The effect of environment on behaviour, plasma cortisol and prolactin in parturient sows

APPLIED ANIMAL BEHAVIOUR

SCIENCE

ELSEV I ER Applied Animal Behaviour Science 39 (1994) 313-330

The effect of environment on behaviour, plasma cortisol and prolactin in parturient sows

A.B. L a w r e n c e *,a, J .C. P e t h e r i c k b, K .A. M c L e a n a, L .A. D e a n s a, J. C h i r n s i d e a, A. V a u g h a n ~, E. C l u t t o n ¢, E .M.C . T e r l o u w d

"Genetics and Behavioural Sciences Department, SAC Edinburgh, West Mains Road, Edinburgh EH9 3JG, UK

bThe Roslin Institute, AFRC Edinburgh, Roslin EH25 9PS, UK CDepartment of Veterinary Clinical Studies, Royal (Dick) School of Veterinary Studies, The University

of Edinburgh, Veterinary Field Station, Easter Bush, Roslin EH25 9RG, UK dlNRA, Institute de Recherches sur la Viande, Centre de Clermont-Ferrand - Their,

63122 St-G enes-Champanelle, France

(Accepted 1 October 1993)

Abstract

Nest-building in the pig is thought to be stimulated by a pre-parturient surge in prolactin. There is concern that sows in crates may experience psychological stress as a result of physical interference with nest-building.

Thirty-three gilts were implanted with jugular catheters approximately 10-14 days before expected date of parturition (EPD). On day 5 before EPD, gilts were moved into either conventional farrowing crates without bedding (treatment C; n = 16) or pens allow- ing freedom of movement with bedding (treatment P; n = 17). The animals were blood sampled on the day before and the day after introduction to the treatments. Blood and behaviour sampling was resumed 48 h before EPD and continued until 4 h post-commencement of farrowing.

As with previous studies gilts in both environments were more active (P<0.001) and performed more substrate-directed behaviour in the pre-parturient period (P< 0.001 ). Gilts in pens spent considerable amounts of time in straw-directed behaviour, and gilts in crates increased amounts of floor- and fixture-directed behaviour (both P<0.001 ). Prolactin was not affected by treatment and increased in both environments over the same time period as this substrate-directed behaviour. However, the causal role of prolactin in nest- building must now be questioned given that a number of individuals showed no increase in prolactin over the 48 h before EPD, but still showed an increase in substrate-directed activity. Cortisol was strongly affected by treatment with treatment C having elevated total cortisol over much of the pre-parturient period (P<0.001). Cortisol also increased in

*Corresponding author.

0168-1591/94/$07.00 © 1994 Elsevier Science B.V. All fights reserved SSDI0168-1591 (93)01010-4

314 A.B. Lawrence et al. / Appl. Anim. Behav. Sci. 39 (1994) 313-330

penned animals during parturition. There were no consistent correlations among behaviour, prolactin and cortisol.

The general increase in cortisol in the early post-partum period may suggest that parturition in itself has stress-inducing aspects. The additional rise in cortisol found in the crated sows may indicate that the close confinement of the crate, by interfering with the expression of maternal behaviour, induces psychological stress. There was no evidence that the elevated cortisol response in crated gilts extended beyond the end of parturition.

Key words: Pig; Endocrinology; Parturition; Housing

1. Introduction

Many species of animal show increased activity and nest-building in the pre- parturient period. A number of studies have shown that when given the opportunity wild boar and domestic sows (Sus scrofa) isolate themselves from the so- cial group as parturition approaches, select a nest-site and then construct a nest of some complexity in which they give birth (Stolba and Wood-Gush, 1984; Gun- dlach, 1968; Jensen, 1986). Nest-building is often regarded as a behaviour sub- ject to strong physiological control (e.g. Widowski and Curtis, 1990), and in mice prolactin is thought to play a central role in motivating nest-building activities. In multiparous sows prolactin is reported to increase sharply on the day before farrowing (Dusza and Krymowska, 1981; Vale and Wagner, 1981 ), and Taverne et al. (1979) suggested a strong temporal relationship between the peak of the prolactin surge and the onset of nest-building. More recently Castren et al. (1994) suggested that it is the pre-parturient rise in prolactin that initiates the behaviour.

