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M. Mihm, A. Baguisi, M. P. Boland and J. F. Roche

After luteolysis, subluteal concentrations of progesterone or treatment with a synthetic progestagen result in an extended period of dominance (persistence) of the dominant follicle in cattle. Two experiments studied (1) the relationship between the duration of dominance of the ovulatory follicle and pregnancy rate and (2) the ability of a persistent dominant follicle to ovulate and form a normal functioning corpus luteum. In Expt 1, beef heifers were either untreated (n = 30) or given a synthetic progestagen implant (3 mg norgestomet) for 12 days starting on day 16 of their cycle (n = 32). The mean duration of dominance of the ovulatory follicle differed (P < 0.05) between treated and control heifers (10.8 ± 1.2 and 3.3 ± 0.8 days, respectively) and 20 of 26 control and 7 of 30 treated heifers were diagnosed pregnant 28 days after artificial insemination (P < 0.01). In Expt 2, on the first day of dominance of the second dominant follicle, heifers received either a PGF analogue alone (controls; n = 18), or prostaglandin and a norgestomet implant for 6 (T6; n = 19) or 10 days (T10; n = 20). Increases in the duration of dominance of the second dominant follicle (controls, 4.1 ± 0.2 days; T6, 8.6 ± 0.2 days; T10, 12.1 ± 0.2 days; P < 0.05) resulted in a decrease in pregnancy rate (controls, 14 of 16; T6, 11 of 19; T10, 0 of 13; P ≤ 0.05). Progesterone concentrations on days 7 and 12 and the area of luteal tissue on day 12 after artificial insemination were not different (P > 0.05) between treatments. It is concluded that (1) treatment with a synthetic progestagen towards the end of the luteal phase causes a variable extension of the period of dominance of the ovulatory follicle with a significant reduction in pregnancy rate, (2) the persistent dominant follicle can ovulate and form a functional corpus luteum, and (3) the pregnancy rate is sequentially decreased as the duration of dominance increases from 4 to 8 days, and is further significantly reduced if the duration of dominance exceeds 10 days.

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M. G. Murphy, M. P. Boland and J. F. Roche

Summary. The ovaries of 18 post-partum beef suckler cows were examined daily, using ultrasound, from Day 5 post partum until a normal oestrous cycle was completed. Periods of growth and regression of medium-sized (5–9 mm) follicles were identified before one medium follicle became dominant (single large follicle ≥ 10 mm). The mean (±s.e.m.) number of days from parturition to detection of the first post-partum dominant follicle was 10·2 ± 0·5. The first post-partum dominant follicle ovulated in 2/18 (11%) cows. The interval from calving to first ovulation (mean ± s.e.m. = 35·9 ± 3·3 days) was characterized by the growth and regression of a variable number (mean = 3·2 ± 0·2; range 1–6) of dominant follicles. The maximum diameter of the dominant follicle increased as the cows approached first ovulation (P < 0·05). Behavioural oestrus was not detected in 16/18 (89%) cows at first ovulation. Following first ovulation, the length of the subsequent cycle was short (mean = 9·7 ± 0·5 days; range 8–15 days) in 14/18 (78%) cows and was characterized by the development and ovulation of a single dominant follicle. During oestrous cycles of normal length (mean = 20·6 ± 0·5 days; range 18–23 days) one (N = 2), two (N = 7) or three (N = 8) dominant follicles were identified. The growth rate, maximum diameter or persistence of non-ovulatory dominant follicles before first ovulation or during oestrous cycles were not different (P > 0·05). These data show that, in beef suckler cows, follicular development and formation of a dominant follicle occur early after parturition and the incidence of ovulation of the first dominant follicle is low. The number of dominant follicles that develop before first ovulation is variable; first ovulation is rarely associated with oestrus and short cycles are common after first ovulation. It is concluded that prolonged anoestrus in post-partum beef suckler cows is due to lack of ovulation of a dominant follicle rather than delayed development of dominant follicles.

