dog sperm, which may also be usable for the assessment of sperm function in vitro for Ž
other canines Hay et al., 1997; Hewitt and England, 1998b,c; Mayenco-Aguirre and .
Peres-Cortez, 1998, Strom Holst, 1999 . Red wolf sperm bind to domestic dog oocytes
¨
Ž .
Goodrowe, 1999, personal communication . 2.3. Artificial insemination and ET
IU artificial insemination may be carried out non-surgically using either endoscopic Ž
visualization or transcervical catheterization, or surgically by laparoscopy for review, .
see Farstad, 2000 . Results in foxes and dogs using IU non-surgical AI are good, and Ž
both birth rates and litter sizes approach those of natural mating see reviews Farstad, .
1996, 1998 . The ET of in vivo-derived embryos has been carried out surgically both in Ž
the silver fox and the dog resulting in live young, but with low success rates Kinney et .
al., 1979; Tsutsui et al., 1989; Jalkanen and Lindeberg, 1998 . Recently, ET has been carried out in the blue fox, using the IU catheter developed for artificial insemination in
Ž .
foxes Lindeberg, 1999, personal communication . To date, the birth of live young from cryopreserved canine embryos has not been reported. However, in the blue fox, freezing
of embryos recently resulted in the observation of implantation sites in naturally Ž
. synchronized recipient females after ET Lindeberg, 1999, personal communication .
The refinement of freezing regimens, improvement of donor-recipient synchronization, in vitro handling of embryos and transfer techniques may soon render both cryobanking
of embryos and ET feasible in foxes.
3. Felines
3.1. Oocyte maturation Generally, cats are seasonally polyestrous carnivores with sexual activity during the
months of increasing day length, and sexual inactivity during the months of declining Ž
. day lengths. Contrary to canines and most other domestic animals, cats are usually
reflex ovulators, i.e. oocytes are ovulated 24–48 h after the post-coital LH release. The Ž
. oocytes are ovulated as secondary oocytes in metaphase II. Goodrowe et al. 1988 first
demonstrated that unovulated follicular oocytes after IVF were able to sustain develop- ment to term with the birth of live kittens. IVM rates are relatively high in cat oocytes
Ž .
40–60 depending on the quality of the oocyte and the type of hormonal supplemen- tation. The stage of the estrous cycle and supplementation of maturation media with
Ž gonadotrophins Goodrowe et al., 1991; Schramm and Bavister, 1995; Wood et al.,
. Ž
1995; Pope et al., 1997 and quality of the cumulus oocyte complex Wood and Wildt, .
1997 influence in vitro development. The highest incidence of MII can be expected after 40–48 h of IVM, similar to the time period from mating to ovulation in the queen
Ž .
Goodrowe et al., 1989 . However, other studies have found that most oocytes reach MII within the first 24 h of IVM, and insemination at 40 h or later does not result in
Ž .
development to blastocysts Luvoni and Oliva, 1993; Wolfe and Wildt, 1996 . The average rate of oocytes that complete maturation to MII in vitro is consistently higher in
Ž felines than in canines, and maturation occurs more rapidly 40–60 MII, 24 h IVM vs.
. 0–58 AIrMIrMII, 48–72 h IVM, respectively , but IVM rates are still lower than in
Ž .
most farm animal IVM systems 80, 24 h IVM . IVM has also been successful in Ž
. some non-domestic felids Johnston et al., 1991 .
3.2. Sperm treatment and assisted fertilization The quality of ejaculated sperm differs within felid species. Some breeds with low
genetic variability have a high incidence of teratospermia and high number of sperm Ž
. Ž .
with acrosomal defects, such as the cheetah Acinonyx jubatus Wildt et al., 1992 . The
structural mechanisms relating to the acrosome and functional defects in protein phosphorylation of some wild feline sperm may be the cause of decreased sperm
Ž .
function Goodrowe, 1999, personal communication . Homologous or heterologous zona binding systems and oocyte penetration assays have been developed for feline sperm
Ž .
Goodrowe and Hay, 1993; Swanson et al., 1998; Nelson et al., 1999 . Fertilization rates after in vitro fertilization of domestic cat oocytes varies between 40 and 50 of in
Ž vitro matured and 60–80 of in vivo matured oocytes Pope, 1999, personal communi-
. cation . In vitro fertilization has been successful in the domestic cat in terms of
Ž production of both embryos and live offspring Goodrowe et al., 1989; Hoffert et al.,
. Ž
1997 . In a few non-domestic felids, such as the tiger Panthera tigris and Indian
Ž .