To our knowledge there have been no reports of the effect of environment on the physiology underlying nest-building. The domestic pig represents a good model for such studies; welfare and production concerns over the widespread use ofbe- haviourally restrictive farrowing crates in pig farming, has led to investigations of the influence of close confinement on the expression of sows' pre-parturient behaviour. Sows in farrowing crates have been reported to show increases in activity and substrate-directed behaviour as parturition approaches, similar to less restricted animals (e.g. Vestergaard and Hansen, 1984; Lammers and De Lange, 1986; Heckt et al., 1988 ). Such behavioural changes in crated sows at parturition are often assumed to represent nest-building activity (e.g. Cronin et al., 1993 ).

It has been suggested that farrowing crates, by interfering with sows' pre-parturient behaviour, may be stress-inducing (Baxter, 1982; Vestergaard and Han- sen, 1984) and that this stress could lead to prolonged parturitions (e.g. Vester- gaard and Hansen, 1984). Lawrence et al. ( 1992 ) recently showed that an acute behavioural stress in mid-partum can induce an opioid-mediated inhibition of oxytocin thereby prolonging parturition. Cronin et al. ( 1991 ) found little evidence that sows housed in farrowing crates around parturition and lactation experienced elevated plasma free cortisol (a commonly used index of behavioural

A.B. Lawrence et al. / Appl. A nim. Behav. Sci. 39 (1994) 313-330 315

stress; Dantzer and Mormede, 1983), but took no measurements over the immediate pre-farrowing period.

In this study we measured the effect of environment on peri-parturient behaviour, on prolactin concentrations as a likely internal stimulus for nest-building and on total cortisol as an index of behavioural stress. Our first objective was to confirm the correlational relationship between prolactin and nest-building, and to test whether physical interference with nest-building affects prolactin concentrations. Second, we tested the hypothesis that physical interference with nest- building is stress-inducing, thereby resulting in elevated plasma cortisol.

2. Animals, materials and methods

2.1. Animals and housing

The subjects were 40 Large White × Landrace primiparous females (gilts; Cot- swold Pig Development Co., Lincoln, UK). The gilts were purchased in four consecutive groups (n = 10) over an 18 month period, and housed in groups in a strawed yard (9.6 m × 6 m) for 90.0_+6.7 days (s.e.m.), where they were floor- fed 2 kg day- ~ of a commercial diet ( 13 MJ (DE) kg- ~ ). The gilts were served by a boar on 2 consecutive days, on average 51.0 _+ 2.1 days after being housed in the pens. The expected parturition date (EPD) was calculated to be 114 days after the first service date. The pens were cleaned weekly and provided with fresh straw for bedding twice a week.

At 79.0-+ 2.0 days before EPD the groups were moved to a strawed yard (9.6 m × 3 m) and fed individually through an electronic sow feeder (Collinson and Co., Lancashire, UK). The feeder was programmed to be open between 08:00 and 24:00 h, and the same commercial ration as above was fed at a rate 2.2-2.6 kg day- 1 per gilt, depending on the individual's weight and body condition. The yard was cleaned and fresh straw provided as above.

2.2. Catheterization

At approximately 14.5_+0.3 days before EPD, all the subjects had a jugular catheter (silastic tubing; Dow Coming Europe, Belgium; internal diameter 1.3 mm; external diameter 2.5 mm) implanted under general anaesthesia (for details of the procedure see Lawrence et al., 1992 ). In brief, the animals were induced to anaesthesia with ketamine (Vetelar: Parke Davis, Pontypool, UK), added to a 5% guaiphenesin solution (J.M. Loveridge, Southampton, UK), following an in- tramuscular injection of azaperone (Stresnil: Janssen Pharmaceutical, Oxford- shire, UK; dose 1 mg kg- 1 body-weight). Anaesthesia was maintained with halothane (Halothane: R.M.B. Animal Health, Dagenham, UK) delivered in oxygen via a semi-closed circle breathing system. A 23 cm length of the catheter was inserted into the vein and the remainder of the catheter, previously strengthened, was exteriorized via the ventral entry wound in the neck. The catheter was then