Keywords: follicle; post partum; beef suckler; cow; ultrasound

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J. D. Savio, M. P. Boland and J. F. Roche

Summary. The resumption of ovarian activity after normal calvings was studied in 18 lactating Friesian cows. Since, in 17 cows, first post-partum ovulation occurred without overt oestrous behaviour being detected, the resultant cycles were called 'ovarian cycles'. The mean (± s.d.) length of the ovarian cycles was 21·0 ± 8·7 days. The duration of cycles tended to be normal (18–24 days) or long (≥25 days) when the ovulatory dominant follicles were identified before Day 10 post partum; they were consistently short (9–13 days) when dominant follicles identified after Day 20 post partum ovulated. When such follicles were detected between Days 10 and 20 post partum, long, normal and short ovarian cycles were detected. The number of waves of follicular growth with associated dominant follicles observed during the ovarian cycles tended to be related to cycle length; short cycles had 1 dominant follicle, normal cycles predominantly 2, and long cycles mostly 3 dominant follicles. The mean (± s.d.) duration of 13 oestrous cycles studied was 23·1 ± 2·1 days. Of these cycles, 7 had 3 and 6 had 2 dominant follicles. The oestrous cycles with 3 dominant follicles had a mean (± s.d.) duration of 24·0 ± 1·2 days and the respective dominant non-ovulatory follicles reached maximum sizes on Days 8 and 18, respectively; oestrous cycles with 2 dominant follicles were 22·2 ± 2·6 days in duration, and the dominant non-ovulatory follicle reached maximum size by Day 8. Ovarian follicular development during the first 45 days of pregnancy was characterized by the growth and regression of successive dominant follicles, each lasting 10–10 days. These results show that the first ovarian cycle was predominantly short when the ovulatory dominant follicle was first detected after Day 20 post partum.

Keywords: dairy cow; dominant follicle; post partum; ovarian activity; ultrasound

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J. D. Murray, M. P. Boland and C. Moran

Summary. Chromosomal analysis was carried out on 48 Day 2–7 embryos collected from superovulated Merino ewes. Three embryos had abnormal chromosome complements (1 × IN, 1 × 1N/2N, 1 × 3N), yielding an incidence of 6·25% abnormal embryos. It is concluded that superovulation does not cause an increase in the incidence of chromosomal abnormalities in embryos of Merino sheep.

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M. Mihm, N. Curran, P. Hyttel, P. G. Knight, M. P. Boland and J. F. Roche

The aim of the present study was to characterize in detail the cytoplasmic and nuclear morphology of cattle oocytes recovered from follicles that are dominant for more than 9 days (with low fertility after ovulation), and to relate morphological changes to intrafollicular markers of follicle health. Beef heifers received prostaglandin F and a synthetic progestagen (3 mg Norgestomet) for 2 or 10 days on the first day of dominance of the second dominant follicle (DF2) of the oestrous cycle, to give a 4 day (n = 19; N2) or 12 day (n = 21; N10) duration of dominance of the dominant follicle at ovariectomy 18 h after implant removal and before the predicted gonadotrophin surge. Ultrasound scanning determined emergence of a new wave of follicles in five N10 heifers the day before (n = 1) or day of ovariectomy (n = 4) (N10-NonDom). Dominant follicles from the remaining N10 heifers (N10-Dom) were larger (P < 0.05) on the day of ovariectomy (17.8 ± 0.6 mm) than those from N2 heifers (13.6 ± 0.4 mm). The oestradiol:progesterone ratio of follicular fluid from N10-Dom heifers was reduced (21.7 ± 3.1 versus 34.1 ± 4.4; P < 0.05), while inhibin A (as measured by immunoradiometric assay) was increased (12.7 ± 1.0 versus 9.0 ± 0.7 μg ml−1; P < 0.05) compared with N2 heifers. Eleven of twelve N2 oocytes demonstrated nuclear activation without germinal vesicle breakdown, while seven of eight N10-Dom oocytes had undergone germinal vesicle breakdown and had progressed to metaphase I (6/8) or II (1/8). In contrast to N2 oocytes, N10-Dom oocytes showed a larger perivitelline space containing more cumulus cell process endings, vacuoles, irregular vesicles, and more mitochrondia and lipid droplets throughout the ooplasm, yet the degree of cumulus cell expansion and atresia was similar. Thus, final oocyte maturation leading to metaphase I is initiated in most dominant follicles with a dominance period of > 9 days before the gonadotrophin surge and is associated with a reduction in dominant follicle health. However, ovulatory ability is maintained and will lead to the ovulation of aged oocytes, markedly reducing subsequent pregnancy rates.

