Laboratory Animals recent issues

The impact of light, noise, cage cleaning and in-house transport on welfare and stress of laboratory rats

Castelhano-Carlos, M J, Baumans, V — 2009-09-18

Human interaction and physical environmental factors are part of the stimuli presented to laboratory animals everyday, influencing their behaviour and physiology and contributing to their welfare. Certain environmental conditions and routine procedures in the animal facility might induce stress responses and when the animal is unable to maintain its homeostasis in the presence of a particular stressor, the animal's wellbeing is threatened. This review article summarizes several published studies on the impact of environmental factors such as light, noise, cage cleaning and in-house transport on welfare and stress of laboratory rats. The behaviour and physiological responses of laboratory rats to different environmental housing conditions and routine procedures are reviewed. Recommendations on the welfare of laboratory rats and refinements in experimental design are discussed and how these can influence and improve the quality of scientific data.

Preclinical testing on insects predicts human haematotoxic potentials

Berger, J. — 2009-09-18

The substitution of insects for laboratory animals in toxicity testing is likely to become a reality in the framework of prescreening. Haematotoxicological studies of newly developed chemicals, such as food components, drugs, etc. performed on insects can offer advantages in, for example, environmental toxicology. Reliable routine predictions should produce an increase in our knowledge of haemocyte physiology. Although the differences between human physiology and morphology and those of insects are great, the basic functions of insect haemocytes and mammalian leukocytes appear not to have changed during evolution. The use of insects in haematotoxicity assays represents a preclinical testing strategy which will lower costs, accelerate screening and offer ethical benefits.

Vascular ultrasound studies for the non-invasive assessment of vascular flow and patency in experimental surgery in the pig

2009-09-18

Vascular ultrasound is a reliable non-invasive tool used for the routine assessment of vascular flow and patency in human recipients. We describe the use at three different time points (immediately, 1 week and 4 weeks postsurgery) of ultrasound studies and its validation by angiographic studies in 37 swine undergoing carotid graft replacement. We calculated predictive values (>92%), sensitivity (>85%) and specificity (>92%) with high results at all time points. Ultrasound appeared as an accessible non-invasive technique, providing rapid, safe, repeatable and reliable results. It is an excellent alternative to angiography, avoiding risks inherent to invasive methods and therefore contributing to animal welfare.

Development of perioperative care for pigs undergoing laryngeal transplantation: a case series

Murison, P J, Jones, A, Mitchard, L, Burt, R, Birchall, M A — 2009-09-18

Pigs are ideal animal models for airway surgical research, facilitating the successful translation of science into clinical practice. Despite their ubiquitous use, there is a paucity of information on the perioperative care of pigs, especially for major procedures. In a series of experiments to investigate laryngeal transplantation, we combined veterinary and medical experience to develop protocols for perioperative management of pigs, including high dependency care. Novel airway management methods were developed. A pain scoring system was used to direct analgesia use. Fluid balance and electrolytes were monitored closely. Recent animals received a central venous line via the femoral vein two days prior to transplantation to facilitate blood sampling and drug delivery. Intensive monitoring and airway management were required to ensure a successful outcome. Methods for optimal perioperative care are proposed. These results will help future groups wishing to use pigs in airway research, will reduce numbers of animals used and improve animal welfare.

Percutaneous external jugular vein catheterization in piglets using a triangulation technique

Flournoy, W S, Mani, S — 2009-09-18

Chronic jugular vein or central venous cannulation is routinely performed in human and animal patients for access to blood circulation. In mature swine, chronic catheter placement techniques have typically involved venous isolation via extensive cut-down, blunt dissection and manipulation of ventral neck tissues prior to catheter placement. More recently, guide-wire-assisted percutaneous techniques have become standard practice in human and veterinary medicine due to the minimization of soft tissue and vessel damages. Laboratory animal piglets are becoming more popular research models because of their immature immunological system, ease of handling and costs. However, external jugular veins are very difficult to catheterize in paediatric animals including freshly weaned piglets. The objective of this study was to develop a simple, safe and efficient method for external jugular vein cannulation in young piglets. In total, 20 piglets were anaesthetized and percutaneously catheterized with a guide-wire technique using palpable anatomical landmarks and triangulation. With this minimally invasive catheterization, it has allowed our veterinarians and veterinary technicians to quickly and easily obtain central venous access in piglets undergoing operative procedures.

Characterization of a rodent model for the study of arterial microanastomoses with size discrepancy (small-to-large)

Rickard, R F, Wilson, J, Hudson, D A — 2009-09-18

Microsurgical autotransplantation of tissues is employed clinically to reconstruct defects following burns, trauma and surgical cancer ablation, and to correct congenital abnormalities. Transplant vessels of <3 mm are anastomosed by hand under the microscope. Experimentally, anastomotic patency rates decrease with increasing vessel diameter mismatch, and clinically, ratios of 3:1 or greater lead to unacceptably low arterial patency rates. A number of surgical techniques for dealing with size mismatch are described, but no one method has found favour, and few controlled studies of technique are reported. In this report, a rodent superficial caudal epigastric artery (SCEA)/femoral artery (FA) model for the study of these techniques is described in detail. The diameter ratio between these vessels lies in the clinically relevant range of 1:1.5–1:2.5. In the male Wistar rat, external vessel diameters were not found to increase markedly in size between animal weights of 300 and 500 g. The length of FA distal to the origin of the SCEA, which is important in allowing undisturbed distal run-off, was found to be negatively associated with animal body weight, implying that a smaller animal would be better for this model. Mean femoral arterial flow rate, measured by transit-time ultrasound, was noted to be statistically and physiologically significantly higher in the right artery when compared with the left. This model has advantages over interposition vein graft models in that it minimizes vessel compliance mismatch, and avoids the need for a second anastomosis.

The analgesic effects of oral paracetamol in two strains of mice undergoing vasectomy

Dickinson, A. L, Leach, M. C, Flecknell, P. A — 2009-09-18

As the production of transgenic mice increases, the need for vasectomized mice also increases. Currently, there is no accurate method of identifying pain in these mice which can be used routinely and therefore no acceptable analgesic regimens can be established. Sixteen male CBA/CaCrl and 16 male DBA/2JCrl mice were randomly allocated to one of four treatment groups (saline, low, medium and high dose oral paracetamol) and then underwent abdominal vasectomy. Their behaviour was videotaped (filmed) preoperatively and at one hour postoperatively and the data were analysed using an automated system – HomeCageScan. HomeCageScan detected significant changes in 16 behaviours following vasectomy. Such behaviours included twitching, rearing and grooming with varying levels of significance between the strains. No significant effects of drug treatment in any of the behaviours analysed by HomeCageScan were detected in the postoperative observation period. Strain-specific changes do occur in behaviour following abdominal vasectomy and HomeCageScan is capable of detecting these changes. The use of orally administered paracetamol was deemed to be an ineffective analgesic for CBA/CaCrl and DBA/2JCrl mice undergoing vasectomy.