316 A.B. Lawrence et al. /Appl. Anim. Behav. Sci. 39 (1994) 313-330

protected within an adhesive neck bandage. Access was by a connecting tap at the back of the neck. The catheters were flushed daily with saline until the sampling procedure began. After flushing, the catheters were primed with saline solution containing 150 I.U. ml- 1 of heparin (Fisons, C.P. Pharmaceuticals, Wrexham, UK) and 5000 I.U. ml-1 of sodium benzylpenicillin (Crystapen: Pitman-Moore (UK), Middlesex, UK. ).

After catheterization, the gilts were moved to a room containing ten straw- bedded pens (2 m × 2 m) where they were individually housed. After full recov- ery from the operation, the food level was increased to 3 kg day- 1 using the same commercial ration as before. Seven gilts developed occluded catheters and were withdrawn from the study.

2.3. Experimental housing

Five days before EPD, the remaining 33 gilts were weighed (average weight: 186.5 _+ 2.3 kg) and moved to the parturition accommodation. Individuals were randomly allocated to either a crate (treatment C; 2 m in length, 0.54 m in width and 1.0 m in height) with a solid floor to the front and slats behind and no bedding, or a pen (treatment P; 2.1 m×3.1 m) again with a solid floor to the front and slats behind, but containing straw bedding. House temperature was controlled (average maximum temperature: 19.8_0.3°C; average minimum: 16.6 ___ 0.3 ° C) and the main lights over a central corridor were on between 08: 00 and 16:30 h. Other lights over side corridors were on throughout the 24 h period to permit observation of animals. The animals were offered two meals of 3 kg each of a food containing 13.75 MJ (DE) kg -1 and 18% protein, at 08:00 and 16: 00 h, and the pens were cleaned and provided with fresh straw after the morn- ing feed between 08:15 and 08: 30 h. The slats at the rear of the crates were also cleaned at this time when necessary.

2.4. Blood sampling

Blood samples were taken throughout the experimental period at intervals de- termined by the expected proximity of parturition (Table 1 ). The samples were taken at 4 h intervals for the 24 h before introduction to the parturition environ- merit (day Pre/In), on the second part of the day of introduction (Post/Inb), between 48 h and approximately 24 h before EPD, and on days l, 2 and 7 following parturition (days F1, F2 and F7; see Table 1 for a definition of these different sample periods). At other times blood sampling was more rapid, at either 15 or 30 min intervals (Table 1 ).

It was intended to blood sample every 30 min from approximately 24 h before the commencement of parturition. In practice, the more frequent blood sampling commenced when there were clear behavioural and/or physical indications (e.g. increased activity; straw- or floor-directed behaviour; secretion of milk; oede- matous udder) that parturition was relatively imminent.

Collection of blood at 4 h intervals was via the neck tap and the catheters were

A.B. Lawrence et al. / Appl. Anim. Behav. Sci. 39 (1994) 313-330

Table 1 Data collection protocol

317

Period Time (h) between samples

Blood Behaviour

Pre/In (08:00-08:00) Post/In a ( 10: 00-11 : 00)

(11:00-12:00) Post/In b ( 12: 00-04: 00) 48 h before EPD 24 h before EPD Farrowing 0-2 h 2-4 h Placenta-3 h Day F 1 (08: 00-04: 00 ) Day F2 (08: 00-04: 00) Day F7 (08: 00-04: 00)

4 0.25 0.5 4 4 0.5

0.25 0.5 0.5 4 4 4

0.25 0.25 4 4 0.5

0.25 0.25 0.5 4 4 4

Pre/In, day prior to introduction into the farrowing environment. Post/In, day of introduction. Day F1, First full day following farrowing. Day F2, Second full day of following farrowing. Day F7, 7 days post-farrowing.

