137-148
Comparison of and Habituation to Four Common Methods of Handling and Lifting of Rats with Cardiovascular Telemetry
by Zivile Baturaite, Hanna-Marja Voipio, Osvaldas Ruksenas, Marja Luodonpää, Hanna Leskinen, Neringa Apanaviciene, Timo Nevalainen

Daily routines in the animal house may influence the results and interpretation of experiments. Handling is one such routine since it is necessary to immobilize animals for even minor procedures. This study assesses the influence of four common handling and lifting methods on cardiovascular parameters (blood pressure, heart rate) and locomotor activity of Sprague-Dawley and Wistar rats. Seven rats were implanted with radio-telemetry transmitters. After a recovery period, they were housed in groups of three with two intact rats. Each instrumented rat was subjected to the four methods of handling and lifting (scruff, encircling, plastic cone, lifting and holding by the tail on the arm) and, the same method was repeated during three consecutive weekdays. The method was changed every second week in a rotational order. Handling increased cardiovascular parameters for about 30 min, these changes being statistically significant (p < 0.05) as compared to control conditions immediately before the procedure. With holding by the scruff, the response duration decreased significantly from day one to days two and three, indicative of habituation to this procedure. Rats did not habituate to the cone handling, nor to encircling or lifting and holding by the tail; with the restraint cone being apparently the most disturbing. In conclusion, we have found that there are measurable differences in the impact of various handling and lifting methods and the correct choice permits refinement (one of the “Three Rs” of animal usage) of the procedure. Cardiovascular telemetry appears to be a useful method when used for refining procedures on animals, such as handling and lifting.

153-158
Influence of Cyclophosphamide on the Haematological Profile of Laboratory Bred African Soft-furred Rats (Mastomys natalensis)
by JM Kagira, NW Maina, JK Thuita, M Ngotho & J Hau

The African soft-furred rat (Mastomys natalensis) has been shown to be a possible model for propagation of Trypanosoma brucei gambiense. This study aimed at determining the baseline biological reference values and reproductive data of a laboratory bred Mastomys colony, which was established at TRC. In addition, the effect of cyclophosphamide (an immunosuppressant) treatment (s) on the haematological profile was investigated. The mean gestation period was 23 days and the mean litter size was eight. At birth, the pups weighed 2.4±0.23 g and the weights increased to 78.0±10.6 g in males and 53.9±4.5 g in females by 90 days. The mean haematological values were significantly (p<0.05) higher in adults than juveniles. However, there was no statistical difference of haematological values between the sexes. Cyclophosphamide treatment caused a macrocytic hypochromic anaemia, which was noted 24 hours after treatment and was more severe in animals treated more than once. Thus, in studies involving a disease that causes anaemia, repeated cyclophosphamide treatment should be limited. Our study is a contribution to the clinical and biological characterization of the disease pattern in this preferred rodent model of T. b. gambiense.

161-166
The Influence of Exposure to Stress of Pregnant Rats on the Adrenal Gland Structure of their Offspring. An Unbiased Stereological Study
by Ali Noorafshan, Mohammad Karimipoor, Soghra Bahmanpoor, Farzaneh Dehghani

Many factors may interact with normal differentiation and growth of tissues and cells. Stress might be experienced during pregnancy and it has been shown that stress may cause low birth weight. The effect of prenatal manipulations on the HPA axis has been focused on physiological and biochemical alteration of the adrenal gland. A stereological examination of the influences of prenatal stress on the structure of the developing adrenal gland of one day and 21 day-old rats has now been performed. In this study experiments were conducted to test the hypothesis that exposure to restraint stress during pregnancy in rat results in structural changes in the developing adrenal gland of their pups. Female rats were exposed to restraint stress from the first day of pregnancy throughout gestation. Male offspring of stressed rats (PS= experimental) and of unstressed mothers (C= control) who were one day and 21 days of age were selected. Their body weight (BW), crown-rump length (CRL), biparietal diameter (BPD), volume of the gland and the cortical layers and medulla were estimated using stereological methods. The results showed that the prenatal stress led to a decrease in BW, but CRL and BPD remained unchanged. Also, a significant increase in volume of the adrenal gland and cortical layers in one day and 21 day-old offspring were observed. The volume of the medulla of the adrenal gland of neonate rats remained unchanged but the volume of the medulla in 21 day-old rats was decreased. Therefore, it can be concluded that prenatal stress alters the structure of the developing adrenal gland.

175-177
Mouse Lung Conditioned Medium Induces Short Term Erythropoiesis in Mouse Long Term Bone Marrow Culture System
by M Salimian*, M Saffari, T Khamehchian, H Nikzad, A Aliasgharzadeh

Dexter-type long-term bone marrow culture is a myelopoietic culture system that allows maintenance of mouse and human hematopoiesis in vitro over a period of several months. In mouse unperturbed long-term bone marrow culture, erythropoiesis activity is limited to the production of immature erythroid progenitors (BFU-E) from primitive hematopoietic stem cells. In this study the effects of mouse lung conditioned medium (MLCM) as a source of myeloid growth factors, on long-term mouse bone marrow cultures was studied. Numbers of cells in adherent and non-adherent layers of cultures were counted weekly and the morphological appearances of mature cells that were produced in non-adherent layers were analyzed. In the presence of MLCM, mature nonnucleated and hemoglobinized red blood cells were produced in the nonadherent layers of the cultures.

183-187
Post-graduate Teaching in Microsurgery Using a Combination of Alternative Non-animal and In Vivo Methods
by Peter J.A. Bollen, René Remie, Willem Kloots, Lis Barfred, Søren Salomon& Merel Ritskes-Hoitinga

Over recent years alternative, non-animal methods have been developed for use in microsurgical training courses, for the training of suture techniques, dexterity and hand-eye coordination. Nevertheless, it is still hard to imagine that alternative methods will be able to replace in vivo teaching completely, because properties of biological tissue are different from those of latex or other artificial materials. Also the dynamics of biological processes, such as a pulsating bloodstream, are difficult to simulate in alternative models. However, the number of animals used in microsurgical training courses can be limited by combining non-animal methods with in vivo training. Moreover, individual techniques can be improved greatly by using non-animal methods. In a five-day introductory course in microsurgical and experimental techniques at the Biomedical Laboratory, University of Southern Denmark, we have limited the average number of animals to one rat per day per participant, by using alternative non-animal methods extensively on the first course day. Nearly all participants were able to produce a successful end-to-end anastomosis of the femoral artery during the second or third day of the course. The alternative methods used included a training device for microsurgical anastomosis and the MD PVC-Rat, an artificial rat with life-size latex abdominal vessels.