Charles River altered Schaedler flora (CRASF(R)) remained stable for four years in a mouse colony housed in individually ventilated cages

2009-09-18

As recommendations for specific pathogen-free housing change, mouse facilities need to re-derive their colonies repeatedly in order to eliminate specified bacteria or viruses. This paper describes the establishment of a new mouse facility using as starting point a small colony of CD-1 mice colonized with the Charles River altered Schaedler flora (CRASF^®) housed in individually ventilated cages (IVCs). The import of new strains was performed exclusively via embryo transfer using CD-1 mice as recipients. The integrity of the CRASF^® in caecum samples of the original CD-1 colony and of three inbred mouse lines imported into the colony was proven by a quantitative realtime polymerase chain reaction approach. Furthermore, we searched for bacterial contaminants in the gut flora using non-specific 16S rRNA primers. The bacterial sequences found were closely related to but not exclusively sequences of altered Schaedler flora (ASF) members, suggesting that the ASF is heterogeneous rather than restricted to the eight defined bacteria. Moreover, no pathogens were found, neither using the non-specific 16S rRNA primers nor in routine quarterly health monitoring. As one effect of this defined gut flora, interleukin-10 knockout mice are devoid of colitis in our facility. In conclusion, our approach building up a mouse facility using foster mothers and embryo transfer as well as a strict barrier system and IVCs is suitable to maintain a colony free from contaminating bacteria over the long term. CRASF^® remained stable for seven mouse generations and was efficiently transferred to the imported mouse strains.

Comparison of polymerase chain reaction primer sets for amplification of rodent Pasteurellaceae

Boot, R, Vlemminx, M J, Reubsaet, F A G — 2009-09-18

Monitoring of rodents for Pasteurellaceae infection may be carried out by the polymerase chain reaction (PCR). We tested which of 17 rodent Pasteurellaceae strains were detected by three PCR primer sets. By phylogenetic analysis, 12 strains were assigned to the Rodent cluster and five strains to other clusters, namely the Somnus cluster, Pasteurella sensu stricto, Actinobacillus sensu stricto, the Mannheimia and Rossii cluster. A primer set developed to detect biotype Heyl [Pasteurella] pneumotropica produced amplicons from three strains and appeared specific for this taxon. A primer set developed to detect biotype Jawetz [P.] pneumotropica produced amplicons from the [P.] pneumotropica type strain and two other strains within the Rodent cluster. A primer set as described by Bootz and his co-workers (Bootz F, Kirschnek S, Nicklas W, Wyss SK, Homberger FR. Detection of Pasteurellaceae in rodents by polymerase chain reaction analysis. Lab Anim Sci 1998;48:542–6) for the detection of all Pasteurellaceae indeed detected all bacterial strains examined. Bootz's primer set should be used to monitor rodents for Pasteurellaceae infection by PCR as FELASA recommends the monitoring of rodents for all Pasteurellaceae taxa. Health monitoring reports should specify the primer set(s) used for PCR testing rodents for Pasteurellaceae infection.

Morphological study on dental caries induced in WBN/KobSlc rats (Rattus norvegicus) fed a standard laboratory diet

2009-09-18

In our previous studies, WBN/KobSlc was characterized as a rat strain in which only males began to develop pancreatitis, and then presented with diabetic symptoms. In the course of studying their pancreatic inflammation, we detected molar caries in prediabetic males feeding on a standard diet (CRF-1) widely used for experimental animals. The purpose of this study is to confirm whether the WBN/KobSlc strain is caries-susceptible to the diet reported to be non-cariogenic, and to examine the effect of a prediabetic condition on their dental caries. For a morphological study, 25 male WBN/KobSlc rats aged 3.2–7.8 months and 24 females of the same strain aged 3.3–6.6 months were used, along with 10 males and 10 females of 8.2-month-old F344 rats. Marked dental caries were detected in the mandibular molars of male and female WBN/KobSlc rats regardless of pancreatitis, although no similar changes were observed in any teeth of the F344 strain fed the same diet. Soft X-ray examination revealed that the caries began in the crown and progressed horizontally and vertically, and that a severe radiolucent lesion extensively expanded to the entire crown, corresponding to a macroscopically deleted molar. The caries had gradually developed mainly in the second mandibular molar from more than 3.5 months of age, while none were seen in any rats before that time. The WBN/KobSlc rats were caries-susceptible even to the standard laboratory diet, and pancreatitis was not directly associated with the onset of dental caries in this strain.

Design and evaluation of a low-cost respiratory monitoring device for use with anaesthetized animals

Devonshire, I M, Preston, M J, Dommett, E J, Murphy, K L, Greenfield, S A — 2009-09-18

This report describes a simple, non-invasive electronic device that employs a compact accelerometer integrated circuit to transduce movements in the chest wall of an anaesthetized animal into an analogue signal that can be used to calculate the rate and relative depth of respiration. The device requires amplification by signal processing hardware/software which are common to most experimental laboratories. We assessed the sensitivity of the device by its ability to detect changes in respiratory patterns produced by modulating the depth of anaesthesia in isoflurane-anaesthetized Wistar rats. It is widely accepted that many anaesthetic agents affect respiratory patterns, especially respiratory rate (RR), which is often used as an important index of anaesthetic depth. Respiratory parameters obtained with the device were compared with concurrently recorded electroencephalographic and cardiac measures. Different concentrations of anaesthetic agent produced four depths of anaesthesia, identified using established electroencephalographic criteria. The accelerometer was attached easily and securely to the location of maximal chest wall movement and produced a strong respiratory signal that was detectable in all four anaesthetic stages. Deepening the anaesthesia produced a gradual decrease in RR, a decrease in dominant spectral frequency of the electroencephalogram (EEG) but no change in the heart rate. There was a significant correlation between RR and the dominant spectral frequency of the EEG, indicating that one useful application of the monitor could be to identify anaesthetic stages. The results demonstrate that respiratory parameters can be recorded using a simply constructed, low-cost device and suggest an application in the monitoring of anaesthetic depth.

DNA analysis from stool samples: a fast and reliable method avoiding invasive sampling methods in mouse models of bleeding disorders

Kalippke, K., Werwitzke, S., von Hornung, M., Mischke, R., Ganser, A., Tiede, A. — 2009-09-18

Mouse models with targeted disruptions of coagulation factor genes are used to study disorders of haemostasis such as haemophilia. Standard protocols to obtain biopsies for genotyping in breeding programmes are based on invasive sampling methods such as tail clipping. These procedures imply a high risk of fatal bleeding, especially in haemophilic mouse models. Here we used a non-invasive sampling method obtaining stool samples for DNA isolation in a breeding programme, aiming to introduce targeted disruptions of Fc receptor genes (Fc gamma receptor IIB and III) into the haemophilia A mouse model (factor VIII deficiency). Faecal pellets were reliably obtained from individual mice and high-quality DNA was extracted with a mean yield of 7.1 µg/pellet. Polymerase chain reaction amplification of wild-type and knockout alleles for Fc receptor and factor VIII genes was similar, comparing stool and peripheral blood as the source of genomic DNA. Definite genotype identification was achieved in a first attempt in 336 of 352 analyses (95%). Repeated analysis of homozygous knockout animals confirmed the first result in all cases. No animal was lost due to bleeding from the procedure. In conclusion, DNA isolation from stool is a preferable method for genotyping in laboratory animals, especially in models of bleeding disorders. Avoiding loss of animals due to bleeding implies a substantial improvement in animal welfare by reducing the number of animals used and may also advance the effectiveness of breeding programmes in these disease models.