then flushed with saline and filled with a heparin solution ( 150 I.U. ml- 1 ). More frequent sampling involved fitting a silastic extension tube to the neck tap which allowed samples to be taken remotely with minimal disturbance to the gilt. After collection, the catheters and extension tubes were again flushed with saline and primed with a heparin solution (75 I.U. ml-1 ). Each sample comprised 10 ml of blood collected in an heparinized luer monovette tube (Sarstedt, Leicester, UK). Subsequent storage, centrifuging and pipetting was at 4 ° C. Plasma for future assay was stored at - 20°C.

2.5. Behavioural observations

Behavioural observations were recorded using a data collection system (Key- behaviour: Deag, 1990) mounted on an Atari Portfolio Personal Computer (Atari Corporation, Sunnyvale, CA, USA). Continuous (focal) observations of individuals' behaviours were made throughout the experimental period, with the timing of observations correlating closely with that of the blood sampling protocol (see above and Table 1 ). Behavioural samples were for 5 min duration when the interval between samples was 4 h or 30 rain. When sampling was more frequent, the continuous recording period was reduced to 3 min. Each behaviour sample recorded changes in the posture and activity of an individual, together with the identification of any substrate used (see Table 2 for details of the behavioural categories used when sampling).


Table 2 Posturai and behaviourai categories used

Postures

Stand Ventrallying Laterallying in between

Behaviours Alert Inactive Move Drink

Straw-directed activity } Floor-directed activity Wall-directed activity Bars-directed activity

Fully upright position Lying on belly with legs tucked under body and udder concealed Lying on side with legs to one side and udder exposed Moving between standing and lying position

Eyes open and individual apparently aware of environment Eyes shut and individual apparently sleeping Stepping movements (only when standing) Drinker manipulation and ingestion of water

Rooting, chewing, licking, biting nosing, pawing at substrate

2.6. Piglets

The birth time of each piglet was recorded as the time when its whole body had been ejected from the gilt. The piglets remained with the gilt until the umbilical cord had been naturally severed. They were then removed, weighed and put into a warm creep area until 2 h after the placenta had been expelled. To aid survival, the piglets were given a colostrum substitute (Littermaker: Fisons Animal Health, Loughborough, UK) within the first 3 h of birth.

2. 7. Hormonal analysis

All blood samples were assayed in duplicate and samples from different treatments and groups were balanced across assay runs.

2. 7.1. Prolactin Plasma prolactin concentrations were measured using a radioimmunoassay

technique based on the method described by McNeilly and Andrews (1974). Modifications to the technique included the use of NIH porcine prolactin for the production of 125I-labelled porcine prolactin, anti-porcine prolactin antibody and standards made by double dilution (concentrations 0.4-200 #1). The assay sensitivity was 1.15 ng ml- 1 and intra- and interassay coefficients of variation were 9.97% and 10.20%, respectively.

A.B. Lawrence et al. / Appl. Anim. Behav. Sci. 39 (1994) 313-330 319

2. 7.2. Cortisol Cortisol was extracted from 50 gl of plasma and concentrations were measured

using radioimmunoassay of the extracted steroid (Duncan et al., 1990). The assay sensitivity was 0.17 ng ml- t and intra- and interassay coefficients of variation were 10.99% and 9.86%, respectively.

2.8. Statistical Analysis

The proportion of time spent in the different behavioural categories at each sample was calculated using Keytime (Deag, 1990). These sample values for each gilt were then averaged for the following time periods: - 48 to - 24 ( - 4 8 / - 24), - 2 4 / - 1 2 , - 1 2 / - 6 , - 6 / - 3 , - 3 / - 0 . 5 h before parturition, 0 (expulsion of first piglet is the start of parturition)/0.75, 1/1.75, 2/4 h after the start of parturition.

