Effects of ultrasound-guided transvaginal follicular aspiration on oocyte recovery and hormonal profiles before and after GnRH treatment.

Endocrine changes and recovered oocytes were evaluated during 16 wk of ultrasound-guided transvaginal follicular aspiration (TVFA) and prior to and following administration of GnRH at the cessation of aspiration. Nonlactating previously aspirated (PAC, n = 4) and non-aspirated, (AC, n = 4) Holstein cows were subjected to 16 wk of twice-weekly aspiration. Four control cows (OAC) were aspirated 1 time only at the final TVFA session (wk 16). Jugular blood samples were collected from all cows during aspiration, before and after the final TVFA session, and during an 18-d period following cessation of aspiration. Ovarian activity was monitored in all cows after cessation of aspiration for 18 d. The PAC and AC cows averaged 3.4 +/- 1.2 (+/- SE) and 6.8 +/- 1.2 oocytes per session, respectively. Progesterone concentrations during TVFA did not differ between the PAC and AC (0.8 +/- 0.1 and 0.9 +/- 0.1 ng/mL, respectively). Progesterone concentration in OAC was 4.5 +/- 0.2 ng/mL before TVFA, while the PAC and AC averaged 0.5 +/- 0.2 and 0.3 +/- 0.2 ng/mL, respectively, at 16 wk. At Week 16 LH was 1.0 +/- 0.2 ng/mL and it increased to 7.5 +/- 0.1 ng/mL after GnRH treatment. The LH concentration before the final aspiration session was higher at peak amplitude in PAC than in AC groups and peak length was longer in OAC than in AC cows (P < 0.07). Between 18 and 24 h after the last aspiration there were more LH peaks and greater peak frequencies in PAC than in OAC cows (P < 0.07), and the interval between peaks was longer in PAC and AC cows (P < 0.10) than in OAC cows. Mean FSH concentrations were lower (P < 0.01) for OAC than for PAC and AC groups at 20 and 24 h after the last aspiration. Follicle numbers after GnRH varied most among treatment groups for follicles < 9 mm, with the PAC, AC and OAC averaging 5.1 +/- 1.0, 5.1 +/- 1.0, and 3.8 +/- 1.0 follicles/d, respectively. Progesterone concentrations increased to 1.1 +/- 0.3 ng/mL in PAC cows and 2.5 +/- 0.3 and 3.4 +/- 0.3 ng/mL in AC and OAC groups, respectively, during the 18-d period. These results suggest that long-term TVFA affects progesterone, LH and FSH profiles and ovarian dynamics in cows.

In vitro embryo production after microinjection and ovarian dynamics following transvaginal follicular oocyte aspiration.

Ultrasound-guided transvaginal follicular aspiration of oocytes from live cows combined with IVM, IVF and in vitro culture (IVC) is a procedure for producing preimplantation-stage bovine embryos and a source of oocytes for pronuclear microinjection of DNA for producing transgenic cattle. This experiment was designed to compare in vitro embryo development rates between oocytes derived from transvaginal follicular aspiration and those obtained from cows at slaughter. Nine cows were subject to a twice-weekly aspiration. Oocytes were aspirated with a 5 MHz ultrasound transducer packaged in a vaginal probe equipped with a dorsal-mounted needle guide (16-ga). All visible follicles (>2 mm) were punctured with a 17-ga, 55-cm needle at each aspiration session and the contents removed under vacuum suction. Oocytes underwent IVM/IVF/IVC. Microinjection of DNA was performed during the pronuclear stage of development, and the zygotes were co-cultured on Buffalo Rat Liver (BRL) cells in modified M199 at 39 degrees C in 5% CO2 and air. After 7 d in culture, embryos were removed and scored for development. A Chi-square analysis was used to compare transvaginal follicular-derived oocytes (microinjected and not) and slaughterhouse-derived, matured in transit oocytes (SHDMT; microinjected and not). Nonmicroinjected embryos resulting from IVF of transvaginal aspiration-derived oocytes developed to blastocysts at a higher rate than SHDMT oocytes (40.0 vs 30.8%; P < 0.05). There was no difference in development rates between the microinjected groups (aspiration = 15.9% vs SHDMT = 12.8%). Higher proportions of the embryos generated from the aspirated oocytes were of excellent or good quality following culture (P < 0.05). In the present experiments the effects of microinjection may overshadow some effects of ova source, but transvaginal follicular aspiration may provide a more consistent, synchronous population of oocytes than those derived from commercial slaughter house sources for use with in vitro systems.