A comparison of two intramuscular doses of xylazine–ketamine combination and tolazoline reversal in llamas

The anesthetic and cardiorespiratory effects of a low dose (LD, 0.4 mg kg^?1 xylazine and 4 mg kg^?1 ketamine) and a high dose (HD, 0.8 mg kg^?1 xylazine and 8 mg kg^?1 ketamine) of IM xylazine–ketamine combination were compared in a randomized cross‐over study using six castrated male llamas. Three llamas in each dosage group (LDT, HDT) were assigned to receive IM tolazoline (2 mg kg^?1) after 30 minutes of recumbency. All IM injections were given in the semitendinosus or semimembranosus muscles. Pulse, respiratory rate, and indirect arterial blood pressure were recorded every 10 minutes, and hemoglobin oxygen saturation was recorded every 5 minutes during lateral recumbency. Samples for arterial blood gas analysis were collected 5 minutes following recumbency and every 30 minutes thereafter. Base‐to‐apex ECG was monitored continuously. Analgesia was evaluated every 5 minutes by both a 30 minutes skin pinch and a needle prick of the toe. Most llamas breathed room air throughout anesthesia. Two llamas that developed severe hypoxemia (SpO₂ < 75%) received 5 minutes of nasal oxygen insufflation, but were maintained on room air for the rest of the anesthetic period. anova for repeated measures and Tukey's test were used to analyze cardiorespiratory data. Fischer's exact test was used to compare the ability of each to provide >30 minutes of lateral recumbency and analgesia. A p‐value < 0.05 was considered significant. Both dosages provided reasonably rapid induction following injection (LD: 10.8 ± 6.3 minutes; HD: 5.0 ± 1.1 minutes; p = 0.07). Duration of lateral recumbency and analgesia were 34.7 ± 6.7 and 27.3 ± 4.6 minutes, respectively, in the LDT llamas. None of the three remaining LD llamas remained in lateral recumbency for longer than 12 minutes. Duration of lateral recumbency and analgesia were 87.3 ± 18.5 and 67.7 ± 16.0 minutes, respectively, for the HD llamas that did not receive tolazoline. The HDT llamas were recumbent for a significantly shorter time (43.3 ± 0.6 minutes; p = 0.05). The ability to provide >30 minutes of recumbency and analgesia was better in the HD group (6/6) than in the LD group (2/6) (p = 0.03). No differences between dosages were seen in pulse rate, respiratory rate, or arterial pressures. No ECG abnormalities were seen. Transient hypoxemia was seen in the first 10 minutes of lateral recumbency in the HD group by both hemoglobin oxygen saturation (84 ± 9.5%) and by blood gas PaO₂ (44.5 ± 5.8 mm Hg). It was concluded that the HD provided more consistent results than the LD, but induced transient hypoxemia. Tolazoline shortened the recovery time in llamas receiving the HD.     

Histological and Immunohistochemical Characterization of Equine Anovulatory Haemorrhagic Follicles (AHFs)

Anovulatory haemorrhagic follicles (AHFs) are often the reason for ovulation failure in the mare. As the underlying factors that lead to AHF development are not well understood, it was of interest to investigate the vascularization of AHFs compared with normal follicles and corpora lutea (controls). In the present study, the ovarian cell populations investigated immunohistochemically included granulosa and luteal cells as well as various vascular structures. None of these cell types showed differences in the expression of vascular endothelial growth factor A (VEGF-A) between control ovaries containing normal follicles and corpora lutea and ovaries with AHFs. In contrast, a considerable reduction in the proportion of Flk-1-expressing cells, together with a decreased intensity of staining, was apparent in the AHFs. This greatly reduced expression of Flk-1 in the luteinized cells and the vascular structures of AHFs may lead to a distinct decrease in the potential pro-angiogenic activity of VEGF-A in these structures compared with the situation in normal follicles and corpora lutea. Furthermore, the authors suspect that the distinct expression of angiopoietin2 and VEGF-A seen in the cells within the inner fibrous layers of the AHFs was caused by hypoxia resulting from deficient vascularization, as suggested by the irregularity of the capillaries present in the luteinized wall of the AHF. In addition, whereas LH-receptor (LH-R) expression occurred uniformly in all stages of development of the corpora lutea in normal control ovaries, there was highly variable labelling for LH-R in all the AHFs examined, thereby indicating a possible numerical deficiency of LH-receptors in AHFs. The authors concluded that, despite the apparent expression of sufficient VEGF-A in the AHFs allows ovulation and corpus luteum formation, a relative lack of receptor, Flk-1, effects the pro-angiogenic activity of VEGF-A which could be a reason for ovulation failure associated with AHF formation.     