The data were normalized by log l° transformation and analyzed using a re- peated measures analysis of variance (ANOVA; Genstat version 5 ), with nested structures for pig and time, and factors treatment (two levels), time (eight levels) and group (four levels). Post-hoc comparison of means was made by the least significance difference (LSD) test. The same procedure was used to analyze the effects of treatment, time and group on rate of standing and moving.

The data for circulating levels of cortisol and prolactin were averaged for the following additional time periods to those used above: Pre/In, Post/In a (first 2 h of the day of introduction), Post/In b, and days F1, F2 and F7. The hormonal data were also normalized by log 1° transformation and analyzed by ANOVA with the same model as used in the behavioural analysis above. Days Pre/In and Post/ In were analyzed separately; days F1 to F7 were analyzed with the data from the peri-parturient phases.

To discriminate animals that showed no rise in prolactin over the pre-parturition period, individual's prolactin values between - 4 8 h and parturition, and - 24 h and parturition were regressed against time. Only those animals with samples taken over the entire 48 h or 24 h before parturition were considered. Ani- mals were classified as having exhibited no rise in prolactin between - 4 8 and -0 .5 h, and between - 2 4 and -0 .5 h, if the slope of the regression was not significant at P< 0.05.

Relationships among behaviour, cortisol and prolactin were analyzed by Pear- son product-moment correlations using the log-transformed data.

3 . R e s u l t s

3.1. Gestation length, litter size and weight

Parturition environment had no overall effect on gestation length (average gestation length (days): 114.19_+0.36 (s.e.m.) vs 114.65 +0.19 for treatment C vs treatment P, respectively). There were also no overall treatment differences in


the total born alive or the average birth-weight of piglets (average litter size: 10.8 + 0.6 vs 10.1 ___ 0.5 for treatment C vs treatment P, respectively; average piglet birth-weight: 1.29 + 0.04 kg vs 1.26 + 0.05 kg for treatment C vs treatment P, respectively).

3.2. Behaviour

There were large differences in behaviour between the pre-parturient phase and parturition itself. Gilts in the periods before parturition stood for longer (proportion of time standing: 0.46 +_ 0.06 vs 0. l 1 _+ 0.03 for time periods - 1 2 / - 6 vs 1/1.75 h(F7,2~4=47.79, P<0.001 )) , and had higher rates of standing (rate of standing: 0 .17+0.03 vs 0.06+0.01 for the same periods ( F 7 , 1 7 2 = 1 7 . 5 1 , P < 0.001 ) ). Time spent in lateral lying was also substantially increased after the start of parturition (proportion of time spent lateral lying: 0.24___0.06 vs 0 .70+0.07 again for the same periods (F7.214= 37.87, P<0.001 )) . Gilts before parturition were also less inactive (proportion time spent inactive: 0.18 + 0.05 vs 0.51 +0.07 for periods - 1 2 / - 6 h vs 1/1.75 (F7,2~4=24.15, P<0.001 )) , spend- ing considerable periods of time in behaviour directed at available substrates (proportion of time in substrate-directed behaviour: 0 .26_ 0.04 vs 0 .07_ 0.02 for the same periods (F7,2~4 = 17.34, P < 0.001 ) ).

Parturition environment was also found to affect a number of behavioural cat-

Table 3a Mean ( _+ se) proportion of time spent in different postures by pre-parturient gilts in crates ( C; n = 16 ) and pens (P; n = 17)

Posture Treatment Time period relative to commencement of parturition (0 h)

- 4 8 / - 2 4 - 2 4 / - 1 2 - 1 2 / - 6 - 6 / - 3 - 3 / - 0 . 5

Stand C 0.49 a 0.37 a,b,x 0.34 b 0.33 b 0.10 c

+0.063 +0.048 +0.068 +0.063 +0.024 P 0.43 a 0.47 a'b'y 0.58 b 0.36 a 0.12 ~

+0.040 +0.057 +0.060 +0.051 +0.033 Lateral C 0.35 ~ 0.26 a 0.23 ~ 0.26 a'x 0.56 b'x

lying _+0.058 +0.046 +0.063 +0.052 +0.074 P 0.46 ~ 0.41 a 0.26 b 0.43,,y 0.76=,y

_+0.042 +0.059 _+0.054 _+0.067 +0.043 Ventral C 0.08 a 0.20 a'b 0.26 b'x 0.26 b 0.22 b'x

lying -+0.025 +0.044 _+0.061 +0.056 +0,056 P 0.09 ~ 0.10 a 0.14 a'y 0.16" 0.07 a'y

