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Four trials were conducted to investigate factors controlling the seasonal onset of reproductive activity in red deer hinds. Firstly (Chapter 4), the role of photorefractoriness to long daily photoperiods in the initiation of the seasonal reproductive activity in breeding red deer hinds was examined. Red deer hinds (n=10) were prematurely exposed to a long daily photoperiod of 15.3 h from 22 July to 8 November 1986 i.e. winter-spring (EPW), or maintained under natural photoperiods (NP). Six hinds experienced the natural changes in daily photoperiod until mid-summer but were exposed to a 15.5 h of light each day from 30 January to 30 April 1987, i.e. summer-autumn (EPS), whilst hinds in the other groups experienced naturally decreasing daily photoperiods. On 5 occasions (July 1986, January, February, March and April 1987), blood samples were collected from 4 NP and 4 EPW hinds every 20 minutes for 4 h to monitor secretion of luteinising hormone and half hourly for another 4 h following an i.v. injection of 2 µg GnRH to measure pituitary responsiveness. In January, March and April 1987 EPS hinds were also intensively sampled for 4 h. Plasma progesterone concentrations and mean date of calving indicated that the onset of breeding activity was not affected by light treatment in the EPW hinds but was delayed by 3 weeks in the EPS hinds. In contrast, supplementary lighting caused a premature elevation of plasma prolactin concentrations and advanced pelage moulting in EPW hinds only. Plasma LH secretion patterns indicated that LH pulse frequency and mean LH concentrations were greater during the breeding season than during pregnancy or, seasonal or postpartum, anoestrus. The reduction in LH secretion was partially explained by a diminished pituitary responsiveness to GnRH. Daily plasma melatonin secretion patterns indicated that the duration of the nocturnal increase in melatonin concentrations was responsive to changes in photoperiod and provided a suitable endocrine signal for measuring day length. The results suggest that, unlike the sheep and prepubertal red deer hind, the onset of seasonal breeding activity in breeding red deer hinds is not affected by long daily photoperiods in spring but is delayed if the autumnal decrease in daily photoperiod is delayed. Therefore neither the development of photorefractoriness nor the spring increase in daily photoperiods initiated the transition from seasonal anoestrus to reproductive activity in the breeding hind. However, long daily photoperiods may have entrained the annual cycle of pelage shedding and prolactin secretion. It is possible that the neuroendocrine pathway by which photoperiodic signals entrain the seasonal cycle of reproduction is separate from those which regulate other seasonal events in the breeding red deer hind. Secondly (Chapter 5), in a study of seasonality of reproduction, 4 pubertal hinds were monitored for live weight and plasma LH and progesterone concentrations from December 1987 to October 1988 (i.e. 12-22 months of age). In addition the pattern of LH secretion was also studied in 4 ovariectomised pubertal hinds implanted s.c. with controlled release implants containing 12 mg oestradiol-17β between 4 March and 25 May and between 15 June and 19 September, 1988. On several occasions (15 December, 29 February, 15 March, 24 April, 14 June, 29 June, 18 September (all hinds) and 3 October (ovariectomised hinds only) blood samples were collected every 20 minutes for 4 h to monitor secretion of luteinising hormone and following an i.v. injection of 2 µg GnRH to measure pituitary responsiveness. Plasma progesterone profiles indicated that 4-6 ovarian cycles, lasting about 19 d each, occurred in each intact