Current topics in artificial insemination of sheep

There have been developments in several aspects of artificial insemination (AI) in recent years, some of which have been directly responsible for proliferation of AI in the sheep-breeding industries of several countries. The most notable advances have probably been associated with the development of intrauterine insemination by laparoscopy. There is potential for refinement of some of the related techniques, particularly in the area of control of ovulation and definition of appropriate times and optimum doses of spermatozoa for insemination. It is unlikely that laparoscopic AI will be developed sufficiently that it will become readily affordable, and therefore widely practised, by commercial producers. Unfortunately, there has been little progress in the past few years in improvement of the methods of cryopreservation of ram semen. There is considerable potential for AI to have a significant impact on the genetic improvement of sheep, though this has yet to be evaluated in practice. However, if the full potential of AI in sheep is to be realized, it will likely only happen when methods of freezing semen are improved sufficiently that cervical or even vaginal insemination can be widely used with frozen-thawed semen, or when practicable methods of deep cervical or intrauterine insemination through the cervix are developed.     

Transcervical artificial insemination of Australian Merino ewes with frozen-thawed semen

We compared conventional methods for laparoscopic and cervical artificial insemination (AI) to a transcervical AI procedure (Guelph System for Transcervical AI; GST-AI) for use with frozen semen in Merino ewes. The GST-AI procedure was performed by an experienced operator in Experiment 1 (771 ewes) and by 2 inexperienced operators in Experiment 2 (555 ewes). In Experiment 1, intrauterine insemination by GST-AI was achieved in 76% of the ewes. The pregnancy rate at Day 70 for ewes inseminated by laparoscopy (48%, 120 251 ) was higher (P<0.01) than for ewes inseminated by either intrauterine GST-AI (32%, 64 201 ) or cervical AI (9%, 24 256 ). The overall (intrauterine and intracervical) pregnancy rate for GST-AI was 26% (68 264 ) and was unaffected by depth of insemination within the cervix. Pregnancy rates were unaffected by ram or day of insemination. In Experiment 2, the operators achieved intrauterine inseminations by GST-AI in 43% (78 182 ) of the ewes, with a significant operator effect (P<0.01) on depth of cervical penetration. The pregnancy rate to intrauterine GST-AI (40%, 31 78 ) did not differ from that to laparoscopic insemination. The total pregnancy rate for GST-AI in Experiment 2 (19%, 34 182 ) was lower (P<0.05) than that for laparoscopic AI (39%, 72 187 ) but superior (P<0.05) to that for cervical AI (1%, 1 186 ). The GST-AI pregnancy rates were affected by depth of AI (P<0.01) and by operator (P<0.05). It is concluded that GST-AI is superior to cervical AI, and may have application in Merinos if cervical penetration rates can be improved.     

Effect of clitoral stimulation after artificial insemination on conception in Zebu-crossbred heifers in the tropics

In order to increase conception rates to AI, an experiment was conducted to determine the effect of clitoral stimulation on pregnancy rate. Seven hundred and thrity-five Zebu-cross heifers were assigned to either of the following treatments: 1) control without clitoral stimulation (n=351) or 2) clitoral stimulation after IA (n=384). The data were analyzed using a linear model procedure. Clitoral stimulation increased pregnancy rates (57 vs 45 % for treated and control groups, respectively; P<0.02). Holstein x Zebu and Brown Swiss x Zebu heifers attained similar rates (P>0.05). There was a significant treatment-by-season interaction (P<0.01), with pregnancy rates being higher using clitoral massage during the rainy season (54.7 and 25.5% for the treated and control group respectively). In this study, manual clitoral stimulation increased conception rates in Zebu cross heifers under tropical conditions.     