_+0.021 +0.027 +0.022 +0.033 +0.022 In between C 0.00" 0.01 ~ 0.03 ~ 0.04 b'x 0.02 ~ standing and + 0.003 + 0.004 + 0.008 + 0.012 _+ 0.008 lying

P 0.01 a 0.01 a 0.02 a 0.01 a,y 0.01 a +0.003 +0,004 +0.008 +0.002 +0.001

Comparisons between time periods with different superscripts "-c or between treaments x-y are significantly different at P < 0.05.

A.B. Lawrence et al. / Appl. Anim. Behav. Sci. 39 (1994,) 313-330 321

Table 3b

Mean ( _+ se) proportion of time spent in different behavioural activities by pre-parturient gilts in crates (C; n = 16) and pens (P; n = 17)

Behaviour Treatment Time period relative to commencement of parturition (0 h )

- 4 8 / - 2 4 - 2 4 / - 1 2 - 1 2 / - 6 - 6 / - 3 - 3 / - 0 . 5

Inactive C 0.38 a'c 0.31 a'b 0.17 b 0.19 b,x 0.48 c,x

+0 ,046 +0 .056 +0 .062 +0 .053 +0 .068 P 0.47 a 0.37 ~ 0.19 b 0.38 a,y 0.71c,y

+0 ,045 +0 .062 +0 .047 +0 .054 +0.041 Alert C 0.11 a,c 0.27 b,~ 0.31 b,x 0.16 a 0.04 a,c

+0.021 +0 .059 +0 .056 +0 .039 +0 .013

P 0.08 a 0.10 a,y 0.12 a,y 0.09 a 0.03 a

+0 ,017 +0 .020 _+0.025 +0 .026 +0 .009 Straw-directed C . . . . .

behav iour . . . . .

P 0.02 ~ 0.10 b 0.27 ~ 0.18 d 0.07 b

+ 0.009 + 0.026 + 0.037 + 0.026 + 0.022 Floor-directed C 0.01 a 0.03 a,b,x 0.07 b,x 0.14 ~,x 0.08 b,~

behaviour + 0.004 + 0.011 + 0.023 + 0.047 + 0.042

P 0.01 a 0.0I a,y 0.02 a,y 0.01 a-u 0.01 a,y

__.0.007 +0 .005 +0 .005 __.0.005 +0.001 Fixture-directed C 0.01" 0.02 a 0.05 b,x 0.09 b,x 0.06 b,x

behaviour + 0.005 _+ 0.005 + 0.072 + 0.024 _+ 0.034

P 0.00 a 0.00 ~ 0.01 a,y 0.00 a'y 0.00 a'y

+0.001 _+0.002 +0 .006 +0 .003 _0 .001

Comparisons between time periods with different superscripts "-a or between treatments x-y are significantly different at P < 0.05.

egories in the pre-parturient period (Tables 3a and 3b). Gilts in treatment P spent more time standing in the pre-parturient phase, particularly at period - 1 2 / - 6 h before parturition (Table 3a; effect of treatment on time standing: Fl,31 = 5.94, P < 0.05 ). In contrast, treatment C gilts had higher rates of standing during the same time period (rate of standing: 0.20 _+ 0.03 vs 0.13 _+ 0.02 for treatment C vs treatment P at period - 1 2 / - 6 : time×treatment interaction: F 7 , 1 7 2 - - 2 . 3 7 ,

P < 0.05 ). Treatment P gilts also showed more lateral lying and treatment C more ventral lying during the pre-parturient phase (Table 3a; effect of treatment on lateral lying: F~,31 = 4.5 l, P < 0.05; time X treatment interaction on ventral lying: F7,2~4 = 2.16, P < 0.05 ). Furthermore, treatment C gilts spent more time in between standing and lying at period - 6 / - 3 (Table 3a; time X treatment interaction: F7,214= 2.89, P < 0.01 ).

