y1 y1
Ž xylazine hydrochloride, 3.2 mg ml
azaperone, and 0.4 mg ml fentanyl citrate 0.02
y1
. ml kg
liveweight Fentazin; Parnell Laboratories, Auckland, NZ . Uterine turgidity Ž
was reduced by an i.v. injection of 0.5 mg clenbuterol hydrochloride 5 ml Planipart; Ž
. .
Boehringer Ingelheim NZ , Auckland 2–3 min prior to laparoscopy. Each ovary was examined for the presence and the number of corpora lutea, luteinised follicles, and
non-luteinised follicles with a diameter greater than or equal to 8 mm. Upon the completion of the ovarian examination, the hinds were given an i.v. injection of 50 mg
Ž .
yohimbine hydrochloride 5 ml Recervyl; Aspiring Animal Services, Wanaka, NZ to reverse the effects of anaesthesia.
2.5. Measurement of oFSH antibody in plasma All hinds in the trial were blood-sampled by jugular venepuncture at the start, middle,
and end of the study. Plasma samples were diluted 1:100 in pH 7.0 phosphate buffered Ž
. Ž
. saline containing 0.1 BSA PBS BSA . Diluted samples 100 ml were incubated
overnight with 100 ml pre-precipitated sheep anti-rabbit second antibody, the samples were then incubated for 1 h and then centrifuged at 3200 rpm for 35 min after the
addition of 1.0 ml of 4 WrV PEG 6000. Samples were decanted and counted for 60 s. The oFSH binding activity of the plasma was determined from the ratio of bound to the
total counts following correction for non-specific binding controls.
2.6. Statistical analyses Ž
. Ovulation rate OR was assessed as the number of corpora lutea and the total
Ž .
follicular stimulation TFS was calculated as the sum of corpora lutea, luteinised Ž
qx
. follicles, and large non-luteinised follicles. OR and TFS data were log 1
trans- Ž
. Ž formed and analysed by residual maximum likelihood REML
Patterson and Thomp- .
son, 1971 with individuals as a random effect, and genotype, date, and their interaction as fixed effects. The assumption of individual randomness in ovarian response over time
was evaluated for each genotype independently by the non-parametric means test Ž
. Friedman’s Test based on ovulation rate rankings. This evaluation was conducted for
the first five consecutive treatments only due to missing values for some animals at subsequent dates.
3. Results
Loss of CIDR devices occurred on four occasions at various times during the study. The data at these observations were excluded from the study. There were highly
significant effects of both the season and genotype on ovarian response to oFSH Ž
. Ž
. treatment P - 0.05; Table 1 and Fig. 1 with minimal interaction P 0.10 . For all of
Ž .
the three genotypes, ovarian responses OR and TFS were greatest from March to Ž
. Ž
November hereafter referred to as the breeding season , and least even non-existent for .
Ž .
some genotypes in the period from December to January, inclusive Fig. 1 . Mean ovulation rates between March and November for red deer ranged from 3.7 in May to
G.W. Asher
et al.
r Animal
Reproduction
Science 59
2000 61
– 70
65 Table 1
Ž .
Summary of the overall ovarian response to FSH treatment by genotype
a b
Genotype Observation
Mean OR Mean TFS
ORrTFS Nil-responses
OR ?3 CL
OR ?10 CL
Ž . Ž
. Ž
. Ž . Ž
. Ž .
Ž . n
S.E.M. S.E.M.
ratio i.e., anovulation
Ž .
Ž .
Red 89
2.15 0.42 3.21 0.51
0.67 58.4
23.6 8.9
Ž .
Ž .
1 4 PD hybrid 77
1.17 0.18 1.61 0.23
0.73 45.5
18.2 Ž
. Ž
. Wapiti hybrid
59 6.32 0.92
7.25 0.96 0.87
35.6 55.9
32.2
a
Ž .
OR s ovulation rate number of CL .
b
TFSs total follicular stimulation.
Ž .
Ž .
Ž .
Fig. 1. Mean qS.E.M. ovulation rates shaded and total follicular stimulation open at each treatment Ž .
Ž . Ž .
period for a red deer, b 1 4 PD hybrids, and c F wapiti hybrids.
1
1.8 in August, with no indication of any trends associated with time during the breeding season. However, all red deer hinds were anovulatory during December and January,
and exhibited little in the way of any follicular stimulation. A similar seasonal pattern was observed for 1 4 PD hybrids, although the overall mean ovulation rates during the
breeding season, ranging from 2.3 in September to 0.5 in November, were consistently lower than for red deer. For F wapiti hybrids, the first two treatment regimens in March
1
Ž and April 1996 were associated with the highest mean ovulation rates 15.6 and 11.7,
. respectively observed at any time during the trial. Thereafter, mean values during the
Ž .
breeding season and including March 1997 ranged between 6.3 and 4.7. In contrast to red deer and 1 4 PD hybrids, the F wapiti hybrids did not all become anovulatory in
1
December and January, with observed mean ovulation rates of 3.0 and 0.5, respectively. Ž
. The overall mean ovulation rate for F wapiti hybrids Table 1 was significantly higher
1
Ž .
than for the other genotypes P - 0.05 . It is noteworthy that while ovulations were
recorded for all of the three genotypes in late September and early November, ; 60 of the CL observed showed signs of premature regression.
Genotype effects were further highlighted by the overall statistics on the proportion Ž
. of non-responders i.e., hinds anovulatory following treatment and the proportion of
Ž .
Ž hinds exhibiting either G 3 CL i.e., a ‘‘superovulation’’ response or G 10 CL i.e., a
. Ž .
‘‘mega-ovulation’’ response Table 1 . For the data pooled for the entire study, F
1
Ž .
wapiti hybrids had the lowest number of non-responders 35.6 and most responders Ž
. 55.9 and 32.2, respectively, for G 3 and G 10 ovulations . Red deer exhibited the
Ž .
Ž highest non-responder rate 58.4 , but 1 4 PD hybrids had the least responders 18.2
. Ž .
and 0, respectively, for G 3 and G 10 ovulations Table 1 . The overall ratio of OR
Ž .
to TFS tended to favour the F wapiti hybrids 0.87 , with the red deer being lowest
1
Ž . Ž
. 0.67
P - 0.05 , indicating a greater tendency for the stimulation of non-ovulating follicles.
Ž The evaluation of individual hind’s ovulatory responses to treatment over time first
. five consecutive treatments
revealed a highly significant non-random pattern for Ž
. individual red deer
Friedman’s Test T s 6.44, P - 0.01 and F wapiti hybrids
2 1
Ž .
Ž .
T s 4.65, P - 0.01 , but not for 1 4 PD hybrids T s 0.23, NS . For the former two
2 2
genotypes, one or two individuals consistently exhibited the highest ovulation rates. Conversely, one red deer hind failed to exhibit any ovulatory responses throughout the
trial. Analysis of plasma samples collected from hinds during the trial failed to reveal any
immunological reactivity to oFSH at all, with all values falling below the non-specific binding threshold.
4. Discussion