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P. Lonergan, H. Khatir, F. Piumi, D. Rieger, P. Humblot and M. P. Boland

In vitro produced bovine zygotes show substantial variation in the time required to complete the first cell cycle and in their in vitro development potential. A number of reports have highlighted the fact that the fastest developing embryos in vitro are most likely to be comparable with their in vivo counterparts. At 24 h after IVF, presumptive zygotes were cultured in droplets of synthetic oviduct fluid medium. Droplets were examined at regular intervals and all cleaved embryos at each time point were transferred into new droplets and cultured separately for the duration of the experiment. All uncleaved zygotes were returned to the incubator and re-examined at the successive time points until 48 h after insemination, at which time the remaining uncleaved oocytes were retained as a group. A representative number of day 7 blastocysts from zygotes that had cleaved by 30 or 36 h were transferred to synchronized recipients and pregnancy was diagnosed by ultrasonography at day 35. Glucose and glutamine metabolism was examined in zygotes and blastocysts and compared retrospectively with time of first cleavage. A representative number of blastocysts from each of the cleavage groups was sexed using PCR. Data were analysed by chi-squared and regression analysis. Development to the blastocyst stage decreased as the time from insemination to first cleavage increased (r = 0.97, P < 0.03). There was no difference in blastocyst hatching, number of blastocyst cells or pregnancy rate between the 30 and 36 h groups. The overall sex ratio was 62% males (n = 258, P < 0.0001) and was not different in the 30 and 36 h groups (61%, n = 155 versus 63%, n = 95, respectively). These results indicate that although time of first cleavage has a major influence on the probability of an embryo developing to the blastocyst stage, once that stage is attained, subsequent developmental characteristics are unrelated to the time of first cleavage.

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D. O'Callaghan, F. J. Karsch, M. P. Boland, J. P. Hanrahan and J. F. Roche

Summary. Photoperiod may regulate seasonal reproduction either by providing the primary driving force for the reproductive transitions or by synchronizing an endogenous reproductive rhythm. This study evaluated whether breed differences in timing of the reproductive seasons of Finnish Landrace (Finn) and Galway ewes are due to differences in photoperiodic drive of the reproductive transitions or to differences in photoperiodic synchronization of the endogenous rhythm of reproductive activity. The importance of decreasing photoperiod after the summer solstice in determining the onset and duration of the breeding season was tested by housing ewes from the summer solstice in either a simulated natural photoperiod or a fixed summer-solstice photoperiod (18 h light:6 h dark; summer-solstice hold). Onset of the breeding season within each breed did not differ between these photoperiodic treatments, but Galway ewes began and ended their breeding season earlier than Finn ewes. The duration of the breeding season was shorter in Galway ewes on summer-solstice hold than on simulated natural photoperiod; duration did not differ between photoperiodic treatments in Finn ewes. The requirement for increasing photoperiod after the winter solstice for initiation of anoestrus was tested by exposing ewes from the winter solstice to either a simulated natural photoperiod or a winter-solstice hold photoperiod (8·5 h light:15·5 h dark). Onset of anoestrus within each breed did not differ between these photoperiodic treatments, but the time of this transition differed between breeds. These observations suggest that genetic differences in timing of the breeding season in Galway and Finn ewes do not reflect differences in the extent to which photoperiod drives the reproductive transitions, because neither breed requires shortening days to enter the breeding season or lengthening days to end it at appropriate times. These findings are consistent with the hypothesis that photoperiod synchronizes an endogenous rhythm of reproductive activity in both breeds and that genetic differences in timing of the breeding season reflect differences in photoperiodic synchronization of this rhythm.