An Overview of Low Dose Insemination in the Mare

The need for relatively high numbers of spermatozoa for artificial insemination limits our application of recently available technologies such as sex‐sorted semen. The fertility of two different methods of low dose insemination using fresh, frozen and sex‐sorted semen are compared in this overview. Satisfactory conception rates are described using very low doses of spermatozoa inseminated by either hysteroscopic or deep uterine insemination methods, proving the stallion is fully fertile. The hysteroscopic method appears to give higher conception rates when inseminating fewer than 5 × 10⁶ spermatozoa and is therefore, the preferred method of insemination for sex‐sorted spermatozoa. However, hysteroscopic deposition of low numbers of spermatozoa from infertile stallions does not appear to improve their fertility.     

The Development and Application of the Modern Reproductive Technologies to Horse Breeding

Although the horse was probably the first animal to experience and benefit from artificial insemination, it trailed the field somewhat with regard to the application of embryo transfer and other oocyte and embryo-related modern breeding technologies. But with a late run it is now back in mid-field and gaining fast on the other large domestic species in the application of the many technological advances of the past 20 years to sound breeding practice. Improvements in extenders and cryoprotectants have resulted in a veritable upsurge in the transport and insemination of cooled and frozen stallion semen, and parallel improvements in ovulation induction and synchrony, exogenous gonadotrophic stimulation of multiple fertile ovulations and simplified, more efficient methods for non-surgical transfer of embryos to recipient mares, coupled with relaxation of breed society registration restrictions, have together contributed to a similar upsurge in the application of embryo transfer to all breeds and athletic types of horses worldwide, with the continuing and notable exception of the Thoroughbred. Although conventional in vitro fertilization remains something of an unjumped fence in equids, other modern breeding technologies like hysteroscopic low-dose insemination, fluorescence-activated sex sorting of stallion spermatozoa, between-species embryo transfer, embryo freezing and bisection, transvaginal ultrasound-guided oocyte collection, intracytoplasmic sperm injection for fertilization (ICSI), gamete intrafallopian transfer (GIFT) and now nuclear transfer (cloning), have all been applied to equids with encouraging success. Cloning, especially, holds enormous promise for the Sporthorse industry to re-create champion geldings in stallion form for breeding purposes.     

Immunohistochemical Localization of Vascular Endothelial Growth Factor (VEGF) and its Two Receptors (Flt-I and KDR) in the Endometrium and Placenta of the Mare During the Oestrous Cycle and Pregnancy

Polyclonal antisera for vascular endothelial growth factor (VEGF) and its two main receptor molecules, VEGF-I (Flt) and VEGF-II (KDR), were used in a conventional immunocytochemical staining method to localize these angiogenic ligand molecules in the endometrium and placenta of the mare during the oestrous cycle and pregnancy. The anti-VEGF and anti-Flt sera both labelled the lumenal and glandular epithelia of the endometrium throughout the oestrous cycle and both the invasive trophoblast cells of the endometrial cups and the non-invasive trophoblast of the allantochorion in pregnancy. The anti-KDR serum likewise stained the maternal and foetal epithelial layers during the oestrous cycle and pregnancy and it also labelled fibroblast-like cells in the endometrial and allantoic stromas and the endothelium of foetal and maternal capillaries. The results demonstrated that constant supplies of the principal vasculogenic and angiogenic factor, VEGF, and its two major receptors, Flt and KDR, are available on both the maternal and foetal sides of the placental barrier throughout gestation in the mare. They are presumed to facilitate the continuing development of the extensive foetal and maternal capillary networks that are such prominent features within the microplacentomes of the diffuse, epitheliochorial equine placenta.     

Immunohistochemical Localization of Oestrogen Receptors α and β, Progesterone Receptor and Aromatase in the Equine Placenta

The functions of placental oestrogens during equine pregnancy are still unclear. Yet, they may act predominantly as local regulators of growth and differentiation in the microplacentomes. Thus, expression patterns of oestrogen receptors (ERs) α and β were investigated in the microcotyledonary placenta from pregnant mares at 110, 121, 179, 199 and 309 days of gestation by immunohistochemistry. In microplacentomes, both the ER isoforms were detected in trophoblast (T) cells, chorionic villous stroma (FS), microcaruncular epithelium (ME) and microcaruncular stroma (MS). Proportions of positive cells were 38–91% (T), 11–41% (FS), 55–89% (ME), 17–51% (MS) for ERα and 66–76% (T), 21–37% (FS), 41–68% (ME) and 24–55% (MS) for ERβ. Between days 110 and 199, proportions of cells positive for progesterone receptor (PR) varied between 19% and 62% (T), 3% and 50% (CS), 15% and 46% (ME), and 4% and 33% (MS). At day 309, PR was virtually absent in T, CS and ME (percentages < 0.1), whereas in MS 14.3% of cells were still positive. The expression of ERs and PR in equine microplacentomes gives evidence for a role of placental steroids as regulators of placental growth, differentiation and function. The detection of ERα, ERβ and PR in foetal and maternal vascular tissue suggests that placental steroids are also involved in the control of placental angiogenesis and /or vascular functions. The co-localization of ERs with aromatase in T suggests auto- or intracrine functions of oestrogens in this cell type.