Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias
Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias
Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias
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16 t h International Congress on Animal <strong>Reproduction</strong><br />
Poster Abstracts 127<br />
P300<br />
Acros<strong>in</strong> activity evaluation <strong>in</strong> chilled/rewarmed dog<br />
sperm dur<strong>in</strong>g different capacitation times<br />
De Los Reyes, M*, Godoy, N; Palom<strong>in</strong>o, J<br />
Animal Production, Animal <strong>Reproduction</strong> Unit, Faculty of Veter<strong>in</strong>ary Sciences,<br />
University of Chile, Chile<br />
Introduction The acrosome reaction causes release of acros<strong>in</strong>, a<br />
proteolytic enzyme that facilitates sperm penetration through the zona<br />
pellucida. The release of acros<strong>in</strong> on <strong>in</strong> vitro capacitated<br />
chilled/rewarmed can<strong>in</strong>e spermatozoa is a time <strong>de</strong>pend<strong>in</strong>g process.<br />
Thus, the aim of the present work was to study the proteolytic acros<strong>in</strong><br />
activity <strong>in</strong> chilled/rewarmed can<strong>in</strong>e spermatozoa dur<strong>in</strong>g <strong>in</strong> vitro<br />
capacitation and acrosome reaction.<br />
Methods Six ejaculates were collected from four adult dogs. Each<br />
ejaculate was liquated <strong>in</strong>to 2 fractions and centrifuged <strong>in</strong> tris buffer<br />
medium. The pellet of one fraction was diluted <strong>in</strong> fert-talp medium<br />
(fresh control sample), and the pellet of the other fraction was diluted<br />
<strong>in</strong> a tris-fructose-citric acid and egg-yolk exten<strong>de</strong>r and then cooled at<br />
4º C for 24 h. Sperm samples (fresh and chilled/rewarmed) were<br />
centrifuged and the pellet rediluted <strong>in</strong> fert-talp medium and then<br />
aliquoted <strong>in</strong>to four tubes which were <strong>in</strong>cubated separately for 0, 1, 2<br />
and 3 h at 20º. At each time of cultured, acros<strong>in</strong> activity was measured<br />
by the gelat<strong>in</strong>-substrate film technique, where enzyme activity is<br />
<strong>de</strong>tected by the presence of halos around s<strong>in</strong>gle sperm result<strong>in</strong>g from a<br />
localized proteolytic digestion of gelat<strong>in</strong>. Gelat<strong>in</strong> suspension was<br />
placed on pre-cooled sli<strong>de</strong>s and fixed <strong>in</strong> 0.05% glutaral<strong>de</strong>y<strong>de</strong>, washed<br />
and kept overnight. Sperm suspensions were placed on the slices and<br />
<strong>in</strong>cubated at 38°C <strong>in</strong> CO2 for 24 h. Slices were sta<strong>in</strong>ed with Comassie<br />
Blue and exam<strong>in</strong>ed with light microscopy for evi<strong>de</strong>nce of digestion.<br />
Results Fresh and chilled dog spermatozoa <strong>in</strong>cubated for up to 3 h <strong>in</strong><br />
fert-talp medium, displayed digestion halos on gelat<strong>in</strong> films. Those<br />
digestion halos were of three different mean sizes: small, medium and<br />
large. Acros<strong>in</strong> activity as measured by halo diameter on sli<strong>de</strong>s coated<br />
with gelat<strong>in</strong>, showed a significant difference between fresh and<br />
chilled sperm. A low proportion of large halos was observed <strong>in</strong> fresh<br />
samples at the beg<strong>in</strong>n<strong>in</strong>g of culture (time 0 and 1), while chilled<br />
sperm showed a high rate of large halos at these times. Dur<strong>in</strong>g the<br />
follow<strong>in</strong>g hours (2 and 3) of culture, fresh sperm showed higher rates<br />
of large halos than fresh sperm.<br />
Conclusion These results <strong>in</strong>dicate that acrosomal proteolytic activity<br />
of chilled/ warmed dog sperm is different throughout the time from<br />
that of the control fresh sperm Supported by Grant FONDECYT<br />
1060602.<br />
P301<br />
Term<strong>in</strong>at<strong>in</strong>g pregnancy <strong>in</strong> bitches: compartive study of<br />
si<strong>de</strong> effects between four treatments<br />
Fila Varela, D 1 *, Berglavaz, A 2 , <strong>de</strong> Leon, J 3 , Navarro, G 3 , Morga<strong>de</strong>s, D 2 ,<br />