Induction of follicular wave emergence for estrus synchronization and artificial insemination in heifers

The objective was to synchronize follicular wave emergence among cattle for synchronization of estrus and ovulation, and to determine pregnancy rate after AI at observed estrus. At random stages of the estrous cycle, a controlled internal drug release device (CIDR-B) was inserted intravaginally (Day 0) in 67 cross-bred beef heifers, and they were randomly allocated to receive either no further treatment (Control; n = 18); 5 mg of estradiol-17beta and 100 mg of progesterone im (E/P; n = 16); 100 microg im of GnRH (GnRH; n = 16); or transvaginal ultrasound-guided follicular ablation of all follicles > or = 5 mm (FA; n = 17). All heifers received a luteolytic dose of PGF (repeated 12 h later), and CIDR-B were removed on Days 9, 8, 6 or 5, in Control, E/P, GnRH or FA groups, respectively, so the dominant follicle of the induced wave was exposed to exogenous progesterone for a similar period of time in each group. Mean (+/- SEM) intervals (and range, in days) from treatment to follicular wave emergence in these groups were 3.5 +/- 0.6 (-2 to 8), 3.4 +/- 0.1 (3 to 4), 1.5 +/- 0.3 (-1 to 4), and 1.0 +/- 0.1 (0 to 2), respectively. Although the interval was longest (P<0.01) in the E/P and Control groups, it was least variable (P<0.01) in the E/P and FA groups. Intervals (and range, in days) from CIDR-B removal (and first PGF treatment) to estrus were 2.3 +/- 0.2 (1.5 to 4.5), 2.2 +/- 0.2 (1.5 to 3.0), 2.1 +/- 0.1,(1.5 to 3.5), and 2.5 +/- 0.1 (2.0 to 3.5), and to ovulation were 3.5 +/- 0.2 (2.5 to 5.5), 3.4 +/- 0.1 (3.0 to 4.5), 3.5 +/- 0.1 (2.5 to 4.5), and 3.8 +/- 0.1 (3.0 to 4.5), for Control, E/P, GnRH and FA groups, respectively (ns). The proportion of heifers displaying estrus was higher in the Control than in the FA group (94% versus 65%, P<0.05) and intermediate in EP and GnRH groups (87% and 75%). Heifers were inseminated approximately 12 h prior to ovulation (based on estrous behavior and ultrasound examinations). Pregnancy rates were 78%, 80%, 69% and 65% for Control, E/P, GnRH and FA groups, respectively (P=0.73). Results support the hypothesis that synchronous follicular wave emergence results in synchronous follicle development and, following progesterone removal, synchronous estrus and ovulation with high pregnancy rates to AI. The synchrony of estrus and ovulation in the E/P, GnRH and FA groups suggest that these treatments, in combination with CIDR-B, could be adapted to fixed-time insemination programs.     

Evaluation of an ovarian synchronization scheme for fixed-time artificial insemination in wapiti

The ovarian response to an empirically derived treatment protocol used commercially for fixed-time insemination in wapiti (Cervus elaphus) was evaluated by transrectal ultrasonography in hinds during transition into the ovulatory season. On September 29, hinds (n=7) were given an intravaginal progesterone-releasing device (CIDR-B, 1.9 g of progesterone) or left untreated (controls, n=9). Fourteen days later, hinds in the treated group were given 200 IU eCG and the CIDR was removed. Hinds in the control group ovulated randomly over a 15 day period. In the treated group, five hinds ovulated 3 days after eCG treatment, one ovulated 7 days after treatment, and one failed to ovulate by November 1. All extant dominant follicles ceased growth and/or began to regress within 2 days of CIDR placement. Two waves of follicular development were detected between CIDR insertion and removal; the first emerged 5.1+/-0.5 days after CIDR insertion and the second at 11.0+/-0.7 days. Serum progesterone concentration was 0.6+/-0.5 ng/mL (range 1.0-0.3 ng/mL) before CIDR placement, remained above 6 ng/mL during CIDR placement, and fell to 0.8+/-0.9 ng/mL after CIDR removal. In the control group, maximal luteal-phase progesterone concentration was lower (1.1+/-0.1 ng/mL; P<0.05) and emergence of the first follicular wave was more variable (P=0.05) than in the treated group. The protocol to synchronize ovulation was effective in 5/7 (71%) hinds, and 4/7 (57%) became pregnant and calved. The pregnancy rate (6/9) and calving rate (5/9) was similar in the control group. In conclusion, synchronization with CIDR-B was effective; however, the protocol may be improved by shortening the interval of CIDR placement to < or = 7 days and by reducing the circulating concentrations of progesterone to physiologic concentrations (< 4 ng/mL).     