Modified PAXgeneTM method allows for isolation of high-integrity total RNA from microlitre volumes of mouse whole blood

Krawiec, J A, Chen, H, Alom-Ruiz, S, Jaye, M — 2009-09-18

Analysis of gene expression is often used to evaluate the effects of experimental manipulations in laboratory animals. Blood is a rich source of potential biomarkers, including gene expression information, which may be obtained from whole blood. When compared with the end of a study, when whole blood samples can be easily obtained for gene expression measurements, the limiting volumes of whole blood obtainable from animals during the course of an experiment requires a method for RNA isolation from a minimal volume of whole blood. The PAXgene^TM Blood RNA Extraction System originally designed for isolation of total RNA from 2.5 mL of human whole blood, was modified and successfully used to isolate high-integrity total RNA from as little as 50 µL of mouse whole blood. Fifty microlitres of mouse whole blood yielded an average of 2.3 µg highly intact total RNA, of sufficient quality and quantity allowing for multiple gene expression determinations. The utility of this method was demonstrated by confirming the time- and dose-dependent upregulation of haem oxygenase-1 (Hmox1) mRNA in response to a single injection of cobalt protoporphyrin. The successful isolation of total RNA from small volumes of mouse whole blood can allow for serial sampling on the same animals, thereby reducing the number of animals required for experimentation.

A simple method for confirming correct endotracheal intubation in mice

Watanabe, A, Hashimoto, Y, Ochiai, E, Sato, A, Kamei, K — 2009-09-18

Endotracheal intubation in mice is both a common and important technique. However, it is a difficult procedure because of the small orotracheal size and the success rate is variable. There have been many reports of refined techniques that facilitate intubation but only a few reports have proposed how to verify the proper placement of the endotracheal tube. We describe a novel, safe and reliable method to confirm endotracheal intubation in mice using an extension tube for intravenous infusion. One drop of water was instilled in the extension tube and connected to the end of the catheter used as an endotracheal tube. When the catheter was inserted correctly into the trachea, the water in the extension tube oscillated in synchrony with the movement of the mouse's thorax, indicating correct placement of the catheter. This method was simple, reliable and use materials that are routinely available. This method is helpful for experimental mouse models that require airway access.

Spontaneous thymic lymphomas in the non-obese diabetic/Shi-scid, IL-2R{gamma}null mouse

2009-09-18

The NOD/Shi-scid, IL-2R^null (NOG) mouse is a severely immunodeficient mouse used for the engraftment of human tissues and cells. In this study, 2406 mice (8–62 weeks old, 503 males and 1903 females) were subcutaneously engrafted with human tissues. In 16 mice (12–26 weeks old, 1 male and 15 females), a mass was seen in the anteroventralis of the thorax on gross examination with an incidence of 0.7%. Histologically, the masses were composed of sheets of lymphoblastic cells. A ‘starry sky’ pattern was observed with numerous mitoses. Immunohistochemically the lymphoblastic cells were positive for Thy 1. The lymphoblastic cells were also seen in the spleen, lung, liver, kidney and heart. The gross and histopathological findings led to the diagnosis of spontaneous thymic lymphoma in NOG mice.

Laboratory Animal Science and Technology (Vol. I/II)

Wilkinson, M. — 2009-09-18

The diet board: welfare impacts of a novel method of dietary restriction in laboratory rats

2009-07-07

Laboratory rats are commonly fed ad libitum (AL). Moderate dietary restriction (DR) decreases mortality and morbidity when compared with AL feeding, but there are several obstacles to the implementation of DR. Traditional methods of restricted feeding disrupt normal diurnal eating rhythms and are not compatible with group housing. We have designed a novel method, the diet board, to restrict the feeding of group-housed rats. Animals fed from the diet board had 15% lower body weight than the AL-fed animals at the age of 17 weeks. The welfare effects of diet board feeding were assessed by comparing the stress physiology of diet board fed animals with that of AL-fed animals. Diet board feeding was associated with higher serum corticosterone levels and lower faecal secretion of IgA, suggesting the diet board causes a stress reaction. However, the AL-fed group had larger adrenal glands with higher adrenaline and noradrenaline content than the diet board animals. No gastric ulcers were found in any of the animals at necropsy. The diet board thus appears to cause a stress reaction when compared with AL-fed rats, but no apparent pathology was associated with this reaction. The diet board could help to solve the health problems associated with AL feeding, while allowing the rats to be group-housed and to maintain their normal diurnal eating rhythms. The diet board can also be seen as a functional cage furniture item, dividing the cage into compartments and thus increasing the structural complexity of the environment. In conclusion, the diet board appears to possess refinement potential compared with traditional methods of DR.

Electroencephalographic responses to tail clamping in anaesthetized rat pups

Diesch, T J, Mellor, D J, Johnson, C B, Lentle, R G — 2009-07-07

We investigated electroencephalographic (EEG) responses to tail clamping in lightly anaesthetized rat pups (5–22 days) in order to determine the ontogeny of EEG activity and at what age they may be capable of experiencing pain. Median frequency (F50) and spectral edge frequency (F95) of the power spectrum in the range of 1–30 Hz were determined before and after the application of a noxious stimulus and power spectra were compared by multivariate analysis. There was a postnatal increase in EEG power as, before clamping, pups aged 5–7 days exhibited isoelectric traces, whereas those aged 12–14 days and 21–22 days had intermittent EEG activity where the power in all frequencies was significantly lower at the former than at the latter age. Pups aged 5–7 days exhibited no EEG response to clamping in view of their isoelectric traces. Pups aged 12–14 days showed a significant decrease in F95 (P = 0.002), whereas those aged 21–22 days showed highly significant reduction in F50 and F95 (P = 0.028 and P < 0.001, respectively) as well as changes in EEG power of specific frequencies after clamping. The results and related literature suggest that rat pups aged 5–7 days and younger are not likely to perceive pain and that the ability to perceive pain develops gradually between postnatal ages 12–14 days and 21–22 days.

Reported analgesic administration to rabbits, pigs, sheep, dogs and non-human primates undergoing experimental surgical procedures

Coulter, C A, Flecknell, P A, Richardson, C A — 2009-07-07

Reported analgesic use following experimental surgery is low in rodents and there has been little published information on the frequency of analgesic use in other laboratory species. A structured literature review was conducted to examine analgesic administration in larger laboratory animals. The Scirus search engine was used to identify studies published in peer-reviewed journals that reported carrying out experimental surgery on ‘large’ laboratory animals, specifically rabbits, pigs, sheep, dogs and non-human primates. Seventy-four studies between 2000 and 2001 and 75 studies between 2005 and 2006 were included in the review. There was an increase in the reported administration of systemic analgesics to these species from 50% in 2000–2001 to 63% in 2005–2006. When all agents with analgesic properties were considered (systemic analgesics, local anaesthetics and anaesthetics with analgesic components), the proportion of papers that reported some form of analgesic administration to ‘large’ laboratory animals increased from 86% in 2000–2001 to 89% in 2005–2006. Overall rabbits, pigs, sheep, dogs and non-human primates were more likely to receive analgesics following potentially painful experimental procedures than has been reported in laboratory rodents but analgesic administration to ‘large’ laboratory species is still not optimal.

