Butocin

The oestrous cycles of each heifer were synchronised using two 500-mg injections of sodium cloprostenol (EstroPlan; Parnell Laboratories NZ Ltd, Auckland, New Zealand), administered intramuscularly 10-12 days apart. Oestrus was monitored behaviourally and recorded twice daily for 4 days after each prostaglandin injection. The day of oestrus, following the second prostaglandin injection was designated Day 0 of the oxytocin challenge cycle. On Day 16 of this oestrous cycle, 100 IU of synthetic oxytocin (Butocin; Bomac Laboratories Ltd, Auckland, New Zealand) was administered intravenously via a jugular vein immediately after a blood sample was taken (Time 0) from the coccygeal vein. Further blood samples from the tail were taken at 10, 20, 30, 60 and 90 min into evacuated blood tubes (Vaccutainer; Becton & Dickinson, Auckland, New Zealand) containing sodium heparin, placed immediately in iced water and centrifuged within 3 h (15 min at 1500g at 48C) after collection. Plasma samples were aspirated and stored at À208C until analysis. Plasma concentrations of 13,14-dihydro-15ketoprostaglandin F 2a (PGFM) were measured using a radioimmunoassay (Mitchell et al. 1976 (link)). The inter-and intra-assay coefficients of variation were 20.4 and 7.2% (high), 24.1 and 10.3% (medium) and 28.1 and 16.4% (low), respectively. The lower detectable limit of the assay was 6.5 pg mL À1 . Total PGFM release (measured as the area under the curve; AUC) and peak PGFM concentrations were calculated.

Training. Before calving, heifers were allocated randomly to 1 of 2 treatment groups (trained and untrained), balanced for predicted calving date and temperament category (n = 10 per group). Heifers allocated to the trained treatment group then received a training program to the rotary milking parlor over 2 consecutive days, whereas those in the other treatment group were left undisturbed in the paddock. All heifers calved within 9 wk of each other, so the period between training and calving ranged from 1 to 10 wk.
On the first day of training, in the morning (session 1), heifers to be trained were brought into the dairy yards. They were encouraged onto the rotary platform using good stockmanship practices; no loud voices or objects were used to force the animals. If necessary, the stockperson would use a quiet voice and gentle, tactile force with the hands to encourage the heifers to move. Heifers traveled around to the exit point, where a stockperson encouraged them to back off the rotary platform by putting an arm in front of the animals' faces and pushing them backward. Each heifer dismounted from the platform, turned around, and moved down an alley back to the main holding yard. This process was repeated 2 more times. In the afternoon on the first day (session 2), heifers were loaded and exited the rotary platform 3 times using the same procedure as in session 1, but during this session, heifers were also introduced to routine noises and objects such as the radio, cup sprayer, and platform washer. On the second day in the morning (session 3), heifers were loaded and exited the rotary platform 3 times, using the same procedures as in session 2, but during this session, they were also introduced to the sounds of the milking machine, had teat spray applied, and were touched gently high on the escutcheon (above the udder) and back legs. Finally, on the second day in the afternoon (session 4), heifers were loaded and exited the rotary platform 3 times using the same procedure as in session 3, but were exposed to further gentle handling or rubbing of the escutcheon, back of the legs, and top of the udder, progressing until the teats and bottom of the udder had been touched.
First Week of Lactation. Each heifer was brought in for milking within 24 h of calving. During the first 5 d of lactation, heifers were kept together in a separate colostrum herd before being moved into a herd that consisted of primiparous and multiparous cows. Behaviors in the milking parlor were observed and recorded during morning milkings for the first 5 d of lactation. During attachment of the milking cluster, cows were given an overall score based on the performance of flinching, stepping, or kicking (FSK) behaviors using a 4-point scale (1 = no hind foot movement, heifer may flinch, shiver or do nothing at all; 2 = hind leg lifted no higher than 20 cm, step or shuffle of a hind leg; 3 = hind leg lifted higher than 20 cm, step or forward kick of a hind leg; 4 = backward kick of hind leg). The most extreme behavior performed during each milking period was the score given to that animal. The order in which the cows entered the milking parlor was also recorded.
On each of d 1 to 5 of lactation, all heifers were given an oxytocin challenge and residual milk volume was measured. Once heifers finished voluntary milk ejection, the milking cluster was removed, and 2 mL (40 USP units) of oxytocin (Butocin, Bomac Laboratories Ltd., Auckland, New Zealand) was given by intramuscular injection. One minute after injection, the milking cluster was reattached, the residual milk was collected, and its volume recorded. Residual milk as a percentage of total volume was also calculated. Milk samples were collected over the entire milking period during the first 5 d of lactation, and somatic cells were enumerated.
Blood samples (10 mL) were collected from all heifers via the coccygeal vein immediately before and after milking on the first and fifth day of lactation. Blood samples were collected from heifers while they were restrained in a holding area that was within close proximity to the milking parlor. Blood was collected into Vacutainer tubes (BD, Franklin Drive, NJ) containing heparin. Blood samples were centrifuged for 12 min at 1,300 × g at 4°C, and plasma was aspirated and stored at -20°C until analyzed. Cortisol concentrations were measured using a commercially available RIA kit (Siemens Cortisol-A-Count Kit, Cruinn Diagnostics Ltd., Dublin, Ireland).
The research farm used a WestfaliaSurge milking system (GEA Farm Technologies, Cambridge, New Zealand) and the associated DairyPlan C21 herd and parlor management software (GEA Farm Technologies). This was used to record production data at each milking (milk yield, milking duration, and average milking flow rate) during the first 8 mo of lactation.