Transcervical artificial insemination in sheep: effects of a new transcervical artificial insemination instrument and traversing the cervix on pregnancy and lambing rates

Cervical anatomy limits the use of transcervical intrauterine artificial insemination (TC AI) in sheep. We have developed an instrument to cope atraumatically with the cervix; although this instrument has not affected fertilization rate or pregnancy rate through Day 3, the effects on sperm transport and pregnancy after Day 3 are not known. The objective of the present study was to determine whether our TC AI instrument affected sperm transport, pregnancy rates, or lambing rate. In Experiment 1, ewes were assigned to two treatments: TC AI using the new TC AI instrument (n=10) or AI via laparotomy using a laparoscopic AI instrument (n=10). Twenty hours after artificial insemination, the uterine horns and oviducts were recovered and flushed to collect spermatozoa. Sperm transport did not differ (P>0.05) between the two treatments. In Experiment 2, ewes were assigned to three treatments: TC AI using the new TC AI instrument+sham intrauterine AI via laparotomy (n=29); sham TC AI+intrauterine AI via laparotomy using a laparoscopic AI instrument (n=29); and sham TC AI+intrauterine AI via laparotomy using the new TC AI instrument (n=30). On Day 14 after AI, uteri were collected and flushed to recover blastocysts. Transcervical deposition of semen reduced (P<0.05) Day 14 pregnancy rate (17.2% versus 61%), but intrauterine deposition of semen using the TC AI instrument via midventral laparotomy increased (P<0.05) Day 14 pregnancy rate (76.6% versus 44.8%). In Experiment 3, ewes were assigned to two treatments: sham cervical manipulation (n=40) or cervical manipulation to mimic TC AI (n=40). Immediately after treatment, each ewe was mated with a ram and watched until the ram mounted and ejaculated into the ewe. Treatment did not affect Day 30 or 50 pregnancy rate (67.5 and 66.2%, respectively), determined ultrasonically, or lambing rate (62.5%). The differences between Days 30 and 50 pregnancy rates and lambing rate were not significant. In Experiment 4, ewes were assigned to two treatments: TC AI (n=99) or laparoscopic AI (n=99). Transcervical AI reduced (P<0.01) Day 30 (TC AI versus laparoscopic AI; 5.0% versus 46.0%) and Day 50 pregnancy rates (4.0% versus 41.0%), determined ultrasonically, and lambing rate (4.0% versus 41.0%). Although the TC AI procedure significantly reduced pregnancy and lambing rates, large numbers of spermatozoa deposited at natural insemination seemed to compensate. Because our TC AI procedure has all but eliminated any visual evidence of trauma, and because the procedure does not seem to affect sperm transport or embryonal survival until Day 3, we speculate that cervical manipulation associated with TC AI may activate pathways that interrupt pregnancy between Days 3 and 14.     

Comparison of prostaglandin- and progesterone-based protocols for timed artificial insemination in sheep

