The domestic dog is a comparative model for studies aimed at the conservation of genetic material of rare canids. Recently, it has been shown that canine oocytes resume meiosis in vitro, although at a much lower rate than other domestic animal oocytes. This could be due either to suboptimal culture conditions or to low developmental competence of oocytes. The physiological role of communications between oocyte and follicular cells is well known to be essential for the acquisition of developmental competence in other domestic species. The purpose of this study was to investigate the possible effect of different stages of the estrous cycle on the presence and persistence of cumulus-oocyte communications through gap-junctions at the time of collection and during IVM as a marker of developmental competence of canine oocytes. Ovaries were harvested from bitches in two different stages of the estrous cycle: anestrous (A) or late proestrous (LP). Ovaries were sliced to release cumulus oocyte complexes (COC). Oocytes with evenly granulated ooplasm and a dense multilayered cumulus were used for culture. The medium used for oocyte maturation was TCM 199 with 0.3% BSA and 0.5 iu FSH, 0.5 iu LH 0.5 ml-1. Oocytes were cultured for 72 h at 39oC in a humidified environment of 5% CO2 in air. After collection (0 h), at 24, 48 or 72 h of IVM, oocytes were fixed and stained with Lacmoid to evaluate the nuclear stage of maturation. At 0, 24, or 48 h of IVM oocytes were microinjected (n= 84) with a 0.3% lucifer yellow solution in order to evaluate the presence of cumulus-oocyte communications through gap-junctions. At time 0, no cumulus-oocyte communications were open in A oocytes as opposed to 89% of t COC collected at LP stage. In LP oocytes communications remained permeable for the first 24 h of maturation (67%) while at 48 h no more fluorescence diffusion was observed. The persistence of cumulus-oocytes communications in LP oocytes was coupled by their ability to resume meiosis and reach the telophaseI/metaphase II stage at a higher rate than oocytes collected at A stage (xx% vs. 0% p<0.05). These results indicate for the first time that during anaestrus oocyte-cumulus cells communications are absent already within the ovary, thus indicating that such oocytes are unsuitable for IVM. Open somatic-oocyte communications, found in LP oocytes, were positively correlated with developmental competence, however the low rate of MII obtained suggests that further efforts to define better culture conditions are still required.

Different stages of the estrous cycle influence cumulus-oocytes communications in canine oocytes: consequences on in vitro maturation efficiency / G.C. Luvoni, A.M. Luciano, S. Modina, L. Marconato, F. Gandolfi - In: Proceedings 4th International Symposium on Canine and Feline Reproduction / [a cura di] W. Farstad, C. Steel. - [s.l] : The Norwegian University Library of Life Sciences, 2000. - ISBN 82-91743-06-1. (( Intervento presentato al 4. convegno International Symposium on Canine and Feline Reproduction tenutosi a Oslo, Norway nel 2000.

Different stages of the estrous cycle influence cumulus-oocytes communications in canine oocytes: consequences on in vitro maturation efficiency

G.C. Luvoni
Primo
;
A.M. Luciano
Secondo
;
S. Modina;F. Gandolfi
Ultimo
2000

Abstract

The domestic dog is a comparative model for studies aimed at the conservation of genetic material of rare canids. Recently, it has been shown that canine oocytes resume meiosis in vitro, although at a much lower rate than other domestic animal oocytes. This could be due either to suboptimal culture conditions or to low developmental competence of oocytes. The physiological role of communications between oocyte and follicular cells is well known to be essential for the acquisition of developmental competence in other domestic species. The purpose of this study was to investigate the possible effect of different stages of the estrous cycle on the presence and persistence of cumulus-oocyte communications through gap-junctions at the time of collection and during IVM as a marker of developmental competence of canine oocytes. Ovaries were harvested from bitches in two different stages of the estrous cycle: anestrous (A) or late proestrous (LP). Ovaries were sliced to release cumulus oocyte complexes (COC). Oocytes with evenly granulated ooplasm and a dense multilayered cumulus were used for culture. The medium used for oocyte maturation was TCM 199 with 0.3% BSA and 0.5 iu FSH, 0.5 iu LH 0.5 ml-1. Oocytes were cultured for 72 h at 39oC in a humidified environment of 5% CO2 in air. After collection (0 h), at 24, 48 or 72 h of IVM, oocytes were fixed and stained with Lacmoid to evaluate the nuclear stage of maturation. At 0, 24, or 48 h of IVM oocytes were microinjected (n= 84) with a 0.3% lucifer yellow solution in order to evaluate the presence of cumulus-oocyte communications through gap-junctions. At time 0, no cumulus-oocyte communications were open in A oocytes as opposed to 89% of t COC collected at LP stage. In LP oocytes communications remained permeable for the first 24 h of maturation (67%) while at 48 h no more fluorescence diffusion was observed. The persistence of cumulus-oocytes communications in LP oocytes was coupled by their ability to resume meiosis and reach the telophaseI/metaphase II stage at a higher rate than oocytes collected at A stage (xx% vs. 0% p<0.05). These results indicate for the first time that during anaestrus oocyte-cumulus cells communications are absent already within the ovary, thus indicating that such oocytes are unsuitable for IVM. Open somatic-oocyte communications, found in LP oocytes, were positively correlated with developmental competence, however the low rate of MII obtained suggests that further efforts to define better culture conditions are still required.
dog; oocytes; cumulus cells
Settore VET/10 - Clinica Ostetrica e Ginecologia Veterinaria
Settore VET/01 - Anatomia degli Animali Domestici
Book Part (author)
File in questo prodotto:
Non ci sono file associati a questo prodotto.
Pubblicazioni consigliate

Caricamento pubblicazioni consigliate

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/185235
Citazioni
  • ???jsp.display-item.citation.pmc??? ND
  • Scopus ND
  • ???jsp.display-item.citation.isi??? ND
social impact