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A. Zaidi, R. G. Lendon, J. S. Dixon and I. D. Morris

Summary. Male rats were injected with 50 mg ethylene-1,2-dimethanesulphonate/kg from Day 5 to Day 16 after birth and control rats received injections of the same volume of vehicle. Testes were studied at various times from Day 6 to Day 108 using histochemistry, light and electron microscopy. Fine structural degenerative changes were observed in the Leydig cells and seminiferous tubules of EDS-treated animals as early as Day 6. By Day 11 no Leydig cells could be detected and the interstitia of EDStreated testes contained large numbers of fibroblast-like cells which formed peritubular collars 3–5 cells thick; the tubules contained Sertoli cells with heterogeneous inclusions and large numbers of lipid droplets. A small number of Leydig cells was found at Day 14 and their numbers increased so that, in animals of 28 days and older, large clusters of Leydig cells were present between severely atrophic tubules. These tubules contained Sertoli cells with few organelles; germinal cells were not observed after 28 days in EDStreated animals.

These results show that EDS destroys the fetal population of Leydig cells postnatally and this mimics the well documented effect of EDS on adult Leydig cells. The seminiferous tubules were permanently damaged by EDS in the present experiments. Tubular damage could have been due to a direct cytotoxic effect of multiple injections of EDS on the tubule before the blood–testis barrier develops or due to withdrawal of androgen support secondary to Leydig cell destruction.

Keywords: testis; Leydig cell; development; EDS; cytotoxicity; rat

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J. D. Savio, W. W. Thatcher, G. R. Morris, K. Entwistle, M. Drost and M. R. Mattiacci

The effects of concentration of progesterone in plasma on development and fertility of the first wave dominant follicle were studied in cattle. To identify a source of exogenous progesterone that would permit extension of the first wave dominant follicle, nonlactating Holstein cows (n = 6) received on day 8 of two successive oestrous cycles an injection of PGF (25 mg) and a new (1.9 g of progesterone (Period 1)) or used (≈ 1.2 g of progesterone (Period 2)) CIDR-B device that was removed on day 17. Control cows (n = 6) received a new CIDR-B device on day 8 that was removed on day 17 and a PGF injection (25 mg) on day 17. Ultrasonography and collection of blood samples were performed on alternate days throughout the experiment. Plasma concentrations of progesterone and oestradiol were different between treatments (P < 0.0001 and P < 0.05, respectively). The dominant follicle was maintained until day 17 and ovulated upon removal of the intravaginal device in 1 of 6, 6 of 6 and 0 of 6 in new CIDR-B, used CIDR-B and control groups, respectively (P < 0.01). The preovulatory dominant follicles were 14.2 ± 1.6 mm, 20 ± 1.3 mm and 10 ± 1.3 mm, respectively (P < 0.001) on day 17. There were fewer 5–9 mm follicles in cows having a persistent dominant follicle (P < 0.01). The interval to onset of oestrus was negatively correlated with size of the dominant follicle on day 17 (P < 0.001). In Expt 2, the fertility of oocytes ovulated from new (PGF on day 7; T1; n = 91) and persistent dominant follicles (PGF on day 7 and a used CIDR-B device inserted on day 7 and withdrawn on day 16; T2; n = 91) was tested using Holstein heifers. Size of the dominant follicle and plasma concentrations of progesterone and oestradiol on days 7 (T1) and 16 (T2) were different between treatments: 11.3 ± 0.2 versus 16.2 ± 0.3 mm (P < 0.001); 4.2 ± 0.2 versus 2.9 ± 0.3 ng ml−1 (P < 0.01) and 3.5 ± 0.3 versus 11.7 ± 1.7 pg ml−1 (P < 0.01), respectively. Pregnancy rates at first artificial insemination were 64.8% (46 of 71) and 37.1% (26 of 70) for new and persistent dominant follicles, respectively (P < 0.01). Pregnancy rates at second service were 50% and 52.8%, respectively. Low plasma concentrations of progesterone, therefore, resulted in persistency of the dominant follicle and temporarily impaired fertility.

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D. G. Morris, M. G. McDermott, M. Grealy, M. G. Diskin, C. A. Morrison, P. J. Swift and J. M. Sreenan

The effects of immunizing cattle against either of two peptides from the amino terminal peptide (αN) of the α43-subunit of bovine inhibin on ovulation rate, gonadotrophin concentration and fertility were investigated. Two peptide sequences from the αN-subunit of bovine inhibin (P1N, bIα-(8-20) and P2N, bIα-(153-167)) were synthesized and conjugated to human serum albumin (HSA). Hereford-cross heifers (n = 5 per group) were given an initial injection of 3 mg of one of the peptide conjugates, followed by three booster injections (1.5 mg) at intervals of 11 weeks. Control heifers (n = 5) were injected with HSA only. Blood samples were taken once a week to measure antibody titre and every hour at about the time of the first oestrus and during the mid-luteal phase after the second booster injection, to measure FSH and LH concentrations. Ovulation rate was measured by ultrasonography. Gonadotrophin concentrations were analysed for four periods relative to the peak (time = 0 h) of the preovulatory LH surge as follows: pre-surge: −16 to −5 h; surge: −4 to 4 h; post-surge: 5 to 16 h and a period of 12 h during the mid-luteal (days 10–12) phase. Antibodies that bound to the individual peptides were generated and the ovulation rate increased (P < 0.05) in immunized heifers. Control heifers had one ovulation at all ovulatory cycles monitored. In group P1N, one heifer had two ovulations at each of the six cycles monitored, while another heifer had two ovulations at one cycle. Three heifers in group P2N had more than one ovulation; one heifer had a superovulation response on seven occasions and another heifer on three occasions while the third heifer had two ovulations at one cycle. The superovulation responses ranged from three to eight ovulations. There was no evidence of a significant correlation between peptide antibody titre, ovulation rate and FSH at any of the periods studied. After mating following the third booster injection, nine of the ten immunized and all five control heifers calved. One heifer from group P1N and two heifers from group P2N gave birth to twin calves. These data show that in cattle immunization against the αN-subunit of bovine inhibin significantly disrupts the mechanism(s) controlling ovulation rate but does not impair fertility.

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D. G. Morris, M. G. McDermott, M. G. Diskin, C. A. Morrison, P. J. Swift and J. M. Sreenan

Three peptide sequences from the bovine inhibin α-subunit (P1: 18–30; P2: 63–72 and P3: 107–122) were synthesized and conjugated to human serum albumin (HSA). Hereford crossheifers (n = 5 per group) were injected with 3 mg of one of the peptide conjugates, followed by three booster injections at intervals of 11 weeks. Control heifers (n = 5) were injected with HSA only. Antibodies recognizing both the individual peptides and 32 kDa bovine inhibin were generated and ovulation rate was increased in peptide immunized heifers. In group P1, 1 of 5 heifers responded with an increased ovulation rate whereas in groups P2 and P3, 5 of 5 and 4 of 5 heifers, respectively, had an increased ovulation rate. In group P2, in the first oestrous cycle following booster injections 2 and 3, 4 of 5 and 3 of 5 heifers, respectively, responded with twin ovulations, whereas a fourth heifer had three ovulations following booster injection 3. After breeding following booster injection 3, 3 of 5 heifers in group P2 and 1 of 5 in group P3 gave birth to twin calves. This study demonstrates the potential of immunizing against synthetic peptide sequences of the α-subunit of bovine inhibin to increase ovulation and twinning rates in cattle.