Improved method for the catheterization of the right ventricle in a rat model of pulmonary artery hypertension.

Ventricle catheterization in the rat is widely practiced in cardiopulmonary research. The catheters deployed are either fluid filled or solid tip pressure or pressure-volume catheters. The access to the right ventricle is through the right jugular vein, most commonly without direct visualization such as fluoroscopy. Advancement of the catheter tip is aided by visualizing the pressure signals of the monitoring/recording systems used. This approach may present challenges due to various reasons, including the stiffness of new catheters, their dimensions or anatomical changes associated with the animal disease model. In this article, we present a novel approach, which has been optimized, successfully validated surgically and adopted in current projects. It has been shown to improve both the overall quality of the signals recorded and the time to access the right ventricle, thus reducing the overall time of surgery. The method presented in this article is safe, easy to reproduce and does not require additional skills compared to a more 'standard' approach.

Evaluation of temperature rise and bonding strength in cements used for permanent head attachments in rats and mice.

In animal models, devices such as indwelling catheters and intracranial cannulae are often fixed on the skull to allow sampling or injection in the freely moving animal. The most commonly used method to fixate these devices is by embedding them in a 'helmet' of cement which is fixed to the skull with screws. Methylmethacrylate cement is commonly used for this purpose. The disadvantages of this cement are the high polymerization temperature, poor bonding to the bone and long hardening time. We have evaluated the use of glass ionomer cement, carboxylat cement and cyanoacrylic glue as alternative for methylmethacrylate cement. Temperature increase during polymerization of methylmethacrylate cement and glass ionomer cement was measured in the cement on the skull and in the brain of 14 rats in an acute model. In a chronic model, 52 rats and 91 mice were equipped with a 'helmet' of one of the cements. The glass ionomer 'helmets' were applied without or with pretreatment of the skull. The attachment of the cement to the skull was checked every day. After four weeks the bonding strengths of the cements were measured. The glass ionomer cement had less temperature increase during polymerization and good bonding capabilities when compared with methylmethacrylate cement. Mechanical pretreatment of the skull resulted in a significant increase in bonding strength of glass ionomer cement in mice and rats as compared with chemical pretreatment. Furthermore, glass ionomer cement had a shorter hardening time than methylmethacrylate cement, and when the glass ionomer cement was used in prepacked capsules, it was possible to apply the cement sterilely and easily. Cyanoacrylic glue had good bonding capabilities to the skull of mice and is also a good substitute for methylmethacrylate cement.

Intraperitoneal versus subcutaneous telemetry devices in young Mongolian gerbils (Meriones unguiculatus).

Radiotelemetry has become a very popular biotelemetric tool for measuring physiological parameters such as heart rate, blood pressure, body temperature and muscle activity, as well as general behavioural activity in undisturbed, freely moving animals. In most studies using this technique, adult subjects are used. However, sometimes an ontogenetic approach is required to clarify whether changes in one parameter are preceeded or followed by changes in another parameter. Tracking physiological changes in young, developing individuals could explain given states of these animals as adults. Implanting telemetry devices can be done subcutaneously and intraperitoneally, the former method posing less of a challenge on the animal and its recovery from surgery. Because telemetry will be used in weanling gerbils during subsequent studies, we needed to investigate whether subcutaneous implantation of telemetric devices is preferable to intraperitoneal surgery with respect to animal welfare. This is a technical paper describing anaesthetic and surgical techniques in detail during a pre-trial involving subcutaneous (n=10, aged 21-29 days) and intraperitoneal (n=10, aged 19-34 days) implantation of dummy telemetry transmitters (1.9 cm3, 3.6 g after shortening of leads) in weanling gerbils, Meriones unguiculatus. Body weight was measured and analysed over four-day intervals. Optimizing anaesthetic dosages was a first step in this pilot trial. This occurred during the first few subcutaneous implantations. Three animals died while anaesthetized during the subcutaneous procedure but none post-surgery. All animals survived anaesthesia during the intraperitoneal implantation, but two died in the first three days post-surgery. In the former method, the tension on the dermal sutures caused by the presence of the transmitters was too great, resulting in the animals opening the sutures by chewing them. The animals died during the latter procedure probably due to strangulation of the intestine by the excess lead that was coiled in the abdomen. Furthermore, placement of the exposed negative lead of the transmitter on the underlying muscle had to be done on the m. pectoralis transversus in order for it to stay in place as the animal developed. This paper showed that the implantation of a telemetric device in weanling gerbils is feasible and is best executed through the intraperitoneal technique.