Determination of the minimum alveolar concentration for halothane, isoflurane and sevoflurane in the gerbil

de Segura, I A G., de la Vibora, J B., Criado, A — 2009-07-07

The present work determined the minimum alveolar concentrations (MAC) of halothane, isoflurane and sevoflurane in adult female gerbils (n = 24). Animals were placed in a chamber for anaesthetic induction before performing tracheal intubation. The tracheal tube was connected to a non-rebreathing circuit with minimal dead space. Body temperature, blood pressure, heart and respiratory rates and end-tidal volatile anaesthetic levels were measured continuously. MAC was determined using a standard noxious stimulus (tail-clamp). All experiments were performed at the same time of the day, body temperature was maintained constant and blood–gas analysis was performed to confirm that values were within normal limits. The mean ± SD MAC values were 1.06 ± 0.11% halothane (n = 8), 1.55 ± 0.08% isoflurane (n = 8) and 2.90 ± 0.12% sevoflurane (n = 7). Cardiovascular parameters at 1x MAC did not differ significantly among anaesthetics but the respiratory rate was significantly higher in the halothane group than in the isoflurane and sevoflurane groups. The SpO₂ values recorded throughout anaesthesia and the pH and partial oxygen pressure values determined at the end of the study did not differ among the studied anaesthetics at 1x MAC. These data suggest that the MAC for halogenated inhalant anaesthetics in gerbils is lower than the average MAC values obtained in rats and mice.

Haemodynamic changes and stress responses of piglets to surgery during total intravenous anaesthesia with propofol and fentanyl

Schoffmann, G, Winter, P, Palme, R, Pollak, A, Trittenwein, G, Golej, J — 2009-07-07

The purpose of the study was to assess the haemodynamic (blood pressure and heart rate) changes and stress responses (serum cortisol and serum amyloid A [SAA] concentrations) to surgery in piglets during total intravenous anaesthesia (TIVA) with propofol and fentanyl. After preanaesthetic medication with intramuscular midazolam (0.5 mg/kg body mass), ketamine (10 mg/kg) and butorphanol (0.5 mg/kg) anaesthesia was induced in five piglets, with intravenous propofol (1 mg/kg) followed by tracheal intubation and mechanical lung ventilation. Soft tissue surgery was performed in the jugular and inguinal regions during TIVA with propofol (8 mg/kg/h) and fentanyl (35 µg/kg/h). Anaesthesia was maintained for 300 min after surgery as the piglets were the control group of a project involving extracorporeal membrane oxygenation. Mean plasma cortisol concentration decreased significantly (P < 0.05) from 59 ± 39.9 nmol/L (mean ± 1 SD) before surgery to 7.5 ± 2.5 nmol/L 300 min after end of surgical procedure. The mean SAA concentrations increased over the same period from 1.6 ± 2.3 µg/mL to 4.2 ± 5.6 µg/mL without statistical significance. The baseline (presurgery) mean arterial pressure (MAP) was 72 ± 9 mmHg compared with 72 ± 11 mmHg 300 min after end of surgery. Neither heart rate nor lactate concentrations changed significantly over the same time points: heart rate was 104 ± 11 and 103 ± 15 beats/min whereas mean lactate concentrations were reduced from 1.14 ± 0.45 mmol/L to 0.90 ± 0.22 mmol/L. Haemodynamic stability, a decrease in serum cortisol and a non-statistically significant rise in mean SAA concentrations suggest that the anaesthetic described suppresses the stress response of piglets to surgery without adverse cardiovascular effects. Therefore, it may prove useful in cardiovascular research.

Effects of streptozotocin-induced diabetes in domestic pigs with focus on the amino acid metabolism

2009-07-07

Streptozotocin (STZ) given intravenously destroys pancreatic beta cells and is widely used in animal models to mimic type 1 diabetes. The effects of STZ on the clinical state of health and metabolism were studied in six high health certified domestic pigs weighing 19 ± 1.3 kg at the start of the experiment. A single STZ dose of 150 mg/kg of body weight successfully induced hyperglycaemia and alterations in amino acid metabolism. Within 9 h after STZ administration, the blood glucose values fell from 5.4–7.5 mmol/L to 0.8–2.2 mmol/L. Hypoglycaemia was treated with 0.5 g glucose/kg body weight. In all pigs, hyperglycaemia was produced 24 h after STZ treatment, and 3 days after STZ injection, the glucose concentration was >25 mmol/L. Mean C-peptide concentration was 0.25 ± 0.16 µg/L since 2 days after STZ injection until the end of the study. The serum concentration of the branched-chain amino acids (BCAA) increased four-fold, and alanine and taurine decreased by approximately 70% and 50%, respectively, after STZ treatment. All but one pig remained brisk and the physical examination was normal except for a retarded growth rate and a reduction of the skeletal muscle. At the end of the study, the pigs were moderately emaciated. Postmortem examination confirmed muscle wasting and a reduction of abdominal and subcutaneous fat. In conclusion, STZ-induced diabetes in pigs fulfils the requirements for a good animal model for type 1 diabetes with respect to clinical signs of the disease and alterations in the carbohydrate and amino acid metabolism.

Blood collection from the sublingual vein in mice and hamsters: a suitable alternative to retrobulbar technique that provides large volumes and minimizes tissue damage

Heimann, M., Kasermann, H. P., Pfister, R., Roth, D. R., Burki, K. — 2009-07-07

Blood examination is a key element in studies of laboratory animals. In rodents, retrobulbar venous plexus puncture is a commonly used method for obtaining a blood sample. Although this technique yields large volumes of blood, the disadvantage is that it can lead to severe tissue damage. The aim of the present study was to develop the puncture of V. sublingualis as a suitable alternative technique for drawing blood in mice and other rodents. In rats, this method has been established for collecting large blood volumes. During the first part of the study, the sublingual bleeding technique was developed for use in mice and hamsters. Guineapigs, however, do not have a sublingual vein; therefore, in this species the method is not possible. In the second part of the study, retrobulbar and sublingual methods were compared using male CD-1 mice. When compared with the retrobulbar method, sublingual venepuncture showed less tissue destruction in mice, with a decreased mean severity in the histological examination. In conclusion, sublingual venepuncture can be recommended as a suitable, alternative blood collection technique, because of the reduced risk of tissue damage in mice and hamsters.