Pereira, O 3 , Elhordoy, D 1 , Cavestany, D 1<br />
1Animal <strong>Reproduction</strong> Department, Veter<strong>in</strong>ary Faculty, Uruguay; 2 Private<br />
Practitioner, Don Quijote Veter<strong>in</strong>ary, Uruguay; 3 Private Practitioner, Zoolymar<br />
Vet, Uruguay<br />
To compare <strong>in</strong>ci<strong>de</strong>nce of si<strong>de</strong> effects <strong>in</strong> four medical treatments to<br />
term<strong>in</strong>ate unwanted pregnancies. 166 bitches were assigned to<br />
different treatments: Group A: (n=38) aglepristone (Aliz<strong>in</strong>e, Virbac,<br />
France) (10 mg/kg), two s.c. <strong>in</strong>jections, 24 hours apart, around 25 to<br />
30 days of gestation; Group D: (n=44) <strong>de</strong>xamethasone orally (0,2<br />
mg/kg two times per day, by 5 days and then 0,1 mg/kg two times per<br />
day, dur<strong>in</strong>g 5 more days) around 28 to 30 days of gestation; Group E:<br />
(n=45) estradiol (0,01mg/kg) <strong>in</strong>tramuscularly with<strong>in</strong> 48 hours of<br />
mated; and Group L: (n=39) lotrifen (Privaprol, FATRO, Italy) (2,5<br />
mg/kg) <strong>in</strong>tramuscularly with<strong>in</strong> 15 days of mated. In group A, there<br />
were no si<strong>de</strong> effects. In group D, the si<strong>de</strong> effects were mild polydipsia<br />
and polyuria that disappeared when treatment was discont<strong>in</strong>ued.<br />
Moreover, 7 of 44 bitches (15,91%) presented bloody vag<strong>in</strong>al<br />
discharge dur<strong>in</strong>g 7 days after treatment en<strong>de</strong>d, without general<br />
symptoms. In group E, 12 of 45 bitches (26,67%) <strong>de</strong>veloped cystic<br />
endometrial hyperplasia-pyometra that forced ovary-hysterectomy. In<br />
group L, 18 of 39 bitches (46,15%) <strong>de</strong>veloped cystic endometrial<br />
hyperplasia-pyometra that forced ovary-hysterectomy. Data were<br />
analyzed us<strong>in</strong>g X2 test and differences with<strong>in</strong> treatments were<br />
consi<strong>de</strong>red significant when p 0.05). They occurred <strong>in</strong> 6/6, 3/3,<br />
5/6 and 5/6, 2/3, 4/5 of the animals of the DA, DA&ACY and<br />
DA&ACY+2 groups, respectively. Estrous response appeared 5.0 ±<br />
1.2, 10 ± 1.0 and 24.2 ± 11.7 days after treatment <strong>in</strong> the same groups<br />
(P = 0.1).<br />
Conclusions The GnRH antagonist, acyl<strong>in</strong>e, adm<strong>in</strong>istered at two<br />
different time po<strong>in</strong>ts after a GnRH agonist failed to prevent estrous<br />
response <strong>in</strong> most of these anestrous bitches. Although, <strong>in</strong> the<br />
antagonist treated groups, estrous response had a ten<strong>de</strong>ncy to appear<br />
later. These f<strong>in</strong>d<strong>in</strong>gs suggest a postponement of the stimulation period<br />
dur<strong>in</strong>g the peak antagonistic effect. Further studies with repeated or<br />
higher doses of antagonists are necessary.<br />
Acknowledgements This study was fun<strong>de</strong>d by the National Agency<br />
for Scientific and Technological Promotion, Argent<strong>in</strong>a (Grant PICT<br />
38376/05). The authors thank to NICHHD, NIH, USA and Peptech,<br />
Australia for drug provision.<br />
References Wright PJ, et al. 2001. The Suppression by Progest<strong>in</strong> of<br />
Oestrus Responses of the Bitch to the GnRH Analogue Deslorel<strong>in</strong>. J<br />
Reprod Fertil. 57: 283-8.<br />
P303<br />
Distribution of active mitochondria <strong>in</strong> can<strong>in</strong>e oocytes is<br />
related to reproductive cycle stage but can be damaged<br />
dur<strong>in</strong>g IVM culture<br />
Iorga, AI*; Valent<strong>in</strong>i, L; De Santis, T; Ambruosi, B; Guaricci, AC; Caira, M;<br />
Dell’Aquila, ME<br />
Department of Animal Production, University of Bari, Italy<br />
Introduction We <strong>in</strong>vestigated the effect of the reproductive cycle<br />
stage on the distribution of active mitochondria <strong>in</strong> can<strong>in</strong>e oocytes<br />
exam<strong>in</strong>ed 1) at collection and 2) after <strong>in</strong> vitro maturation (IVM).<br />
Methods Cumulus-oocyte complexes (ooplasmic size >120 µm <strong>in</strong><br />
diameter) were recovered from 20 bitches divi<strong>de</strong>d <strong>in</strong>to five groups