Havemeyer Foundati<strong>on</strong> M<strong>on</strong>ograph Series No. 3TABLE 1: Rates <str<strong>on</strong>g>of</str<strong>on</strong>g> successful embryo collecti<strong>on</strong> knowing <str<strong>on</strong>g>the</str<strong>on</strong>g> exact age <str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>the</str<strong>on</strong>g> embryoFirst collecti<strong>on</strong> attemptSec<strong>on</strong>d and third collecti<strong>on</strong> attemptsMoment <str<strong>on</strong>g>of</str<strong>on</strong>g> 144 h 147 h 150 h 147 h 150 h 150 h 156 h 156 hcollecti<strong>on</strong> (after 144) (after 144) (after 147) (after 150) (after 144(h after ovulati<strong>on</strong>) and 147)Successful 2/16 3/14 5/15 0/2 1/9 0/9 0/80/7collecti<strong>on</strong>s (16%) (21%) (33%) (11%)No. <str<strong>on</strong>g>of</str<strong>on</strong>g> embryos 2 4 a 7 b 1Twin embryos were recovered in 1 a or 2 b cases (double ovulati<strong>on</strong>s were synchr<strong>on</strong>ous)TABLE 2: Morphology <str<strong>on</strong>g>of</str<strong>on</strong>g> embryos recovered 144, 147 or 150 h after ovulati<strong>on</strong>Moment <str<strong>on</strong>g>of</str<strong>on</strong>g> recovery 144 h 147 h 150 h(h after ovulati<strong>on</strong>)No. <str<strong>on</strong>g>of</str<strong>on</strong>g> embryos n = 2 n = 4 n = 8Diameter (µm):mean + sd (range) 186 + 21 163 + 11 166 + 9.8(171-201) (146-168) (153-183)Cell number: 407 + 227( n = 2) 440 (n = 1) 317 + 60 (n = 4)mean + sd (range) (247-568) (274-403)% inner cell mass 37 + 9 29 35 + 7% mitosis 5.6 + 0.7 5.2 5.1 + 3For <str<strong>on</strong>g>the</str<strong>on</strong>g> first collecti<strong>on</strong> attempts, rates <str<strong>on</strong>g>of</str<strong>on</strong>g> succesdid not significantly differ according to <str<strong>on</strong>g>the</str<strong>on</strong>g>moment <str<strong>on</strong>g>of</str<strong>on</strong>g> collecti<strong>on</strong>. Most <str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>the</str<strong>on</strong>g> time, at sec<strong>on</strong>dand third collecti<strong>on</strong> attempts, <str<strong>on</strong>g>the</str<strong>on</strong>g> cervix wasrelaxed, and embryos have probably been lost,ei<str<strong>on</strong>g>the</str<strong>on</strong>g>r during flushing or in <str<strong>on</strong>g>the</str<strong>on</strong>g> time elapsedbetween 2 successive collecti<strong>on</strong>s.In c<strong>on</strong>trast with our previous results (Battut etal. 1997), some embryos were recovered from <str<strong>on</strong>g>the</str<strong>on</strong>g>uterus 144 h (exactly 6 days) after ovulati<strong>on</strong>.Recovery rates at 150 h were lower than thatpreviously reported at 156 h: all <str<strong>on</strong>g>the</str<strong>on</strong>g> embryos d<strong>on</strong>ot seem to have reached <str<strong>on</strong>g>the</str<strong>on</strong>g> uterus 150 h (exactly6.25 days) after ovulati<strong>on</strong>. These data corroborateprevious results (Colchen et al. 2000), indicatingthat <str<strong>on</strong>g>the</str<strong>on</strong>g> moment <str<strong>on</strong>g>of</str<strong>on</strong>g> arrival <str<strong>on</strong>g>of</str<strong>on</strong>g> equine embryos in<str<strong>on</strong>g>the</str<strong>on</strong>g> uterus after ovulati<strong>on</strong> varies am<strong>on</strong>g embryos,within an interval about 12 h. The possible reas<strong>on</strong>sexplaining this variability are: i) variable delaybetween ovulati<strong>on</strong> and fertilisati<strong>on</strong>, depending <strong>on</strong>individual oocytes; ii) maternal factors: noinfluence <str<strong>on</strong>g>of</str<strong>on</strong>g> maternal breed was observed, andprogester<strong>on</strong>emia has no influence (Colchen et al.2000); iii) envir<strong>on</strong>mental factors (o<str<strong>on</strong>g>the</str<strong>on</strong>g>r thanseas<strong>on</strong>); and iv) embry<strong>on</strong>ic factors: sex; moment<str<strong>on</strong>g>of</str<strong>on</strong>g> PGE 2 secreti<strong>on</strong>; o<str<strong>on</strong>g>the</str<strong>on</strong>g>r individual factors.MorphologyUnder inverted microscope, all <str<strong>on</strong>g>the</str<strong>on</strong>g> embryoslooked like morula or early blastocyst, <str<strong>on</strong>g>the</str<strong>on</strong>g>difference between <str<strong>on</strong>g>the</str<strong>on</strong>g> 2 stages being difficult toevaluate. Diameters and cell numbers arepresented <strong>on</strong> Table 