Factors stimulating or obstructing the implementation of the 3Rs in the regulatory process.

Approximately 30% of animal use within the European Union (EU) is done to meet regulatory requirements. The tests are often repetitive in nature and may cause severe suffering, due to the procedures used and to rigidly predefined end points. In addition, product evaluation procedures often take long and are very expensive. Over the last decades the heavy reliance on animal experimentation in this area has met serious objections, both ethical and economical in nature. This study describes obstacles and opportunities to implement the 3Rs in regulatory animal testing. The findings are based primarily on interviews with legislators, regulators, industry, science and animal welfare organisations and reflect shared perceptions of these respondents. In order to increase the application of the 3Rs in regulatory testing a number of technical, political and social obstacles must be overcome. This study offers insight into the persistent character of regulatory animal testing and can function as a starting point for further discussion on how to tackle these problems. To this end, several recommendations are made ranging from strategic test approaches and data sharing to strengthening the policy network and improving communication between 3Rs experts and regulators. The study is an initiative of the national project group "Regulatory Animal Testing", which consists of a group of Dutch experts on animal testing working for a variety of organisations in the field.1 They felt the need for cooperation to initiate a discussion at relevant levels and to identify possible solutions in order to implement the objectives of the three R's in testing for regulatory purposes without loss of scrutiny in safety and/or efficacy evaluation needed for product release.

A guinea pig model of acute and chronic asthma using permanently instrumented and unrestrained animals.

To investigate mechanisms underlying allergen-induced asthmatic reactions, airway hyperresponsiveness and remodeling, we have developed a guinea pig model of acute and chronic asthma using unanesthetized, unrestrained animals. To measure airway function, ovalbumin (IgE)-sensitized animals are permanently instrumented with a balloon-catheter, which is implanted inside the pleural cavity and exposed at the neck of the animal. Via an external cannula, the balloon-catheter is connected to a pressure transducer, an amplifier, an A/D converter and a computer system, enabling on-line measurement of pleural pressure (P(pl))-closely correlating with airway resistance-for prolonged periods of time. Using aerosol inhalations, the method has been successfully applied to measure ovalbumin-induced early and late asthmatic reactions and airway hyperresponsiveness. Because airway function can be monitored repeatedly, intra-individual comparisons of airway responses (e.g., to study drug effects) are feasible. Moreover, this model is suitable to investigate chronic asthma and airway remodeling, which occurs after repeated allergen challenges. The protocol for establishing this model takes about 4 weeks.

Measuring blood pressure in small laboratory animals.

The most common techniques currently employed for monitoring blood pressure (BP) in conscious rats and mice are the tail cuff and the exteriorized catheter that feeds a pressure transducer located outside the cage. There are, however, considerable drawbacks associated with these methods, which in many respects make each of these techniques undesirable as an accurate means of obtaining pressure measurements. Recent studies have shown that measurements of physiological variables, such as electrocardiogram (ECG), heart rate (HR), and body temperature (BT), from freely moving rats and mice by using implantable radiotelemetry were more efficient, reliable and less labour intensive when compared to measurement techniques described in the literature so far. Nowadays, measurement of BP by radiotelemetry has been described and validated for many laboratory animal species, including rats and mice. The implantable radiotelemetry technique can circumvent many of the problems associated with conventional methods (tail cuff; exteriorized catheters) of BP monitoring in mice and rats. This chapter describes the surgical aspects of the radiotelemetry techniques currently used to monitor and measure blood pressure in awake animals.

Effect of conditioning on the increase of heart rate and body temperature provoked by handling in the mouse.

To assess the effect of procedures on animal welfare, various physiological parameters, such as body weight, hormone levels in plasma and/or urine, heart rate (HR), blood pressure and body temperature (BT), can be used. When measuring physiological parameters with techniques involving restraint of the animals, the results must be interpreted with caution, since restraint itself may have an effect on those parameters. Radio-telemetry, using an implantable transmitter, provides a way to obtain more accurate and reliable physiological measurements from freely moving animals in their own environment. In this study, we have used radio-telemetry to investigate the influence of conditioning on the increase of HR and BT as provoked by handling of mice. It was found that, after a conditioning period of 12 days, the increase of HR due to handling was significantly reduced.