Keywords: breeding season; sheep; photoperiod

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M. A. Crowe, D. Goulding, A. Baguisi, M. P. Boland and J. F. Roche

There is a low incidence of ovulation of the first dominant follicle that develops in the early postpartum period of beef suckler cows, which prolongs the interval from calving to first ovulation. The objective of this study was to determine whether a single injection of a GnRH analogue would ovulate the first postpartum dominant follicle. Limousin × Friesian beef suckler cows were assigned at parturition, over two years (16 cows in year 1; 19 cows in year 2), to one of three treatments: (1) untreated (control; n = 12), (2) GnRH analogue (20 μg buserelin i.m.) administered in the growing–plateau phase of the first postpartum dominant follicle (GnRH-G; n = 12) and (3) GnRH analogue administered in the declining phase of the first postpartum dominant follicle (GnRH-D; n = 11). From day 8 or 9 post partum, the ovaries of each cow were examined daily by ultrasound to determine the time of GnRH injection and ovulation. Blood samples were collected daily for progesterone measurement to confirm ovulation and in year 2 to determine the duration of the first oestrous cycle. The mean (± sem) number of days from parturition to development of the first dominant follicle was 11.0 ± 0.3, 10.3 ± 0.5 and 10.1 ± 0.7 for cows assigned to treatments 1–3, respectively (P > 0.05). The proportion of cows ovulating the first dominant follicle was higher (P < 0.05) following GnRH treatment (12 of 12 and 7 of 10; GnRH-G and GnRH-D, respectively) than with controls (2 of 12). The mean interval from parturition to first ovulation in all cows in the GnRH-G treatment was reduced (16.1 ± 0.8 days; P < 0.01) compared with the interval for cows in the GnRH-D treatment (24.5 ± 3.6 days) or controls (27.1 ± 2.5 days). There was no difference in the duration of the first luteal phase post partum in control (9.3 ± 3.0 days) and GnRH-treated cows (8.0 ± 1.0 and 9.6 ± 3.7 days; GnRH-G and GnRH-D, respectively) or in the proportion of cows with short cycles (4 of 7, 5 of 6 and 4 of 5; control, GnRH-G and GnRH-D, respectively). In conclusion, a single injection of GnRH analogue during the growing–plateau or declining phase of the first postpartum dominant follicle of beef suckler cows induced ovulation in most cows but did not alter the proportion of cows with short cycles compared with controls.

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S. J. Sunderland, M. A. Crowe, M. P. Boland, J. F. Roche and J. J. Ireland