An in vivo magnetic resonance imaging technique for measurement of rat lumbar vertebral body blood perfusion

2009-07-07

The technical feasibility of dynamic contrast-enhanced magnetic resonance imaging (MRI) measurement of rat vertebral blood perfusion using a 1.5 T clinical scanner and a small surface coil was investigated in this study. Nine male 7-month-old Wistar-Kyoto rats were used. Computed tomographic assessment of lumbar vertebral bone mineral density (BMD) and MRI assessment of lumbar vertebral blood perfusion were performed twice with 8 weeks' interval. A 4.7 cm surface coil was used for receiving radiofrequency signal. Gadolinium-DOTA (0.3 mmol/kg) was injected through the tail vein of the rat as a quick bolus and dynamic MRI scan was carried out on the central sagittal plane of the lumbar spine for 8 min with a temporal resolution of 0.6 s. Blood perfusion parameters of wash-in rate, maximum enhancement and wash-out rate were derived from the dynamic MR images. No vertebral body size and vertebral BMD difference was detected between the two time points (P = 0.222 and 0.123, respectively). Wash-in rate was 0.105 (±0.016)% and 0.111 (±0.018)%, maximum enhancement 126.5 (±10.6)% and 129.2 (±13.4)%, and wash-out rate 0.788 (±0.182)% and 0.792 (±0.182)% for the two time points. With these parameters, no difference was found between the two time points with a P value of 0.575, 0.889 and 0.754, respectively. We conclude that dynamic contrast-enhanced MRI measurement of rat vertebral blood perfusion using a 1.5 T clinical scanner and a small surface coil is feasible and reliable. This technique offers a non-invasive means to probe vertebral blood perfusion changes associated with rat disease models.

Longitudinal as well as age-matched assessments of bone changes in the mature ovariectomized rat model

Leitner, M M, Tami, A E, Montavon, P M, Ito, K — 2009-07-07

In the past, bone loss in the ovariectomized (OVX) osteoporotic rat model has been monitored using in vitro micro-computed tomography (micro-CT) to assess bone structure (bone volume/total volume, BV/TV). The purpose of this study was to assess the importance of baseline control and sham groups in 12–16-week-old, reproductively mature rats. Measurements were carried out in a longitudinal and age-matched fashion using newer in vivo peripheral quantitative computed tomography (pQCT), which measures apparent bone mineral density (BMD). BMD was measured at the distal femoral metaphysis of 12-week-old female Wistar rats with pQCT. Subsequently, animals were either OVX or sham operated, and pQCT measurements were repeated four weeks later. Then, all rats were euthanized and in vitro BMD and BV/TV were obtained by micro-CT imaging. Results from three consistently differentiated regions of interest showed that there was significant bone loss and growth during the four weeks in the OVX and sham group, respectively. Taking this into account, i.e. a posteriori superimposing growth to loss, no differences resulted between BMD values measured in a longitudinal fashion with pQCT and that measured in comparison with an age-matched sham group with micro-CT and pQCT. In addition, there was a strong linear correlation between BMD measured with pQCT and BV/TV obtained from micro-CT. In conclusion, this outcome provides new insights into individual bone changes due to OVX and growth in Wistar rats during the age period of 12–16 weeks, which is often applied in osteoporosis research as the ‘mature’ rat model. Data can be used as baseline information upon which future in vivo study designs with this rat model can refer to reduce and minimize animal use.

A new small animal model for the study of spine fusion in the sand rat: pilot studies

Gruber, H. E, Gordon, B., Williams, C., Ingram, J. A, Norton, H J., Hanley, E. N — 2009-07-07

Spine fusion is used to treat traumatic or degenerative lumbar instability in the cervical or lumbar spine. Although degenerative radiological changes in discs adjacent to a fusion have been well-recognized, histopathological changes in adjacent discs have not been studied and are poorly understood. An economical small animal model for lumbar fusion would be a useful research tool. Study objectives were to: (1) develop a model of non-instrumented spine fusion in the sand rat, a rodent with spontaneous, age-related disc degeneration; (2) use radiological–histological analyses to study fusion and disc degeneration in cranial and caudal discs adjacent to the fusion. Studies were approved by our Institutional Animal Care and Use Committee. A small segment of outer annulus tissue was surgically removed from lumbar discs, radiographs obtained and the animal allowed to recover and age. At surgical harvest, radiographs of 28 spine fusion specimens were scored and statistically analysed for adjacent disc space narrowing, wedging, endplate sclerosis and irregular disc margins. At harvest, the incidence of these radiological indices of disc degeneration were significantly greater than at time of surgery. Pilot studies presented here indicate that this model provides a novel addition to basic science approaches studying the clinically important topic of spinal fusion and adjacent segment changes. The resulting fusion site can be assessed statistically with radiological scoring to determine development/progression of disc degeneration in adjacent segments, and correlative histological features can be examined. The sand rat is well-suited for use in spine fusion studies because of reliable/reproducible progression of disc degeneration and the favourable economics of small rodent studies.

An interview study of phenotypic characterization of genetically-modified mice

Thon, R, Vondeling, H, Lassen, J, Hansen, A K, Ritskes-Hoitinga, M — 2009-07-07

An interview study was carried out with the aim of clarifying the reasons for the limited use of phenotypic characterization of genetically-modified mice (GMM) and identifying issues hindering its implementation. A total of 15 users of GMM participated in semi-structured face-to-face interviews, which were audio-taped and transcribed. The results were extracted using content analysis by theme. The investigation confirmed that few animals were systematically phenotyped and an observational approach was found to be widespread. The primary interest of the interviewees was phenotyping for impaired animal welfare. The concept of phenotyping was widely understood and perceived as a scientific advantage. The comprehensiveness of the protocols and the resources required for phenotyping were seen as problematic. All participants addressed this issue, be it regarding lack of time, money or expertise. Also, among the negative statements were worries about the capability of the available protocols to produce the information needed by the individual scientist. Phenotyping was predicted to become much more widespread in the future and its success was expected to depend on the development of reliable, fast and inexpensive methods. The study identified different aims of phenotyping and the suitability of the published protocols for these purposes was discussed. The contradiction between the limited use of characterization and its advantages was also discussed and proposals for the improvement of future phenotyping strategies are formulated.

Limited sampling strategy in rats to predict the inhibited activities of hepatic CYP3A

Zhu, X.-H., Jiao, J.-J., Zhang, C.-L., Lou, J.-S., Liu, C.-X. — 2009-07-07

The present study was undertaken in order to evaluate feasibility of a limited sampling strategy (LSS) to predict the systemic clearance of midazolam (MDZ), which is a hepatic CYP3A activity phenotyping probe. Groups of rats pretreated with or without serial doses of ketoconazole, which is a selective inhibitor on CYP3A, were used as training set. Linear regression analysis and a Jack-knife validation procedure were performed based on plasma MDZ concentrations at specific time points after sublingual vein injection of MDZ to establish the most informative LSS equations for accurately estimating the clearance of MDZ. Another group of rats in the same setting was used as the validation set to confirm the individual values of estimated clearance (Cl_est) that were derived from the predictive equations developed in the training set. LSS that were derived from one, two or three sampling times, namely 90 min, 60–90 min, 30–60–90 min and 30–60–120 min, gave the best correlation and acceptable errors between the values of observed clearance (Cl_obs) and Cl_est and were chosen to evaluate hepatic CYP3A activity. Our results supported the hypothesis that using limited plasma sampling is simpler than the usual method of estimating CYP3A phenotyping by predicting the systemic clearance of MDZ when the hepatic activity of CYP3A is reduced in the rat. This experimental design offers opportunities to reduce animal use in the study of drug metabolism.