2. There was no significantdifference according to <str<strong>on</strong>g>the</str<strong>on</strong>g> moment <str<strong>on</strong>g>of</str<strong>on</strong>g> recovery.Only 8 embryos could be subjected tohistological analysis. One was degenerated, 6 wereearly blastocysts, and <strong>on</strong>e was a morula (150 h,168 µm, 313 cells). Fragments <str<strong>on</strong>g>of</str<strong>on</strong>g> capsule werevisible <strong>on</strong> all <str<strong>on</strong>g>the</str<strong>on</strong>g> embryos, but a c<strong>on</strong>tinuous andvery thin capsule was observed <strong>on</strong>ly around <strong>on</strong>eembryo (150 h, 153 µm, 279 cells). Cell numbersare presented <strong>on</strong> Table 2. There was no correlati<strong>on</strong>between diameter, cell number, percentage <str<strong>on</strong>g>of</str<strong>on</strong>g>inner cell mass, and percentage <str<strong>on</strong>g>of</str<strong>on</strong>g> mitosis.These results indicate that morphology <str<strong>on</strong>g>of</str<strong>on</strong>g>equine embryos, between Day 6 and Day 6.25,does not depend <strong>on</strong> <str<strong>on</strong>g>the</str<strong>on</strong>g>ir age in relati<strong>on</strong> toovulati<strong>on</strong>. Hypo<str<strong>on</strong>g>the</str<strong>on</strong>g>sis <str<strong>on</strong>g>of</str<strong>on</strong>g> explanati<strong>on</strong> are:i) variability in <str<strong>on</strong>g>the</str<strong>on</strong>g> interval ovulati<strong>on</strong>-fertilisati<strong>on</strong>;ii) variability in development rate; iii) variability in<str<strong>on</strong>g>the</str<strong>on</strong>g> interval : arrival in <str<strong>on</strong>g>the</str<strong>on</strong>g> uterus - recovery. In <str<strong>on</strong>g>the</str<strong>on</strong>g>uterus, capsule begins to form, and cell number67
Equine Embryo Transferseems to increase 2-fold every 6 h (Colchen et al.2000). Embryos were recovered at a fixed intervalafter ovulati<strong>on</strong>, but <str<strong>on</strong>g>the</str<strong>on</strong>g> moment <str<strong>on</strong>g>of</str<strong>on</strong>g> transport to <str<strong>on</strong>g>the</str<strong>on</strong>g>uterus is variable with respect to ovulati<strong>on</strong>. Thesereas<strong>on</strong>s explain <str<strong>on</strong>g>the</str<strong>on</strong>g> large possible variati<strong>on</strong> inembry<strong>on</strong>ic morphology (cell number, aspect <str<strong>on</strong>g>of</str<strong>on</strong>g>capsule) according to <str<strong>on</strong>g>the</str<strong>on</strong>g> time spent in <str<strong>on</strong>g>the</str<strong>on</strong>g> uterusbetween arrival and recovery.CONCLUSIONM<strong>on</strong>itoring ovulati<strong>on</strong> hourly does not help torecover an homogenous group <str<strong>on</strong>g>of</str<strong>on</strong>g> embryos, forfreezing or micromanipulati<strong>on</strong>, for example.Factors related to individual oocytes or embryos,and resp<strong>on</strong>sible for this variati<strong>on</strong>, have to beidentified.When ovulati<strong>on</strong>s are checked by ultrasoundexaminati<strong>on</strong> <strong>on</strong>ly <strong>on</strong>ce or twice a day, embryocollecti<strong>on</strong> must be performed 156 h after <str<strong>on</strong>g>the</str<strong>on</strong>g> firstobservati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a corpus luteum, to be sure that <str<strong>on</strong>g>the</str<strong>on</strong>g>embryo has reached <str<strong>on</strong>g>the</str<strong>on</strong>g> uterine cavity.REFERENCESBattut, I., Colchen, S., Fieni, F., Tainturier, D. andBruyas, J.F. (1997) Success rate when attempting t<strong>on</strong><strong>on</strong> surgically collect equine embryos at 144, 156and 168 hours after ovulati<strong>on</strong>. Equine vet. J. Suppl.25, 60-62.Bruyas, J.F., Bézard, J., Lagneaux, D. and Palmer, E.(1993) Quantitative analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> morphologicalmodificati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> day 6.5 horse embryos aftercryopreservati<strong>on</strong>: differential effects <strong>on</strong> inner cellmass and trophoblast cells. J. reprod. Fert. 99,15-23.Colchen, S., Battut,I., Fieni, F., Tainturier, D., Siliart B.and Bruyas J.F. (2000) Quantitative histologicalanalysis <str<strong>on</strong>g>of</str<strong>on</strong>g> equine embryos at exactly 156 and 168 hafter ovulati<strong>on</strong>. J. reprod. Fert.Suppl. 52, In press.68
Change language