrated. The implantation was followed by the induction of alloxan diabetes. Clinical diabetic compensation has been shown in a number of experimental animals (maximum time of experiments--45 days). The formation of islet-like structures in the frontal chamber of the eye was confirmed microscopically. The structures contained B cells in their cytoplasm. Cellular differentiation leading to the appearance of functionally active endocrine cells has been noted in the implants.     

Hemocompatibility and hemodynamic comparison of two centrifugal LVADs: HVAD and HeartMate3

Mechanical circulatory support using ventricular assist devices is a common technique for treating patients suffering from advanced heart failure. The latest generation of devices is characterized by centrifugal turbopumps which employ magnetic levitation bearings to ensure a gap clearance between moving and static parts. Despite the increasing use of these devices as a destination therapy, several long-term complications still exist regarding their hemocompatibility. The blood damage associated with different pump designs has been investigated profoundly in the literature, while the hemodynamic performance has been hardly considered. This work presents a novel comparison between the two main devices of the latest generation–HVAD and HM3–from both perspectives, hemodynamic performance and blood damage. Computational fluid dynamics simulations are performed to model the considered LVADs, and computational results are compared to experimental measurements of pressure head to validate the model. Enhanced performance and hemocompatibility are detected for HM3 owing to its design incorporating more conventional blades and larger gap clearances.

     

Task Design Influences Prosociality in Captive Chimpanzees (Pan troglodytes)

Chimpanzees confer benefits on group members, both in the wild and in captive populations. Experimental studies of how animals allocate resources can provide useful insights about the motivations underlying prosocial behavior, and understanding the relationship between task design and prosocial behavior provides an important foundation for future research exploring these animals'' social preferences. A number of studies have been designed to assess chimpanzees'' preferences for outcomes that benefit others (prosocial preferences), but these studies vary greatly in both the results obtained and the methods used, and in most cases employ procedures that reduce critical features of naturalistic social interactions, such as partner choice. The focus of the current study is on understanding the link between experimental methodology and prosocial behavior in captive chimpanzees, rather than on describing these animals'' social motivations themselves. We introduce a task design that avoids isolating subjects and allows them to freely decide whether to participate in the experiment. We explore key elements of the methods utilized in previous experiments in an effort to evaluate two possibilities that have been offered to explain why different experimental designs produce different results: (a) chimpanzees are less likely to deliver food to others when they obtain food for themselves, and (b) evidence of prosociality may be obscured by more “complex” experimental apparatuses (e.g., those including more components or alternative choices). Our results suggest that the complexity of laboratory tasks may generate observed variation in prosocial behavior in laboratory experiments, and highlights the need for more naturalistic research designs while also providing one example of such a paradigm.     

Combined Internal Mammary Artery Implantation and Free Omental Graft Operation: A Highly Effective Revascularization Procedure: (A Study of 17 Cases)

One hundred and forty patients with coronary artery disease treated by internal mammary artery implantation were followed up from six months to 13 years. The mortality rate was 3%; improvement was obtained in 70%. Occlusive disease of all three major coronary arteries, however, requires more blood than can be supplied by the implant procedure, and the free omental graft operation was developed for the management of selected patients with this condition. Normally in animals triple coronary artery ameroid constriction results in 100% mortality. When the free omental graft operation, with or without internal mammary artery implantation, was performed, 80% of such animals survived. The free omental graft forms capillary anastomoses in three days and arteriolar vessels in eight days, which leave the base of the aorta, enter the omentum, thence to the myocardium. A combined internal mammary artery implantation and free omental graft operation was performed in 17 patients with triple coronary artery disease as shown by cine coronary arteriography. There was no operative mortality, and 12 of the patients are free of pain and have returned to work.