Cardiac arrhythmias related to continuous thermodilution cardiac output measurement in an anaesthetized sheep

Coppens, P, Moens, Y — 2009-07-07

A case of cardiac arrhythmias related to continuous thermodilution cardiac output (CCO) is reported. A sheep anaesthetized for experimental purpose was instrumented with a special Swan-Ganz catheter-type to be used for CCO measurements. One hour after starting the CCO monitoring, isolated ventricular extrasystoles were noticed on the electrocardiogram with an increasing frequency. Subsequently bursts of extrasystoles occurred. Atrioventricular dissociation was also observed. The peaks of temperature of the thermal filament were within the normal range and their presence was noticed when arrhythmias appeared. Mean blood pressure and cardiac output did not change during this episode. When the CCO was switched off, no more arrhythmias were observed. The CCO Swan-Ganz by itself did not generate any arrhythmia. The sheep recovered uneventfully. When arrhythmias occur during anaesthesia where CCO is used, a thermal filament induced origin of the arrhythmia must be considered.

A new method for induction and synchronization of oestrus and fertile ovulations in mice by using exogenous hormones

Pallares, P, Gonzalez-Bulnes, A — 2009-07-07

Four consecutive experiments were conducted to design and establish the effectiveness of different protocols for induction and synchronization of oestrus and ovulation in mice. Results showed that the highest synchronization degree and the highest fertility rates were obtained using two intraperitoneal doses of 0.5 µg of cloprostenol, three days apart, plus a single subcutaneous dose of 3 µg of progesterone coincidentally with the first injection of cloprostenol. Of the main advantages of the new method, we have to highlight the short time elapsed for appearance, and the high degree of synchronization of oestrus and ovulations (almost 100% of the animals responding to the treatment in 48 h; 78.4% with fertile mates at 24 h), plus the high fertility rate obtained after a programmed mating (100%). Overall, these yields are superior to those obtained by classical methods based on the use of male pheromones; hence the proposed protocol arises as an adequate alternative for reproductive management in mice.

Mycobacterium gordonae infection in a colony of African clawed frogs (Xenopus tropicalis)

Sanchez-Morgado, J. M, Gallagher, A., Johnson, L. K — 2009-07-07

Mycobacterium gordonae is an occasional human pathogen associated with cutaneous infections and nodular granulomatous skin lesions. A case of cutaneous nodular infection caused by M. gordonae in a colony of African clawed frogs (Xenopus tropicalis) is described and confirms this organism to be an opportunistic frog pathogen.

Clostridial typhlitis associated with topical antibiotic therapy in a Syrian hamster

Alworth, L, Simmons, J, Franklin, C, Fish, R — 2009-07-07

A Syrian hamster that had been treated with topical antibiotic ointment for 10 days following injuries sustained during fighting was presented moribund. Postmortem and microscopic examinations revealed lesions consistent with clostridial typhlitis and enteritis. Anaerobic culture of caecal contents resulted in the isolation of two Clostridium species, and caecal contents contained Clostridium difficile enterotoxins. Based on these findings, a diagnosis of acute C. difficile enterotoxaemia was made. This report discusses the pathogenesis of C. difficile enterotoxaemia and the potential role of topical antibiotic ointment therapy in initiating the disease.

Refinements in husbandry, care and common procedures for non-human primates: Ninth report of the BVAAWF/FRAME/RSPCA/UFAW Joint Working Group on Refinement

2009-03-13

Preface

Whenever animals are used in research, minimizing pain and distress and promoting good welfare should be as important an objective as achieving the experimental results. This is important for humanitarian reasons, for good science, for economic reasons and in order to satisfy the broad legal principles in international legislation. It is possible to refine both husbandry and procedures to minimize suffering and improve welfare in a number of ways, and this can be greatly facilitated by ensuring that up-to-date information is readily available. The need to provide such information led the British Veterinary Association Animal Welfare Foundation (BVAAWF), the Fund for the Replacement of Animals in Medical Experiments (FRAME), the Royal Society for the Prevention of Cruelty to Animals (RSPCA) and the Universities Federation for Animal Welfare (UFAW) to establish a Joint Working Group on Refinement (JWGR) in the UK. The chair is Professor David Morton and the secretariat is provided by the RSPCA. This report is the ninth in the JWGR series. The RSPCA is opposed to the use of animals in experiments that cause pain, suffering, distress or lasting harm and together with FRAME has particular concerns about the continued use of non-human primates. The replacement of primate experiments is a primary goal for the RSPCA and FRAME. However, both organizations share with others in the Working Group, the common aim of replacing primate experiments wherever possible, reducing suffering and improving welfare while primate use continues. The reports of the refinement workshops are intended to help achieve these aims.

This report produced by the British Veterinary Association Animal Welfare Foundation (BVAAWF)/Fund for the Replacement of Animals in Medical Experiments (FRAME)/Royal Society for the Prevention of Cruelty to Animals (RSPCA)/Universities Federation for Animal Welfare (UFAW) Joint Working Group on Refinement (JWGR) sets out practical guidance on refining the husbandry and care of non-human primates (hereinafter primates) and on minimizing the adverse effects of some common procedures. It provides a valuable resource to help understand the physical, social and behavioural characteristics and needs of individual primates, and is intended to develop and complement the existing literature and legislative guidelines. Topics covered include refinements in housing, husbandry and common procedures such as restraint, identification and sampling, with comprehensive advice on issues such as primate communication, assessing and facilitating primate wellbeing, establishing and maintaining social groups, environmental and nutritional enrichment and animal passports. The most commonly used species are the key focus of this resource, but its information and recommendations are generally applicable to other species, provided that relevant individual species characteristics are taken into account.

Key issues concerning environmental enrichment for laboratory-held fish species

Williams, T D, Readman, G D, Owen, S F — 2009-03-12

An improved knowledge and understanding of the fundamental biological requirements is needed for many of the species of fish held in captivity and, without this knowledge it is difficult to determine the optimal conditions for laboratory culture. The aim of this paper is to review the key issues concerning environmental enrichment for laboratory-held fish species and identify where improvements are required. It provides background information on environmental enrichment, describes enrichment techniques currently used in aquatic ecotoxicology studies, identifies potential restrictions in their use and discusses why more detailed and species-specific guidance is needed.

A review of the need and possible uses for genetically standardized Atlantic salmon (Salmo salar) in research

Grimholt, U, Johansen, R, Smith, A J — 2009-03-12

Large numbers of Atlantic salmon (Salmo salar) are used as research animals in basic research and to solve challenges related to the fish-farming industry. Most of this research is performed on farmed animals provided by local breeders or national breeding companies. The genetic constitution of these animals is usually unknown and highly variable. As a result, large numbers of fish are often needed to produce significant results, and results from one study are often impossible to reproduce in another facility. The production of standardized salmon could in many cases reduce the number of animals used in research and at the same time provide more reproducible results. This paper provides an overview of the methods available for the production of standardized Atlantic salmon, and discusses the pros and cons of each technique. The use of zebrafish and other well-defined laboratory fish species as a model for salmon is also discussed. Access to genetically defined fish would greatly benefit the scientific community, in the same way as genetically defined lines of rodents have revolutionized mammalian research.