The objective was to compare the reproductive performance of a new PGF_2α-based timed artificial insemination (TAI) protocol in sheep (Synchrovine®: two doses of PGF_2α, 7 d apart) to a traditional progesterone-eCG (P4-eCG) protocol, considering the effects of seminal state, AI-times, and AI-pathway. Three experiments involving 1297 multiparous Australian Merino ewes were done during the physiologic breeding season (location 32 °S-57 °W). Reproductive performance was assessed as non-return rate to service 21 d after AI (NRR21d), based on detection with androgenized wethers, as well as Fertility (pregnant/inseminated ewes), Prolificacy (fetuses/pregnant ewe), and Fecundity (fetuses/inseminated ewe), which were based on transabdominal ultrasonography 50 d after TAI. In Experiment 1, Synchrovine® treated ewes TAI cervically with fresh semen at 42, 48, or 54 h had similar NRR21d (0.51, 0.46, 0.57), Fertility (0.27, 0.31, 0.26), and Fecundity (0.29, 0.32, 0.27), all of which were lower (P < 0.05) than in a control P4-eCG group inseminated at 54 h (0.61, 0.48, 0.52, NRR21d, Fertility and Fecundity respectively). In Experiment 2, using chilled semen and cervical TAI, Synchrovine® treated ewes inseminated at 42 h yielded lower (P < 0.05) NRR21d, Fertility and Fecundity (0.28, 0.06, 0.06) compared to 48 (0.43, 0.24, 0.24) and 54 h (0.44, 0.22, 0.23). In Experiment 3 with chilled semen, Synchrovine® treated ewes TAI into the cervix at 51 or 57 h were similar in NRR21d (0.16 vs 0.20), Fertility (0.12 vs 0.14), and Fecundity (0.12 vs 0.15), respectively; but lower (P < 0.05) than P4-eCG treated ewes TAI at 54 h (0.34, 0.28, and 0.33 for NRR21d, Fertility and Fecundity respectively). Synchrovine® treated ewes intrauterine TAI at 51 or 57 h yielded similar NRR21d (0.51 vs 0.58), Fertility (0.43 vs 0.51), and Fecundity (0.45 vs 0.56) respectively, but lower (P < 0.05) results compared to P4-eCG treated ewes (0.75, 0.71, and 0.88 for NRR21d, Fertility and Fecundity respectively). In conclusion, AI-time in Synchrovine® treated ewes with fresh semen might be extended (42 to 54 h after the second PGF_2α), but should be delayed to 48-54 h with chilled semen and cervical AI. Independent of the seminal state, AI-time or AI-pathway, Synchrovine® yielded lower reproductive results than a conventional P4-eCG protocol.     

Luteolysis and cycle synchronization with a new prostaglandin analog for artificial insemination in the mare

Over a period of three years, 165 cyclic or anestrous Hanoverian mares received 177 treatments with 2, 3 or 4 mg of a novel luteolytic prostaglandin analog, K 11941. Heat and ovulations, indicating luteolysis, were observed after an average of 3.98 and 7.62 days, respectively, in 88.04% of 142 treated cyclic, postpartum and anestrous mares, and mares after an early loss of the conceptus. All doses tested were effective in inducing luteolysis as confirmed by determination of progesterone blood levels in samples collected daily, in 70 of 80 mares studied (87.5%). Attempts to achieve control of the cycle by various methods revealed that with K 11941 given once or twice, alone or in combination with hCG and/or an GnRH analog (Hoe 766), one can effectively concentrate estrus periods and follicular growth patterns, but can neither synchronize nor concentrate ovulations. Since most of the mares treated were confirmed problem mares, the pregnancy rate of 40.0% from first insemination at drug induced estrus, was regarded as satisfactory when compared to a pregnancy rate of 43.8% obtained with natural breeding in the same population. No drug related clinical signs of side-effects were observed.     

Factors influencing the success of vaginal and laparoscopic artificial insemination in churra ewes: a field assay

Pregnancy rate following artificial insemination (AI) in sheep is variable depending on several factors. The Churra breed (milk breed of the North-West of Spain) yields lower fertility results compared with other local and European breeds (Manchega, Latxa, Merino, Lacaune, Sarde, etc.). In this work we studied the influence of many factors on the fertility of the Churra breed (insemination technique, year, farm, age, male, number of inseminations per ewe, lambing-insemination interval and technician), analyzing lambing data obtained after 44448 inseminations (39.67% cervical AI via vagina, AIV, and 60.33% intrauterine AI using laparoscopy, AIL) in a categorical model. The most important factors influencing fertility after AI were farm, year, season, AI technique, and technician. AIL showed significantly higher fertility results than AIV (44.89% versus 31.25%). Season significantly affected fertility in both cases, but differences were more evident in AIV. Fertility dropped 1.74% (AIV) and 2.07% (AIL) per year as the ewes aged. Finally, AI fertility decreased when the lambing-insemination interval was lower than 10 weeks.     