This study examined the correlation between measurement of follicle growth by ultrasound, and measurement of intrafollicular ratios of oestradiol and progesterone concentrations and the serum concentrations of FSH during selection, dominance and atresia or ovulation of dominant follicles in heifers. Heifers were ovariectomized on days 0 (before LH surge), 1 (after LH surge, preovulation), 1 (postovulation), 3, 6 and 12 of the oestrous cycle. Blood samples were collected at 4–6 h intervals. After ovariectomy all follicles ≥ 5 mm were measured and follicular fluid was aspirated. Follicles were classified by size according to ultrasound (F1, largest; F2, second largest; F3, all remaining follicles ≥ 5 mm) and by the ratio of oestradiol:progesterone concentrations. During the follicular phase, a single dominant oestrogen-active follicle increased in diameter while serum concentrations of LH increased and FSH decreased (P < 0.05). On day 1 (after LH surge, preovulation), serum LH and FSH decreased to pre-surge concentrations (P < 0.0001), while follicle size and intrafollicular progesterone concentration increased and oestradiol concentration decreased (P < 0.05). A dominant nonovulatory follicle, classified as oestrogen-active on days 1, 3 and 6 and oestrogen-inactive on day 12, increased in size from day 1 to day 7 and lost dominance during days 10–12, coincident with the growth of multiple oestrogen-active follicles. The serum FSH concentration increased transiently (P < 0.05) before each new wave of dominant follicular growth. The overall correlation of ultrasound measurements of follicle diameter with measures of follicle size after ovariectomy was high. The ratio of oestradiol:progesterone concentrations, but not of size, reliably distinguished potential dominant from atretic follicles. The size of the follicle and the oestradiol concentration were not determinants of subsequent dominance during a selection phase. We conclude that: (1) ovarian follicles go through selection, dominance and atresia phases coincident with transient increases and decreases in FSH; and (2) ultrasound is an accurate measure of follicle growth, but that size alone is not a sufficient measure to ascribe dominance and both ultrasound and the intrafollicular ratio of oestradiol:progesterone concentrations are needed to monitor selection, dominance and atresia of follicles accurately.

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R. M. McKeown, D. O'Callaghan, J. F. Roche and M. P. Boland

The effects of inhibin immunization on inhibin antibody titres, semen characteristics, scrotal size, fertility, FSH, LH and testosterone concentrations were determined by immunizing adult rams against bovine inhibin α1–26-Gly-Tyr conjugated to human serum albumin (n = 16) in non-ulcerative Freund's adjuvant and DEAE:dextran (1:1) or adjuvant alone (n = 16) on days 0 (29 June), 30, 60, 191, 303 and 394. Blood samples were collected and bovine inhibin α1–26-Gly-Tyr antibody titres and serum testosterone concentrations were determined. Each month, between days 174 and 417, semen was collected every 30 min to a maximum of 15 ejaculates over 7 h and scrotal circumference was measured. Ram fertility was recorded during natural service. FSH, LH and testosterone concentrations and GnRH-induced FSH and LH release were measured in a subgroup of immunized (n = 5) and control (n = 5) rams at frequent intervals. Antibody titres were variable among immunized rams (0–46% I-labelled bovine inhibin α1–26-Gly-Tyr at 1:1600 serum dilution) but mean titres were consistently higher than in control rams (P ≤ 0.001). Immunization did not alter the semen volume, output or quality of spermatozoa or ram fertility, but increased the mean scrotal circumference (37.6 ±0.8 cm versus 34.4 ± 0.7 cm, P < 0.001). Mean FSH concentrations were higher in immunized rams during two intensive blood sampling periods (in June and August) (5.8 ± 0.7 ng ml−1 versus 3.0 ± 0.3 ng ml−1, P < 0.001 in June; and 4.8 ± 0.9 ng ml−1 versus 2.0 ± 0.3 ng ml−1, P < 0.02 in August), and were correlated with antibody titres (r 2 = 0.3, P < 0.05 in June; and r 2 = 0.8, P < 0.001 in August). Discrete FSH pulses were not detected. Immunization did not alter mean or basal testosterone or LH concentrations, or LH pulse frequency; LH pulse amplitude was increased (1.6 ± 0.2 ng ml−1 versus 0.8 ± 0.2 ng ml−1, P < 0.02) and was correlated with antibody titres (r 2=0.6, P < 0.01). Immunization enhanced GnRH-induced FSH (P <0.05) but not LH release. In conclusion, immunization of adult rams against bovine inhibin α1–26 Gly-Tyr increased scrotal circumference, mean FSH concentrations and LH pulse amplitude, without altering semen characteristics, fertility, mean LH concentrations, LH pulse frequency or mean testosterone concentrations.