Echocardiographic assessment of global left ventricular function in mice

2009-03-12

Doppler-echocardiographic assessment of cardiovascular structure and function in murine models has developed into one of the most commonly used non-invasive techniques during the last decades. Recent technical improvements even expanded the possibilities. In this review, we summarize the current options to assess global left ventricular (LV) function in mice using echocardiographic techniques. In detail, standard techniques as structural and functional assessment of the cardiovascular phenotype using one-dimensional M-mode echocardiography, two-dimensional B-mode echocardiography and spectral Doppler signals from mitral inflow respective aortal outflow are presented. Further pros and contras of recently implemented techniques as three-dimensional echocardiography and strain and strain rate measurements are discussed. Deduced measures of LV function as the myocardial performance index according to Tei, estimation of the mean velocity of circumferential fibre shortening, LV wall stress and different algorithms to estimate the LV mass are described in detail. Last but not least, specific features and limitations of murine echocardiography are presented. Future perspectives in respect to new examination techniques like targeted molecular imaging with advanced ultrasound contrast bubbles or improvement of equipment like new generation matrix transducers for murine echocardiography are discussed.

A novel dietary restriction method for group-housed rats: weight gain and clinical chemistry characterization

2009-03-12

Laboratory rodents are usually fed ad libitum. Moderate dietary restriction decreases mortality and morbidity compared with ad libitum feeding. There are, however, problems in achieving dietary restriction. Traditional methods of restricted feeding may interfere with the diurnal rhythms of the animals and are not compatible with group-housing of rodents. We have invented a novel method, the diet board, for restricting the feed intake of laboratory rats. The use of the diet board moderately decreased weight gain of rats when compared with ad libitum-fed animals. The diet board retarded skeletal growth only minimally, whereas major differences were found in body fat depositions. Serum free fatty acid, triglyceride and cholesterol values were lower in diet-restricted rats, while the opposite was true for serum creatine kinase. There were no differences in total protein, albumin or alanine aminotransferase. Moreover, differences in interindividual variances in parameters were not detected between the groups; hence this study could not combine the diet board with reduction potential. The diet board provides mild to moderate dietary restriction for group-housed rats and is unlikely to interfere with the diurnal eating rhythm. The diet board can also be seen as a cage furniture item, dividing the open cage space and increasing the structural complexity of the environment. In conclusion, the diet board appears to possess refinement potential when compared with traditional methods of dietary restriction.

Reported analgesic and anaesthetic administration to rodents undergoing experimental surgical procedures

Stokes, E L, Flecknell, P A, Richardson, C A — 2009-03-12

A structured literature review was carried out to assess recent trends in the administration of analgesics and anaesthetics to laboratory rats and mice undergoing surgical procedures. The ScienceDirect database was used to systematically identify studies published in peer-reviewed journals over two periods (2000–2001 and 2005–2006), 86 studies from each time period were included in the review. The total number of animals that underwent surgery, species used, type of procedure, anaesthetic regimen and analgesic administration were noted for each study. There was an increase in the reported administration of systemic analgesics from 10% in 2000–2001 to 20% in 2005–2006. Buprenorphine was the most commonly reported analgesic in both periods (2000–2001: 78%, 2005–2006: 35%) and reporting the use of non-steroidal anti-inflammatory drugs increased from 11% to 53%. There was also a change in reported anaesthetic practices, notably a decrease in the use of pentobarbital and an increase in the use of isoflurane and ketamine/xylazine. Although reported administration of analgesics has increased and there has been some refinement in the selection of anaesthetic agents used, the findings of this review suggest that there is still significant scope for improvement with respect to the perioperative care of laboratory rodents.

Reducing variation in a rabbit vaccine safety study with particular emphasis on housing conditions and handling

2009-03-12

This paper describes the results of a study of the effects of modified housing conditions, conditioning and habituation on humans using a rabbit model for monitoring whole-cell pertussis vaccine (pWCV)-induced adverse effects. The study has been performed with reference to previous vaccine safety studies of pWCV in rabbits in which results were difficult to interpret due to the large variation in experimental outcome, especially in the key parameter deep-body temperature (T_b). Certain stressful laboratory conditions, as well as procedures involving humans, e.g. blood sampling, inoculation and cage-cleaning, were hypothesized to cause this large variation. The results of this study show that under modified housing conditions rabbits have normal circadian body temperatures. This allowed discrimination of pWCV-induced adverse effects in which handled rabbits tended to show a dose-related increase in temperature after inoculation with little variance, whereas non-handled rabbits did not. Effects of experimental and routine procedures on body temperature were significantly reduced under modified conditions and were within the normal T_b range. Handled animals reacted less strongly and with less variance to experimental procedures, such as blood sampling, injection and cage-cleaning, than non-handled rabbits. Overall, handling had a positive effect on the behaviour of the animals. Data show that the housing modifications have provided a more robust model for monitoring pWCV adverse effects. Furthermore, conditioning and habituation of rabbits to humans reduce the variation in experimental outcome, which might allow for a reduction in the number of animals used. In addition, this also reduces distress and thus contributes to refining this animal model.

Contemporary prevalence of infectious agents in laboratory mice and rats

Pritchett-Corning, K. R, Cosentino, J., Clifford, C. B — 2009-03-12

Periodic health screening of rodents used in research is necessary due to the consequences of unwanted infections. One determinant of the risk of infection for any given agent is its prevalence; other factors being equal, a prevalent agent is more likely than a rare one to be introduced to a research facility and result in infection. As an indicator of contemporary prevalence in laboratory populations of rats and mice, the rate of positive results in the samples received at a major commercial rodent diagnostic laboratory was compiled for this paper. Although samples from laboratory rodent vendors have been excluded, results are tabulated from samples from more than 500,000 mice and 80,000 rats submitted over several years from pharmaceutical, biotechnology, academic, and governmental institutions in North America and Europe, allowing meaningful determination of which agents are common in the research environment versus which agents are rare. In mice, commonly detected infectious agents include mouse norovirus, the parvoviruses, mouse hepatitis virus, rotavirus, Theiler's murine encephalomyelitis virus, Helicobacter spp., Pasteurella pneumotropica, and pinworms. In rats, commonly detected infectious agents include ‘rat respiratory virus’, the parvoviruses, rat theilovirus, Helicobacter spp., P. pneumotropica, and pinworms. A risk-based allocation of health-monitoring resources should concentrate frequency and/or sample size on these high-risk agents, and monitor less frequently for the remaining, lower-risk, infectious agents.