hind. Regular ovarian cycles commenced in late April (26 April ± 3.4 d, mean ± s.e.m.) and ceased 3 months later in July (21 July ± 7.2 d). The number of LH pulses in the intact hinds was higher in June (1-2 pulses/4 h) than in the non-breeding season (< 1 pulse/4 h) probably due to a seasonal increase in GnRH secretion. It appears that the seasonal increase in LH pulsatility and onset of reproductive activity in the entire hinds were temporally related to a reduction in the sensitivity of LH secretion to the negative feedback effects of oestradiol in ovariectomised pubertal hinds. In the third trial (Chapter 6), 20 male and 20 female red deer calves were immunised at birth against a melatonin conjugate or injected with adjuvant only (controls). Booster injections were given on 5 occasions over the next 2 years. Stags which produced significant melatonin binding activity in response to immunisation, were heavier than the controls between 7 and 11 months of age and at 16, 20, 30 and 34 months of age. A similar but smaller effect on live weight was seen in the immunised hinds. Immunisation against melatonin did not affect the calving date of the hinds or antler development and time of casting of antlers in the stags. These results indicated that disruption of the photoperiodic signal by immunisation against melatonin may have prevented the entrainment of annual rhythms in feed intake and growth but without affecting the seasonal cycles in antler growth and ovarian activity. Finally in a series of 3 trials (Chapter 7), anoestrous red deer hinds were induced to ovulate with the GnRH analogue, buserelin. Hinds were pre-treated with intravaginal devices containing 0.6 g progesterone (CIDR-Type S) for 14 days prior to CIDR withdrawal on 4 March. In Year 1, 15 hinds were treated with 1 CIDR each and 8 hinds were injected i.m. with 4 µg buserelin (a GnRH analogue) at CIDR removal followed by 2nd injection of 10 µg 48 h later. In Year 2, 16 hinds were treated with 2 CIDRs each and 8 hinds were injected Lm. with 4, 3, 2, 2 and 10 µg buserelin at -48, -24, 0, 24 and 48 h respectively from CIDR withdrawal. In Year 3, 15 hinds were treated with 2 CIDRs and 11 hinds (Groups Band BO) injected with buserelin identical to the protocol in Year 2. At CIDR withdrawal 6 of these buserelin treated hinds (Group BO) and 4 other progesterone-primed hinds (Group 0) were injected with 500 µg oestradiol benzoate. In addition, during the breeding season, 4 hinds (Group C) were treated with double CIDRs only for 14 d from 1 April. When oestrous behaviour and the pattern of plasma LH secretion were monitored in Year 3, oestrous behaviour was less noticeable and delayed in Group B hinds and peak LH levels were lower and increased later relative to Group 0, BO and C hinds. Two weeks after CIDR withdrawal, 6 buserelin-treated hinds in both Year 1 and Year 2, and 3/5 B, 2/4 0, 3/6 BO and 4/4 C group hinds in Year 3 had a single corpus luteum present. Plasma progesterone concentrations were elevated for about 12 d in most hinds with a corpus luteum in Year 2 and in most B, BO and C Group hinds in Year 3. However, progesterone secretion was low in several Year 1 and all Group 0 hinds in Year 3 indicating that the induced corpora lutea in these hinds were functionally subnormal. There was no evidence that any of the buserelin-induced ovulations resulted in pregnancy, probably because the induced ovulations were not accompanied by normal hormonal and behavioural patterns. The experiments described in this thesis have contributed to our understanding of the regulation of seasonal breeding in red deer hinds. This knowledge may eventually enable reproductive activity to be effectively manipulated, thereby improving the efficiency of deer production systems.