Ž Desert cat F. silÕestris , offspring have been obtained Pope et al., 1989; Donoghue et
. al., 1990; for review, see Howard, 1999 .
Lately, blastocysts have been obtained from in vivo matured oocytes collected from gonadotrophin stimulated domestic queens, fertilized in vitro by any of the following:
Ž . Ž .
Ž .
Ž . Ž
1 co-incubation with sperm, 2 subzonal insemination SUZI or 3 ICSI Pope et al., .
Ž 1998 . Earlier attempts at comparing SUZI and ICSI were in favor of SUZI Pope et al.,
. 1995 , but improvements in sperm injection technique and visualization of ooplasm by
centrifugation of oocytes improved results considerably in favor of ICSI. ICSI is the most invasive of the assisted fertilization techniques: it allows fertilization with immo-
Ž .
bile teratogeneic sperm, and also enables fusion between nuclei at different stages of development. Thus, refining methods for injection of sperm or other DNA containing
material into the oocyte may spur the development of nuclear transfer techniques in felids, and thereby provide the possibility for genetic modification, as well as for
conservation of nuclear material. Cat ooplasm could be a potential host for somatic cell nuclei from endangered species of felids, as suggested by the bovine model, which
demonstrated that bovine oocyte cytoplasm supports embryo development of nuclear
Ž .
transfer produced embryos from many species Dominko et al., 1999 . 3.3. Embryo culture in Õitro
In vivo matured oocytes readily develop to the blastocyst stage after in vitro fertilization and culture. The developmental rate of in vivo matured in vivo fertilized
Ž .
embryos to the morula stage in TCM 50–90 , and the rate of blastocyst formation Ž
. was 50–66
mean rate 64.7 depending on the developmental stage of the
zygoterearly cleavage stage embryo at flushing of the gonadotrophin stimulated donors
Ž .
Kanda et al., 1995 . Blastocysts have been produced by IVC of in vitro matured and Ž
IVF cat oocytes, and the rate of blastocyst formation varied between 10 and 50 Wolfe and Wildt, 1996; Wood and Wildt, 1997; Pope et al., 1997; Freistedt et al., 1999;
. Swanson et al., 1999 . Live kittens have been born after transfer of embryos produced
Ž .
by complete in vitro generation, i.e. IVM, IVF and IVC Pope et al., 1997 . Until recently, a high percentage of felid embryos produced in vitro experienced a
developmental block at the morula to blastocyst stage, i.e. somewhat later than the in Ž
vitro block observed in other mammals Donoghue et al., 1990; Johnston et al., 1991; .
Roth et al., 1994; Swanson et al., 1996b; Hoffert et al., 1997 . A difference in Ž
. metabolism glycolysis was demonstrated between in vivo and IVM cat oocytes that
may partially explain why IVM oocytes may have developmental difficulties after IVF Ž
. was compared with in vivo matured oocytes Spindler and Wildt, 1999 . A recent study
showed that both supplementation of cysteine to the maturation medium and reduction of the O atmosphere significantly improved in vitro development to the blastocyst stage
2
Ž .
Pope et al., 1999 . Thus, as in bovine IVF, in vitro matured cat oocytes do not develop to blastocysts as readily as their in vivo matured counterparts, but the difference is no
Ž .
longer strikingly large due to improvements in culture conditions Swanson et al., 1998 . Results in the vicinity of 30–40 blastocysts from IVM oocytes vs. 40–50 from in
Ž .
vivo matured oocytes can be expected on day 7 Pope, 1999, personal communication . 3.4. Short- and long-term preserÕation of gametes, follicles and embryos
Reports on the of use of frozen semen for the exchange of feline genetic material has Ž
. been limited Howard et al., 1997b . Cat semen may be chilled to 4
8C and stored for Ž
24–48 h in a TesT buffer based on a trishydroxymethyl amino methane sulphonic acid .