Comparing ovulation synchronization protocols for artificial insemination in the scimitar-horned oryx (Oryx dammah)

Ovarian response and pregnancy success in scimitar-horned oryx (n=28) were compared, following treatment with two synchronization protocols and fixed-time artificial insemination (AI) with frozen-thawed semen. Each oryx received two injections of 500 microg of prostaglandin-F(2alpha) analogue (PGF(2alpha)-only) 11 days apart, and half received PGF(2alpha) in combination with an intravaginal progesterone-releasing device (CIDR11+PGF(2alpha)). Semen was collected by electroejaculation from anaesthetised adult oryx and cryopreserved. Anaesthetised females were transcervically inseminated 56.0+/-1.1 h (+/-S.E.M.) after PGF(2alpha) injection and/or device withdrawal using 28.0+/-1.5x10(6) motile thawed sperm. Ovarian endocrine response was monitored in 20 females by analysing faecal oestrogen and progesterone metabolites. Periovulatory oestrogen peaks were detected in 19/20 (95%) females after synchronization. There were no between-treatment differences in oestrogen concentrations or peak characteristics (P0.05). Luteal development after synchronization was delayed in half the progesterone treated (CIDR11+PGF(2alpha)) females, and faecal progestin excretion profiles indicated that the ovulatory follicle associated with synchronization either failed to ovulate or to fully lutenise. Pregnancy was diagnosed by ultrasonography and/or rectal palpation and was monitored by faecal progestin excretion. More (P=0. 013) pregnancies resulted from the PGF(2alpha)-only treatment (37.5%, 5/14) than from the CIDR11+PGF(2alpha) treatment (0/14), and four healthy scimitar-horned oryx calves were born, three after gestation intervals of 247 days and one after 249 days.     

Alternatives to improve a prostaglandin-based protocol for timed artificial insemination in sheep

The objective was to improve the reproductive performance of a prostaglandin (PG) F_2α-based protocol for timed artificial insemination (TAI) in sheep (Synchrovine®: two doses of 160 μg of delprostenate 7 d apart, with TAI 42 h after second dose). Three experiments were performed: Experiment 1) two doses of a PGF_2α analogue (delprostenate 80 or 160 μg) given 7 d apart; Experiment 2) two PGF_2α treatment intervals (7 or 8 d apart) and two times of TAI (42 or 48 h); and Experiment 3) insemination 12 h after estrus detection or TAI with concurrent GnRH. Experiments involved 1131 ewes that received cervical insemination with fresh semen during the breeding season (32/34 °S–58 °W). Estrous behaviour, conception rate, prolificacy, and fecundity (ultrasonography 30–40 d), were assessed. In Experiment 1, ewes showing estrus between 25 and 48 h or at 72 h after the second PGF_2α did not differ between 80 and 160 μg of delprostenate (73 vs 86%, P = 0.07; and 92 vs 95%, P = NS, respectively). Conception rate and fecundity were lower (P < 0.05) using 80 vs 160 μg (0.24 vs 0.42, and 0.27 vs 0.47, respectively). In Experiment 2, giving PGF_2α 7 d apart resulted in higher (P < 0.05) rates of conception (0.45 and 0.51) and fecundity (0.49 and 0.53) than treatments 8 d apart (conception: 0.33 and 0.29; fecundity: 0.33 and 0.34) for TAI at 42 and 48 h, respectively. In Experiment 3, rates of conception, prolificacy and fecundity were similar (NS) between Synchrovine® with TAI at 42 h (0.50, 1.13, and 0.56) and AI 12 h after estrus detection (0.47, 1.18, and 0.55), and Synchrovine® plus GnRH at TAI (0.38, 1.28, and 0.49). However, all TAI treatments had lower (P < 0.05) prolificacy and fecundity compared to AI following detection of spontaneous estrus (1.39 and 0.83, respectively). In conclusion, the Synchrovine® protocol was: a) more successful using 160 vs 80 μg delprostenate; b) more successful with a 7 d than 8 d PGF_2α interval; c) similarly effective for TAI versus AI 12 h after estrus detection; and d) not improved by giving GnRH at TAI.     