Two experiments were conducted to examine the interrelationships between body composition at parturition, herbage allowance during lactation and milk production. Detailed measurements of feed intake and changes in body composition allowed estimates of maintenance energy requirement and efficiency of use of energy for milk production. Dorset ewes were offered low (L-) or high (H-) herbage allowances during pregnancy to achieve post-partum body weights, respectively, of about 50 and 60-65 kg. During the first 6 weeks of lactation herbage allowances were approximately 2 (-L), 5 (-M and -Mm) and 8 (-H) kg DM/ewe/d and feed intake was determined using chromic oxide dilution. In the first experiment (Expt.A) ewes were machine-milked and in the second (Expt.B) they were suckled by twin lambs with a further group (-Mm) machine-milked. Samples of ewes were slaughtered during early pregnancy, post-partum, and after 6 weeks of lactation. Body weight differences post-partum between L- and H- groups were 10 and 15 kg in Expts.A and B respectively. During lactation body weight changes appeared erratic, particularly in Expt.B, owing to the effects of variation in gut fill. Daily feed intake of machine-milked ewes on -M and -H allowances reached a maximum (1.9-2.2 kg OM/ewe/d) 2-3 weeks after parturition but ewes rearing lambs on similar allowances showed maximum intake (2.5-2.8 kg OM/ewe/d) during the first week. Mean intakes during lactation were 58, 69 and 73 g DOM/kg W•⁷⁵/d respectively for -L, -M and -H groups machine-milked and 51, 75 and 83 g DOM/kg W•⁷⁵/d respectively for -L, -M and -H groups rearing twin lambs. In both experiments ewes in L- groups had approximately 14% greater mean intake (g DOM/kg W•⁷⁵/d) during lactation compared with those in H- groups. Mean daily milk production of ewes rearing lambs was 2.0, 2.5 and 2.7 kg/d respectively in -L, -M and -H groups and was 33-52% greater than that for machine-milked ewes offered similar herbage allowances. In both experiments L- ewes had 14% greater milk yield (g/kg W•⁷⁵/d) compared with H- ewes. Mean lamb growth rates in Expt.B averaged 209, 254 and 268 g/lamb/d in -L, -M and -H groups, respectively. Lambs in the H- group were approximately 11% heavier at birth than those in the L- group and their mean growth rate was 20 g/d greater. Body fat and energy content of live ewes at the start of lactation was predicted using regression relationships from ewes slaughtered post-partum. There was large variation in the energy content of body weight loss during lactation (-37 to +140 MJ/kg) owing to variation in weight of gut fill and changes in chemical composition of the empty body. In Expt.A the body fat content of ewes during early pregnancy was 16.9 kg. Predicted values post-partum were 10.0 and 15.7 kg in L- and H- groups, respectively. During lactation all groups showed body fat losses which ranged from 53 (LH) to 120 g/d (HL). Body fat content during early pregnancy in Expt.B was slightly greater (19.0 kg) than in Expt.A and predicted values post-partum were 11.5 (L-) and 19.7 kg (H-). Fat mobilization during lactation was greater than in Expt.A, and ranged from 157 (LH) to 287 g/d (HL). In both experiments there were losses in body protein during pregnancy in L- ewes (5 and 15 g/d in Expts.A and B respectively). During lactation, protein losses were most evident in H- ewes, being greatest in HL groups (26 and 43 g/d in Expts.A and B respectively). Water: protein ratio in the empty body showed a progressive increase during pregnancy and lactation in both experiments. Estimated maintenance energy requirement for machine-milked ewes tended to be greater for H- (.236 MJ ME/kg W/d) compared with L- ewes (.205 MJ ME/kg W/d) but was similar for both groups of ewe rearing lambs (average of.238 MJ ME/kg W/d). Efficiency of use of ME above maintenance for milk production (K₁) in L- and H- groups was, respectively, .69 and .95 in machine-milked, and .69 and .64 in suckled ewes. Energy from mobilized body tissues was utilized for milk production with respective efficiencies in L- and H- ewes of .23 and .35 in machine-milked, and .40 and .50 in suckled ewes. Efficiency of conversion of total energy available (i.e. ME above maintenance and mobilized tissue energy) to milk energy (K₁(t)) when compared among all groups in both experiments, tended to decrease (from .84 to .51) with increasing body energy mobilization. There was a positive relationship, however, between K₁(t) and the proportion of mobilized energy derived from body protein. Metabolizable energy intakes in the present experiments appeared to be 10-15% greater than calculated requirements from ARC (1980). Estimates of ME requirements for lactating ewes at pasture, in relation to body weight, level of milk production and body energy change, have been calculated based on the present data.

