Effect of embryo age on the viability of equine embryos after cooled storage using two transport systems

The aim of this study was to compare the viability of 7- and 8-day-old equine embryos cooled and stored for 6 or 24 hours in two different transport systems. Embryos (n = 97) were recovered on day 7 or 8 and assigned to 10 groups (n = 10/group). Embryos within the same age group (D7 or D8) were evaluated immediately after collection (Group-0h) or after storage in an Equitainer at 5°C for 24 hours in 5 ml Emcare Holding Solution (EHS) (Group-E-24h) or 5 ml Ham's F10 (Group-H-24h) or in a refrigerator at 5°C in 500 ml Emcare Flushing Solution (EFS) for 6 hours (Group-B-6h) or 24 hours (Group-B-24h). After collection or storage, embryos were incubated in 1 μg/ml DAPI to determine the percentage of dead cells per embryo (DAPI positive, fluorescent cells). Subsequently, embryos were fixed in 4% paraformaldehyde and re-stained with DAPI to determine the total number of cells. The percentage of dead cells in group-0h and B-6h was similar and significantly lower than for embryos stored for 24 hours in groups B-24h, E-24h, and H-24h. The percentage of dead cells was similar for embryos stored in an Equitainer (groups E-24h and H-24h) and was significantly higher for embryos stored 24 hours in EFS (Group B-24h). Within each storage system (0h, B-6h, B-24h, E-24h, and H-24h) no significant difference in the percentage of dead cells was observed between 7- and 8-day-old embryos. Storage in 500 ml EFS at 5°C for 6 hours resulted in embryos of better quality than after the traditional 24-hour storage in an Equitainer, suggesting that this simplified system offers a good alternative for short-term storage and transport.     

The Use of Equine Follicle Stimulating Hormone to Increase Equine Chorionic Gonadotropin in the Pregnant Mare

Equine chorionic gonadotropin (eCG), obtained from pregnant mares, is used for assisted reproductive technologies in laboratory rodents and livestock. The objective of the present study was to use equine follicle-stimulating hormone (eFSH) to increase the incidence of twin pregnancies, through multiple ovulations, and increase eCG. Nineteen light horse–type mares were enrolled in the study. The control group (n = 9) was bred with fresh or cooled semen and given human chorionic gonadotropin (hCG) at the time of breeding. The second group (n = 10) was given 12.5 mg of eFSH intramuscularly twice a day beginning 5–7 days after ovulation. Prostaglandin F2α was administered intramuscularly the second day of eFSH treatment. Treatment with eFSH continued until follicles were >35 mm in diameter, and mares were then given no treatment for 36 hours. The mares were then bred with fresh or cooled semen from the same stallion as the control group and given hCG. Blood samples were taken weekly from day 35 to day 105 after ovulation. Serum concentration of eCG was obtained, and data were analyzed with multivariate analysis using the mixed procedure. Significance was set at P < .05. Data were combined for all mares carrying twins and compared with those carrying singletons. The group of mares carrying twins had higher peak concentrations of eCG and higher values for area under the curve compared with mares carrying singletons (P < .05). These results suggest inducing twins could be a method used to increase eCG production.     

Effects of Carbon Dioxide Insufflation Combined With Changes in Body Position on Blood Gas and Acid-Base Status in Anesthetized Llamas (Llama glama)

Objective— To study the combined effects of intra-abdominal CO₂ insufflation with changes in body position during laparoscopy in xylazine-ketamine-halothane anesthetized llamas. Study Design— Prospective, controlled study. Animals— Nine castrated, male llamas weighing 114 ± 23 kg, 3 to 13 years old. Methods— Three llamas (preliminary study [PS] group) were used to study the effect of right lateral, dorsal, and left lateral recumbency on gas exchange and acid-base status. The other six (experimental study [ES] group) were used to study the combined effects of changes in body position and CO₂ insufflation to an intraabdominal pressure of 10 to 12 mm Hg. Heart rate, respiratory rate, and indirect arterial blood pressures (systolic [SAP], mean [MAP], and diastolic [DAP]) were recorded every 5 minutes during anesthesia. Arterial blood gases (PaO₂ and PaCO₂) and acid-base status (pHa and HCO₃) were measured immediately after induction of anesthesia and before each change of position. Results— In the PS group, significant decreases in SAP, MAP and PaCO₂ and increases in PaO₂ and pHa were observed when the llamas were turned from right lateral to dorsal recumbency. Values for HCO_-3 were lower than the postinduction values, but they remained unaffected by the changes in position. In the ES group, values for MAP were significantly lower when the llamas were placed in dorsal and left lateral recumbency than those observed during right lateral recumbency. Arterial O₂ tension during right lateral recumbency was lower but returned to preinsufflation values when the llamas were placed in the dorsal position. All llamas recovered uneventfully within 30 minutes after termination of anesthesia. Conclusions— Insufflation of CO₂ and changing body position induce minor and transient changes in cardiovascular and respiratory function. Clinical Relevance— Laparoscopy with mild intra-abdominal CO₂ insufflation (10 to 12 mm Hg) can be used safely in spontaneously breathing llamas anesthetized with xylazine, ketamine, and halothane.     