Kinetic analysis of apolipoproteins in postprandial hypertriglyceridaemia rabbits

Hata, M, Ito, T, Ohwada, K — 2009-03-12

The postprandial hypertriglyceridaemia (PHT) rabbit, developed as a new animal model of metabolic syndrome, is characterized by PHT, central obesity and glucose intolerance. For detailed investigation of lipid metabolism characteristics in PHT rabbit, the plasma levels of apolipoproteins A-I, B, C-II, C-III and E were measured. Movements of apolipoproteins B100 and B48 were investigated using sodium dodecyl sulphate–polyacrylamide gel electrophoresis to determine whether postprandially increased triglyceride is exogenous or endogenous. The level of apolipoproteins A-I, B, C-II and E were increased in PHT rabbit after feeding. Apolipoproteins B100 and B48 were detected in the plasma fraction of d < 1.006 g/mL of the PHT rabbit. The postprandial increase in apolipoprotein B in the PHT rabbit reflects a numerical increase in lipoprotein particles in the blood; the increase in apolipoproteins C-II and E suggests some disturbance in lipoprotein catabolism. Apolipoprotein B48 was detected postprandially in PHT rabbits. These results suggest that delayed catabolism of exogenous lipids caused the retention of chylomicron remnants in the blood. Results also suggest that activities of the lipolytic enzyme lipoprotein lipase and hepatic triglyceride lipase were deficient and that the hepatic uptake of exogenous lipoproteins was delayed in the PHT rabbit. Especially, for examining remnant hyperlipoproteinaemia in humans, PHT rabbit is an excellent animal model for hypertriglyceridaemia research.

Effects of environmental enrichment on the amyotrophic lateral sclerosis mouse model

2009-03-12

The manner in which an animal's environment is furnished may have significant implications for animal welfare as well as research outcomes. We evaluated four different housing conditions to determine the effects of what has been considered standard rodent enrichment and the exercise opportunities those environments allow on disease progression in the amyotrophic lateral sclerosis mouse model. Forty-eight copper/zinc superoxide dismutase mice (strain: B6SJL-TgN [SOD1-G931]1Gur) (SOD1) and 48 control (C) (strain: B6SJL-TgN[SOD1]2Gur) male mice were randomly assigned to four different conditions where 12 SOD1 and 12 C animals were allotted to each condition (n = 96). Conditions tested the effects of standard housing, a forced exercise regime, access to a mouse house and opportunity for ad libitum exercise on a running wheel. In addition to the daily all-occurrence behavioural sampling, mice were weighed and tested twice per week on gait and Rotor-Rod^TM performance until the mice reached the age of 150 days (C) or met the criteria for our humane endpoint (SOD1). The SOD1 mice exposed to the forced exercise regime and wheel access did better in average lifespan and Rotor-Rod^TM performance, than SOD1 mice exposed to the standard cage and mouse house conditions. In SOD1 mice, stride length remained longest throughout the progression of the disease in mice exposed to the forced exercise regime compared with other SOD1 conditions. Within the control group, mice in the standard cage and forced exercise regime conditions performed significantly less than the mice with the mouse house and wheels on the Rotor-Rod^TM. Alpha motor neuron counts were highest in mice with wheels and in mice exposed to forced exercise regime in both mouse strains. All SOD1 mice had significantly lower alpha neuron counts than controls (P < 0.05). These data show that different enrichment strategies affect behaviour and disease progression in a transgenic mouse model, and may have implications for the effects of these strategies on experimental outcomes.

The effect of defect localization on spontaneous repair of osteochondral defects in a Gottingen minipig model: a retrospective analysis of the medial patellar groove versus the medial femoral condyle

Jung, M., Breusch, S., Daecke, W., Gotterbarm, T. — 2009-03-12

Various animal models for experimental osteochondral defect healing have been used in orthopaedic research. Two main defect locations were chosen: the patellar groove or the central part of the medial femoral condyles (MFC). To date, it is not clear whether both locations display similar patterns in critical size osteochondral defect healing. We retrospectively analysed both locations in our minipig model hypothesizing that they show similar healing pattern. Thirty-five defects were analysed after three or 12 months. Osteochondral defects were 10 mm deep and 6.3 mm (MFC, n = 19) in diameter or 8 mm and 5.4 mm, respectively (trochlear groove [TG], n = 16). Semi-quantitative histological scoring and histomorphological evaluation were carried out. Both defect locations showed fillings of fibrous and fibrocartilage-like repair tissue. The osseous defect was closed by endochondral bone formation in the MFC. Semi-quantitative scoring did not show differences, whereas qualitative histomorphological analysis more frequently showed cartilaginous repair tissue in MFC defects. There was more frequent subchondral bone cyst formation in MFC location (P = 0.05), TG defects resulted in lower postoperative pain. Both defect localizations are suitable for studies on osteochondral healing. Since regenerating with less hyaline-like repair tissue and less subchondral cyst formation, TG is more favourable for experimental osteochondral defect healing in this model.

Experimental middle ear surgery in rabbits: a new approach for reconstructing the ossicular chain

Stieve, M, Hedrich, H J, Battmer, R D, Behrens, P, Muller, P, Lenarz, T. — 2009-03-12

This experimental animal study aimed at evaluating a new prosthesis to replace the ossicular chain; we developed a new technique for surgical implantation into the middle ear of rabbits. The rabbit middle ear is, owing to the relative anatomical dimensions involved, an ideal environment for implantation procedures involving the ossicles, as the surgical conditions are similar to those of the human middle ear. This study included a total of 34 approximately six-month-old female white rabbits (New Zealand) weighing between 3.2 and 4.4 kg. The implants used were constructed of ceramic materials (titania, TiO₂) of various pore sizes. Directly prior to implanting the total ossicular reconstruction prostheses (TORPs), as well as at 28, 84 and 300 days after implantation, electric response audiometry was used to determine the hearing thresholds of the animals (bone conduction; click stimulus nHL). An erbium:YAG laser was used to excise the original ossicular chain. Following implantation, we were unable to detect any stenosis of the outer ear canal or perforation of the tympanic membrane. The conductive hearing threshold was in the range of 4.21 ± 6.68 dB nHL (n = 131). The hearing level showed no significant difference before and after surgery (P < 0.05).

Soft-hydrothermal processing of red cedar bedding reduces its induction of cytochrome P450 in mouse liver

2009-03-12

Red cedar-derived bedding materials cause changes in cytochrome P450-dependent microsomal enzyme systems in laboratory animals. We examined the effect of essential oil of red cedar (EORC), as well as the effect of bedding from which it had been removed, on the hepatic expression cytochrome P450s in mice. EORC was obtained from liquid extracts of red cedar bedding by a soft-hydrothermal process and was administered orally to mice. Between days 1 and 2 after administration, hepatic P450s were significantly induced as follows: CYP3As, 7.1x; CYP1As, 1.6x; CYP2E1, 1.5x; CYP2Cs, 1.6x. A housing study of mice indicated that red cedar bedding increased the levels of these P450s in mouse liver, whereas mice housed in cedar bedding from which EORC had been removed (ST-cedar bedding) showed significantly lower levels of P450s, especially CYP3As, CYP1As and CYP2E1. Soft-hydrothermal processing partially removed many components of EORC. In particular, several volatile sesquiterpenes, naphthalene-derived aromatics and 4,4-dimethyl-13-androst-5-ene were decreased in the ST-cedar bedding, suggesting that these may be responsible for P450 induction. This study demonstrated that the removal of these volatile compounds by soft-hydrothermal processing can decrease the hepatic P450-inducing effect of red cedar bedding.