A series of eight experiments was conducted to investigate the requirement for thyroid hormones in neuroendocrine processes which lead to the seasonally anoestrous state in red deer hinds. The first two experiments used thyroidectomized, ovariectomized, oestradiol-treated hinds which received various thyroid hormone replacement treatments (n=5 per group) to investigate the timing and dose-responsiveness of thyroid hormones in bringing about seasonal oestradiol-induced suppression of plasma LH concentration. A significant seasonal decline in mean plasma LH concentration during September (coinciding with the onset of anoestrus in entire cycling hinds in New Zealand) was observed in all thyroidectomized hinds in both experiments regardless of T₄ or T₃ treatment. When oestradiol implants were removed in November or December, mean plasma LH concentrations increased significantly in all but one of hinds in which T₄ had been administered at very low doses by subcutaneous implants, and mean plasma LH concentrations and LH pulse amplitude increased in approximately half of hinds administered T₃ at varying doses by subcutaneous injections over a one-week period in October. These results suggested that thyroid hormones are not required for steroid-dependent reproductive suppression, but could possibly play a role in steroid-independent suppression of LH secretion. Because problems were encountered in delivering appropriate doses of thyroid hormones in both experiments, further confirmation of these findings was required. Therefore in the next experiment the role of thyroid gland secretions was examined in euthyroid (n=5) and thyroidectomized (n=4) ovariectomized hinds treated with oestradiol implants. These implants were removed for about one month on three occasions to examine the effect of thyroidectomy on steroid-independent control of seasonal LH secretion. During the non-breeding season basal and GnRH-induced plasma LH concentrations declined in all hinds in the presence of oestradiol, but returned to breeding season levels when oestradiol was withdrawn in November. In a concurrent experiment, thyroidectomy of ovary-entire hinds (n=7) during the breeding season prevented the cessation of oestrous cyclicity in spring; this was in contrast to oestrous cyclicity in T₄replaced (n=4) or euthyroid control (n=5) hinds which ceased to occur in early September. Collectively, these results indicate that thyroid hormones are required for the termination of the breeding season in cycling red deer hinds and that this action occurs via steroid-independent neuroendocrine pathways. Two experiments were conducted using neurotransmitter receptor agonists and antagonists to identify neural pathways in the brain which mediate LH suppression by oestradiol and by steroid-independent mechanisms, and to test if the thyroid gland is required for activation of these pathways during the non-breeding season. It was concluded from the lack of plasma LH responses to dopaminergic and opioidergic agonists and antagonists in ovariectomized and ovariectomized, thyroidectomized hinds (n=5) that neural pathways involving dopamine-D₂receptors do not mediate oestradiol-induced seasonal suppression of plasma LH concentrations, and neither dopaminergic or opioid neural pathways mediate non-steroidal suppression of plasma LH concentrations. However preliminary evidence was obtained for a stimulatory role of serotonergic neural pathways in controlling LH secretion. Another experiment was conducted to identify when the steroid-independent mechanisms which suppress LH concentrations during the non-breeding season are responsive to thyroid hormones. T₄treatment at the beginning of or during the non breeding season was effective in bringing about suppression of plasma LH concentration in thyroidectomized, ovariectomized hinds (n=5 per group), but this action of thyroid hormones did not occur during the breeding season. These results show that the steroid-independent mechanisms which contribute to seasonal suppression of plasma gonadotrophin concentrations require thyroid hormones to be present only from around the time of the end of the breeding season for their normal expression, and they remain responsive to thyroid hormones after this period. Lastly, the feasibility of achieving out-of-season breeding using thyroidectomized hinds (n=9) was evaluated by comparing oestrous behaviour, ovulation and pregnancy rates to those of euthyroid control hinds (n=7) following synchronization of oestrous cycles. There was a non-significant trend for a greater occurrence of oestrous behaviour and ovulation in thyroidectomized hinds compared with euthyroid controls during the non-breeding season, but the pregnancy rate following out-of season mating with a thyroidectomized stag was low, suggesting that a side effect of thyroidectomy may be impaired fertility. Six out-of-season pregnancies were obtained from eight matings, however because three of these pregnancies occurred in euthyroid control hinds no improvement in out-of-season reproductive performance could be attributed to thyroidectomy. It is likely that if the actions of the thyroid glands are to be exploited as a tool for achieving out-of-season breeding in this species, techniques will have to be developed for specifically blocking or overcoming the effects of thyroid hormones on the reproductive neuroendocrine centres without causing general hypothyroidism and its associated side-effects.