Training. Before calving, heifers were allocated randomly to 1 of 2 treatment groups (trained and untrained), balanced for predicted calving date and temperament category (n = 10 per group). Heifers allocated to the trained treatment group then received a training program to the rotary milking parlor over 2 consecutive days, whereas those in the other treatment group were left undisturbed in the paddock. All heifers calved within 9 wk of each other, so the period between training and calving ranged from 1 to 10 wk.
On the first day of training, in the morning (session 1), heifers to be trained were brought into the dairy yards. They were encouraged onto the rotary platform using good stockmanship practices; no loud voices or objects were used to force the animals. If necessary, the stockperson would use a quiet voice and gentle, tactile force with the hands to encourage the heifers to move. Heifers traveled around to the exit point, where a stockperson encouraged them to back off the rotary platform by putting an arm in front of the animals' faces and pushing them backward. Each heifer dismounted from the platform, turned around, and moved down an alley back to the main holding yard. This process was repeated 2 more times. In the afternoon on the first day (session 2), heifers were loaded and exited the rotary platform 3 times using the same procedure as in session 1, but during this session, heifers were also introduced to routine noises and objects such as the radio, cup sprayer, and platform washer. On the second day in the morning (session 3), heifers were loaded and exited the rotary platform 3 times, using the same procedures as in session 2, but during this session, they were also introduced to the sounds of the milking machine, had teat spray applied, and were touched gently high on the escutcheon (above the udder) and back legs. Finally, on the second day in the afternoon (session 4), heifers were loaded and exited the rotary platform 3 times using the same procedure as in session 3, but were exposed to further gentle handling or rubbing of the escutcheon, back of the legs, and top of the udder, progressing until the teats and bottom of the udder had been touched.
First Week of Lactation. Each heifer was brought in for milking within 24 h of calving. During the first 5 d of lactation, heifers were kept together in a separate colostrum herd before being moved into a herd that consisted of primiparous and multiparous cows. Behaviors in the milking parlor were observed and recorded during morning milkings for the first 5 d of lactation. During attachment of the milking cluster, cows were given an overall score based on the performance of flinching, stepping, or kicking (FSK) behaviors using a 4-point scale (1 = no hind foot movement, heifer may flinch, shiver or do nothing at all; 2 = hind leg lifted no higher than 20 cm, step or shuffle of a hind leg; 3 = hind leg lifted higher than 20 cm, step or forward kick of a hind leg; 4 = backward kick of hind leg). The most extreme behavior performed during each milking period was the score given to that animal. The order in which the cows entered the milking parlor was also recorded.
On each of d 1 to 5 of lactation, all heifers were given an oxytocin challenge and residual milk volume was measured. Once heifers finished voluntary milk ejection, the milking cluster was removed, and 2 mL (40 USP units) of oxytocin (Butocin, Bomac Laboratories Ltd., Auckland, New Zealand) was given by intramuscular injection. One minute after injection, the milking cluster was reattached, the residual milk was collected, and its volume recorded. Residual milk as a percentage of total volume was also calculated. Milk samples were collected over the entire milking period during the first 5 d of lactation, and somatic cells were enumerated.
Blood samples (10 mL) were collected from all heifers via the coccygeal vein immediately before and after milking on the first and fifth day of lactation. Blood samples were collected from heifers while they were restrained in a holding area that was within close proximity to the milking parlor. Blood was collected into Vacutainer tubes (BD, Franklin Drive, NJ) containing heparin. Blood samples were centrifuged for 12 min at 1,300 × g at 4°C, and plasma was aspirated and stored at -20°C until analyzed. Cortisol concentrations were measured using a commercially available RIA kit (Siemens Cortisol-A-Count Kit, Cruinn Diagnostics Ltd., Dublin, Ireland).
The research farm used a WestfaliaSurge milking system (GEA Farm Technologies, Cambridge, New Zealand) and the associated DairyPlan C21 herd and parlor management software (GEA Farm Technologies). This was used to record production data at each milking (milk yield, milking duration, and average milking flow rate) during the first 8 mo of lactation.

The oestrous cycles of each heifer were synchronised using two 500-mg injections of sodium cloprostenol (EstroPlan; Parnell Laboratories NZ Ltd, Auckland, New Zealand), administered intramuscularly 10-12 days apart. Oestrus was monitored behaviourally and recorded twice daily for 4 days after each prostaglandin injection. The day of oestrus, following the second prostaglandin injection was designated Day 0 of the oxytocin challenge cycle. On Day 16 of this oestrous cycle, 100 IU of synthetic oxytocin (Butocin; Bomac Laboratories Ltd, Auckland, New Zealand) was administered intravenously via a jugular vein immediately after a blood sample was taken (Time 0) from the coccygeal vein. Further blood samples from the tail were taken at 10, 20, 30, 60 and 90 min into evacuated blood tubes (Vaccutainer; Becton & Dickinson, Auckland, New Zealand) containing sodium heparin, placed immediately in iced water and centrifuged within 3 h (15 min at 1500g at 48C) after collection. Plasma samples were aspirated and stored at À208C until analysis. Plasma concentrations of 13,14-dihydro-15ketoprostaglandin F 2a (PGFM) were measured using a radioimmunoassay (Mitchell et al. 1976 (link)). The inter-and intra-assay coefficients of variation were 20.4 and 7.2% (high), 24.1 and 10.3% (medium) and 28.1 and 16.4% (low), respectively. The lower detectable limit of the assay was 6.5 pg mL À1 . Total PGFM release (measured as the area under the curve; AUC) and peak PGFM concentrations were calculated.