Ovsynch synchronization and fixed-time insemination in goats

This study assessed the efficacy of an Ovsynch protocol (vs. the classical cronolone containing vaginal sponge+eCG treatment) to generate fixed-time insemination in goats during the breeding season. Each regimen was applied to 24 Boer goat does. Onset and duration of estrus were determined with an aproned male and follicular development was monitored by ultrasonography. Ovulation and quality of the corpora lutea were established from progesterone concentrations. In 10-11 goats per group, LH concentrations were determined throughout the preovulatory period. Does were inseminated at pre-determined times (16 h after the second GnRH injection and 43 h after sponge removal). Estrus was identified in 96% of the Ovsynch-treated goats (at 49 h after prostaglandin injection) and in 100% of the goats synchronized with sponges (at 37 h after sponge removal). Low progesterone concentrations at the time of AI were observed in 21/24 and 24/24 goats synchronized by Ovsynch and sponges, respectively. Synchronization of the LH surge was tighter following Ovsynch compared to sponge treatment. Kidding rates (at 58 and 46% in the Ovsynch and sponge groups, respectively) and prolificacy (at 1.86 and 1.83 in the Ovsynch- and sponge-treated goats) were similar for both groups, as were the number of ovulations (2.9 and 3.3) and the proportion of does with premature corpus luteum regression (29 and 17%). When excluding does with premature luteal regression and those with low progesterone levels when receiving prostaglandins, kidding rate reached 87.5% (14/16) after Ovsynch. During the breeding season, the Ovsynch protocol may thus be an useful alternative to the sponge-eCG treatment.     

Behavioral and endocrine responses of hair sheep ewes exposed to different mating stimuli around estrus

Hair sheep ewes were used to evaluate the influence of various levels of mating stimuli on the duration and timing of estrus and LH concentrations around estrus. Ewes were treated with PGF2alpha (15 mg, im) 10 d apart. At the time of the second PGF2alpha treatment (Day 0) ewes were placed in groups and exposed to different types of mating stimuli. One group of ewes (n = 16) was exposed to an epididymectomized ram (RAM), a second group of ewes (n = 16) was exposed to an epididymectomized ram wearing an apron to prevent intromission (APRON) and a third group of ewes (n = 17) was exposed to an androgenized ovariectomized ewe (T-EWE). Jugular blood samples were collected from ewes at 6-h intervals through Day 5. Plasma was harvested and LH concentration was determined by RIA. The ewes were observed at 6-h intervals to detect estrus. A ewe was considered to be out of estrus when she no longer stood to be mounted by the teaser animal. There was no difference (P > 0.10) in the proportion of ewes expressing estrus (79.6%) or having an LH surge (85.7%) among the treatments. Neither the time to estrus nor the duration of estrus were different (P > 0.10) among APRON, RAM or T-EWE groups (41.6+/-3.8 vs 43.6+/-3.6 vs 46.1+/-3.6 h, respectively, and 26.5+/-2.2 vs 24.8+/-2.3 vs 30.5+/-2.2 h, respectively). The time to LH surge was similar (P > 0.10) among APRON, RAM and T-EWE groups (51.2+/-4.5 vs 51.2+/-4.7 vs 52.7+/-4.5 h, respectively). The magnitude of the LH surge was similar (P > 0.10) in the T-EWE, APRON and RAM ewes (99.7+/-4.9 vs 87.2+/-4.9 vs 85.8+/-5.0 ng/mL, respectively). The time from estrus to the LH surge was not different (P > 0.10) among APRON, RAM or T-EWE ewes (10.1+/-2.2 vs 9.8+/-2.3 vs 11.6+/-2.3 h, respectively). These results show that the expression and duration of estrus are not influenced by different types of mating stimuli in hair sheep ewes. In addition, the timing and the magnitude of LH release does not appear to be influenced by mating stimuli around the time of estrus.     