Clinical observations on changes in concentrations of hormones in plasma of two stallions with thermally-induced testicular degeneration

To investigate effects of thermally-induced testicular degeneration on hormonal and seminal parameters in stallions, the scrotum was insulated for 36 hours in two mature (5-year-old mixed breed and 11-year-old Throughbred) stallions. Semen was collected daily for 10 days (DSO) prior to, and at intervals after, scrotal insulation. When DSO determinations were not being made, semen was collected 3 times weekly. Jugular blood samples were collected at 15-minute intervals for 6 hours from each stallion prior to, and at intervals after, scrotal insulation. A mouse interstitial cell testosterone assay was modified to quantify biologic activity of equine luteinizing hormone (BLH) in plasma samples. Immunoactive luteinizing hormone (ILH) and testosterone (T) concentrations were determined in plasma samples by routine RIA procedures. Percentages of progressively motile and morphologically normal spermatozoa began to decrease by 1 to 2 weeks postinsulation, reached nadir values at 3 to 3-1/2 weeks postinsulation, and returned to preinsulation values by 7 weeks postinsulation. Total number of spermatozoa and total number of progressively motile, morphologically normal spermatozoa in ejaculates at DSO returned to normal by 8 weeks postinsulation in stallion 2 and 12 weeks postinsulation in stallion 1. Concentrations of BLH and ILH increased, and while T concentrations decreased, immediately postinsulation. The increase in ILH concentrations was greater than the increase in BLH concentrations, resulting in a decrease in the BLH:ILH (B:I) ratio. Following the peak in LH secretion immediately postinsulation, LH concentrations gradually decreased while T concentrations increased. The B:I ratio was elevated from 1 to 13 weeks postinsulation compared to immediately postinsulation. In addition to changes in spermatozoal quality in ejaculates, stallion response to scrotal insulation included increased secretion of luteinizing hormone and impaired Leydig cell function (as determined by reduced testosterone concentration in circulating plasma). The proportion of biologically active LH secreted in response to thermal testicular injury increased during the recovery phase.     

Effect of Prosthesis Number and Position on Rima Glottidis Area in Equine Laryngeal Specimens

Objective— To assess the effect of 1 or 2 laryngeal prosthetic sutures on rima glottidis areas in equine laryngeal specimens. Study Design— Experimental, randomized design. Animals— Cadaveric equine larynges (n=16). Methods— Larynges were collected from 10 horses; 2 sutures each were preplaced in the right and left sides of each larynx. A dorsal suture (DS) was placed through the caudal rim of the dorsal midline of the cricoid cartilage, under the cricopharyngeus muscle and through the proximal and rostral aspect of the muscular process. A lateral suture (LS) was placed 1.5 cm lateral to the DS and through the muscular process more distal and caudal to the 1st suture. Larynges were positioned in a customized stand and the rima glottidis photographed after each suture (LS or DS) or suture combination (CS) was tied in random sequence. An additional 6 larynxes were used to determine whether the tension applied to the sutures was repeatable. Sutures were preplaced in both the right and left side of each larynx as described above and each suture and CS was tied and released 3 times in each larynx. Photographs were taken of the rima glottidis after each suture or CS was tied generating 3 replicates for each suture configuration on each side of the 6 larynges. Results— Mean rima glottidis area was not different between DS and LS when tied alone (P=.85); however, mean area after CS (DS+LS) was greater than DS (P<.001) and LS (P<.001) alone. The coefficient of variation for the 6 suture patterns were low (1–7%) and the intraclass correlation coefficient estimates were very high (0.997–0.998) demonstrating excellent repeatability between replicates for each of the 3 suture configurations. Conclusion— Our results suggest that laryngoplasty using 2 prostheses; 1 placed dorsally in the cricoid and through the rostral and proximal muscular process and 1 placed 1.5 cm lateral to the 1st and more caudal and distal in the muscular process results in a greater cross sectional area of the rima glottidis than either suture used alone. Clinical Relevance— Seemingly prosthetic sutures contribute independently to each other in determining the contour of the rima glottidis. Use of 2 prosthetic sutures improves crosssectional area of the rima glottidis compared with each suture alone and may improve surgical outcome in laryngoplasty.     

Effect of a Nonsurgical Embryo Transfer Procedure and/or Altrenogest Therapy on Endogenous Progesterone Concentration in Mares

Endogenous progesterone levels may decline after transcervical embryo transfer in some mares. Progestogen therapy is commonly used to support endogenous progesterone levels in embryo transfer recipient mares or those carrying their own pregnancy. The goal of this study was to determine the effects of the transcervical transfer procedure and/or altrenogest therapy on luteal function in mares. Mares were assigned to one of six treatment groups: group 1 (untreated control; n = 7 cycles), group 2 (sham transfer, no altrenogest; n = 8 cycles), group 3 (sham transfer plus altrenogest; n = 8 cycles), group 4 (pregnant, no altrenogest; n = 9 mares), group 5 (pregnant plus altrenogest; n = 9 mares), and group 6 (nonpregnant plus altrenogest; n = 10 cycles). Mares in groups 4-6 were bred and allowed an opportunity to carry their own pregnancy. Blood samples were collected for 22 days beginning on the day of ovulation. Sham embryo transfer (groups 2 and 3, combined) did not result in a decline in endogenous progesterone levels compared with control mares (group 6). However, sham embryo transfer did result in luteolysis and an abrupt decline in endogenous progesterone levels in one of the 16 (6.2%) sham-transferred mares. Altrenogest therapy in sham-transferred mares (group 3) was associated with lower endogenous progesterone levels on days 10, 12, and 13 postovulation when compared with sham-transferred mares that did not receive altrenogest (group 2). Administration of altrenogest to pregnant mares (group 5) was associated with lower concentrations of endogenous progesterone from days 14 to 18 and on day 21 compared with endogenous progesterone levels in pregnant mares not administered altrenogest (group 4). In conclusion, a transcervical embryo transfer procedure can cause luteolysis in a low percentage of mares. Altrenogest therapy may be associated with a reduction in endogenous progesterone secretion, presumably mediated by a reduction in pituitary luteinizing hormone (LH) release and a decrease in luteotropic support.