Ž buffer, Tes at pH 7.4, and subsequently used for AI or in vitro insemination Axner,
. Ž
. 1998 . Buffers, such as TesT and Tris trishydroxy methylamino methane , have been
Ž .
used with 4 glycerol or dimethylsulphoxide DMSO and 20 egg yolk, yielding no Ž
. differences between the tested extenders. High cryoprotectant concentrations i.e., 8
Ž .
compromised cat sperm Nelson et al., 1999 . Pelleted freezing has often been the Ž
. standard method Howard, 1986 . Freezing in straws has been found to be equal to
Ž .
freezing in pellets Wood et al., 1993 . A pregnancy rate of only 10 was obtained in Ž
cats after the use of frozen–thawed semen with vaginal deposition of semen Platz et al., .
Ž .
1978 , and vaginal inseminations in wild felids have been unsuccessful Howard, 1999 . Offspring from IU laparoscopic AI with frozen–thawed semen have been obtained in
Ž .
Ž .
Ž .
ocelot F. pardalis , leopard cat F. bengalensis , cheetah Aci. jubatus , snow leopard Ž
. Ž
. Ž
P. uncia , clouded leopard Neofelis nebulosa and tiger for review see, Howard,
. 1999 .
Cat oocytes collected from ovaries and exposed to up to 72 h in refrigerated storage Ž
. matured to MII at normal rates 50–60 . Oocytes collected from ovaries that had been
stored for 24 h developed to blastocysts, showing that cold storage of feline oocytes Ž
does not compromise their ability to sustain development in vitro Wolfe and Wildt, .
1996 , and cold storage up to 48 h results in moderate gross degeneration of oocytes Ž
. Ž
. Wood et al., 1997 . Immature oocytes germinal vesicle stage have been successfully
cryopreserved and have resumed and completed meiosis after slow freezing with
DMSO, although the meiotic resumption rate was half the rate of unfrozen controls Ž
. Luvoni et al., 1997 . Isolated preantral ovarian cat follicles also have been cryopre-
served, showing that a small subpopulation of these follicles survive and are functionally Ž
. intact after freezing with conventional cryoprotectants Jewgenow et al., 1998 . The first
report on the birth of live kittens after ET of cryopreserved, in vivo-derived embryos Ž
. was reported in 1988 Dresser et al., 1988 and later, from cryopreserved embryos
Ž .
produced in vitro from in vivo matured oocytes Pope et al., 1994 and after in IVM, Ž
. IVF and IVC-derived cryopreserved cat embryos Pope et al., 1997 . The cryopreserva-
tion of domestic cat embryos can be carried out in a variety of cryoprotectants. Modifications of freezing regimens for bovine and mouse embryos have provided
Ž .
promising results Pope et al., 1994, 1997; Swanson et al., 1999 . 3.5. Artificial breeding techniques
Artificial insemination in domestic cats has mostly been used in research, in which Ž
cats have served as model species. Laparoscopic insemination is often used Howard et al., 1992, 1997; Donoghue et al., 1993, 1996; Barone et al., 1994; Swanson et al.,
. 1996a . Deep vaginal insemination may be done in domestic cats by inserting a French
Ž .
Tom cat catheter as far as possible into the vagina see Axner, 1998 . A method for
´
non-surgical IU ET has been described, which may equally well be used for intrauterine Ž
. Ž
. AI Swanson and Godke, 1994 . Pregnancy rates are higher after IU laparoscopic AI
Ž .
both in domestic and captive wild cats see Axner, 1998; Howard, 1999 . The time of
´
insemination in relation to gonadotrophin stimulation and anaesthesia influences preg- nancy rates. The best time for AI when anaesthesia is used, is after ovulation has
Ž .
occurred Howard, 1999 . ET has been carried out in both domestic and wild cats with both fresh and
Ž cryopreserved in vivo and in vitro-derived embryos Dresser et al., 1988; Pope et al.,
. 1989, 1994, 1997; Donoghue et al., 1990; Kanda et al., 1995; Swanson et al., 1998 .
Surgical, laparoscopic and transcervical ETs have resulted in live offspring, albeit at a Ž
. relatively low rate in terms of survival of transferred embryos Swanson et al., 1998 .
Transfer to either the oviduct or the uterus has resulted in pregnancies. Lately, transferring 5–8 morulae or blastocysts per recipient improved overall pregnancy rate as
Ž .
well as pregnancy rate per recipient Swanson et al., 1999 .
4. Conclusion