Short-term dietary effects on reproductive wastage after induced ovulation and artificial insemination in primiparous lactating Sarda ewes

Short-term effects of nutrition on conception rate (CR), ovulation rate (OR), ova and embryo losses (OEL) during the first 50 days following insemination and total reproductive wastage after ovulation (TRW), were investigated in primiparous lactating Sarda ewes after oestrous synchronisation and cervical [corrected] artificial insemination (AI). Eighty ewes grazing a green high-quality pasture were offered one of three iso-energetic supplements from day 14 before to day 2 after AI: whole maize grain (M); soyabean meal (S); maize gluten meal (G); or served as controls (C). Supplements G and S were iso-nitrogenous but provided different amounts of rumen undegradable digestible protein. The intake of herbage and digestible dry matter, measured by the n560 mg/l was associated with lower CR. Ranking by ovulation groups of CR was single相似文献   

Caffeine and artificial insemination

We studied the reactivation of cells and the repair of photomutagenic damage induced by xanthotoxin and visnagin plus NUV in arg-1 cells of Chlamydomonas reinhardii. Maintenance of liquid cultures in the dark resulted only in a slight reactivation of cells, even after 24 h. Repair of photomutagenic damage was more efficient: within 24 h the number of Arg⁺ revertants was reduced by 50% in cells cultured in the dark at 20°C. The repair was more efficient at 30°C. At the beginning of dark cultivation an after-effect could be observed. Cultivation in standard white light instead of dark after treatment resulted in a very strong after-effect. Therefore it was not possible to detect any photoreactivation.After treatment with xanthotoxin plus standard white light (24 h) neither reactivation of cells nor repair of photomutagenic damage was found. The after-effect was higher than after xanthotoxin plus NUV. It is possible that a small amount of repair could be masked by the after-effect.Treatment with visnagin yielded similar results. The photomutagenic effect of visnagin is described for the first time in this paper. The drug is a much less effective photomutagen than xanthotoxin. The photomutagenesis of visnagin may be attributable to photoproducts similar to those formed after treatment with furocoumarins.No definite conclusion can be drawn from the present results regarding the basis for the observed lack of repair (or reduced repair) after standard white light treatment; a possible cause might be a preferential formation of bi-adducts under these conditions.     

Ram-induced ovulation to improve artificial insemination efficiency with frozen semen in sheep.

Ram effect, defined as shortening of seasonal anestrus in ewes by exposure to the ram, is now well recognized but the underlying mechanisms are still unclear. Little information also exists whether the ram is able to influence the estrus cycle and ovulation. Three experiments were conducted to investigate endocrine response, time of ovulation and pregnancy rate of ewes in proestrus, exposed to the ram (treated) or an adult ewe (control). In the first experiment, ewes (n = 20) were treated with fluorgestone acetate pessaries for 12 days and were given eCG and cloprostenol one day before withdrawal of pessaries. On the day after removal of the pessaries ewes in the treated group (n = 10) were exposed to the ram and those in the control group (n = 10) were exposed to an adult ewe. Blood samples were taken for LH assay every 20 min from 2 h before to 24 h after ram exposure. In the second experiment, ewes (n = 120) were induced into proestrus and on the day after removal of the pessaries were exposed to either a ram (n = 60) or a ewe (n = 60) as described above and were laparoscoped 50, 60 or 70 h after pessary withdrawal (n = 20 at each time interval). In the third experiment ewes (n = 90) were induced and exposed to the ram (n = 45) or an adult ewe (n = 45) and inseminated via a laparoscope whit frozen-thawed semen at 50 or 60 h after pessary removal, respectively. Exposure to the ram was followed in 2 h by a marked rise in LH, equivalent to a preovulatory surge in duration and amplitude. It was also followed by concentrated ovulation within 25 to 30 h and by an increased pregnancy rate in exposed ewes (73.3 vs. 53.3%).     

Fertility of ewes following artificial insemination with semen frozen in pellets or straws, a preliminary report

In two trials involving the artificial insemination of 194 ewes, the fertility of ram semen was examined following freezing, either in pellet form or in straws, and after storage in a chilled state (15 degrees C) for up to 16 hours. Estrus was synchronized in ewes by intravaginal sponge (MAP) treatment for 14 days. At sponge removal 600 IU PMSG was injected and the ewes received two inseminations 50 and 60 hours later. Fertility was assessed at lambing. In trial 1, the mean lambing rate of 52% (16 31 ) for semen frozen in pellets was higher than 29% (9 31 ) for semen frozen in straws but this difference was not significant. In trial 2, ewes inseminated with chilled semen and semen frozen in pellets had lambing rates of 83% (44 53 ) and 55% (44 79 ) respectively (P<0.001).     

Development of a new transcervical artificial insemination method for sheep: effects of a new transcervical artificial insemination catheter and traversing the cervix on semen quality and fertility

The difficulty of traversing the cervix severely limits transcervical artificial insemination (TC AI) in sheep. Cervical trauma and poorly designed instruments can reduce fertility after AI. To overcome problems associated with TC AI, we developed a new TC AI catheter. Three bench experiments were conducted to determine the effects of the new TC AI catheter on semen quality independent of the effects of moving the catheter through the cervix. In each of the three bench experiments, the standard laparoscopic instrument for intrauterine AI in sheep was used as the control for the TC AI catheter. In Experiment 1, the total volume of semen extender expelled and void volumes for both types of AI instrument (TC versus laparoscopic) were determined. In Experiment 2, the effects of each type of AI instrument (TC versus laparoscopic) on semen quality, estimated as percentage motility and percentage forward progressive motility, of frozen-thawed semen was determined. In Experiment 3, the effects of both types of AI instrument (TC versus laparoscopic) on number of spermatozoa expelled was determined. The type of AI instrument affected neither semen quality nor the number of spermatozoa expelled. However, void volume differed (P < 0.01) between the two instruments. After differences in void volume were taken into account, an in vivo experiment was conducted to determine whether using our new TC AI catheter for TC or surgical intrauterine AI affected fertilization and pregnancy rates. For this, ewes were assigned to one of three treatments: (1) TC AI using the new TC AI catheter + sham AI via laparotomy (n = 9); (2) sham TC AI + AI via laparotomy using a laparoscopic AI instrument (n = 8); and (3) sham TC AI + AI via laparotomy using the new TC Al catheter (n = 10). To synchronize estrus, progestogenated pessaries were inserted and left in place for 12 days. On Day 5 after pessary insertion, PGF2alpha (15 mg) was given i.m. At pessary removal, 400 IU of eCG were administered i.m. Ewes were inseminated 48-52 h after pessary removal using fresh diluted semen (200 x 10(6) to 350 x 10(6) spermatozoa per 0.2 ml) pooled from the same four rams each day during the experiment. At 72 h after AI, uteri were collected postmortem and flushed. Oocytes and embryos were recovered and evaluated. Treatments did not affect (P > 0.01) ovum and embryo recovery rate (mean = 87.3%), fertilization rate (59.3%), or Day 3 pregnancy rate (mean = 66.6%). We conclude from these data that the use of our new TC AI catheter for TC AI or intrauterine AI should not impair the success of AI in sheep.