In terms of specific behavioural activities, gilts on treatment P spent considerable amounts of time in straw-directed behaviour, which reached a peak at - 12/ - 6 h before parturition, declining thereafter (Table 3b; time effect on straw- directed behaviour: F7,2x4 = 14.24, P < 0.001 ). For much of the same time period, treatment C showed increased amounts of floor- and fixture-directed behaviour, the treatment difference for both activities persisting up until period - 3 / - 0.5


(Table 3b; t ime×trea tment interactions on floor- and fixture-directed behaviour: F7,214=4.95 and F6,183=4.62, respectively; both P<0.001 ). Over much of the time that treatment C gilts were directing behaviour towards the floor and bars, they were also scored as being more alert than P gilts (Table 3b; time × treatment interaction on time alert: F7,214 = 4.12, P < 0.001 ). In contrast as parturition approached, treatment P were increasingly scored as inactive relative to treatment C (Table 3b; treatment effect on time inactive: Fi,31 ----- 5.77, P< 0.05 ).

Not all behavioural categories were affected by treatment. Gilts on both treatments increased time spent in movement and drinking during the pre-parturient phase to the same extent (proportion of time spent moving [pooled data for both treatments ]: 0.009 +_ 0.004 vs 0.044 _ 0.017; proportion of time spent drinking: 0.012 _ 0.003 vs 0.325 + 0.010 both for periods - 4 8 / - 24 vs - 1 2 / - 6; time effect on moving and drinking: F7,214=2 .57 , P<0.05; F6,183=7 .02 , P<0.001, respectively).

3.3. Prolactin

There was no difference in prolactin concentrations between treatments on day Pre/ In (Fig. 1 ). Prolactin was elevated on day Post/In a, but there was no treatment difference and concentrations had returned to the Pre/ In baseline by Post/ In b (overall time effect on prolactin: F2,50 = 6.57, P < 0.01 ).

Prolactin was strongly affected by parturition (Fig 2; overall effect of time on pro lac t in : F10,242 = 66.65, P < 0.001 ). There was a strong rise in concentrations at period - 4 8 / - 24 relative to Pre/ In (paired t-test: t3o = 17.77, P < 0.001 ). There were further significant rises between periods - 4 8 / - 24 and - 2 4 / - 12, and between - 2 4 / - 12 and - 1 2 / - 6, and significant decreases between periods 2/4 and F1, F1 and F2, and F2 and F7 (all LSD: P < 0.05 ). However, not all individuals showed increases in prolactin over the pre-parturient period. Nineteen and 25 individuals had complete data sets from - 4 8 h to parturition, and from - 2 4 h to parturition, respectively. Of these individuals, six (two in treatment C; four in treatment P) were classified (see Statistical methods), as having no rise prolactin from - 4 8 h to parturition, and 13 (six in treatment C; seven in treatment P) as having no rise from - 2 4 h to parturition (Fig. 3 ). Treatment had no effect on prolactin concentrations (F~,24=0.40, ns; Fig. 2), and neither was there a time × treatment interaction (F1o,242 = 0.95, ns ).

3.4. Cortisol

Circulating plasma cortisol concentrations did not differ between treatments on day Pre/In, but were elevated in treatment C on day Post /In a, after introduction to the parturition environment (Fig. 1 ). This small, but significant effect of treatment on cortisol was sustained over Post /In b (Fig. 1; overall effect of treatment on cortisol on days Pre/ In and Post/In: F1,24=15.49, P<0.001; time × treatment interaction: F2,so = 6.04, P < 0.001 ).

Treatment had a strong effect on cortisol concentrations with treatment C gilts

A.B. Lawrence et al. /AppL Anim. Behav. Sci. 39 (1994) 313-330 323

lo Prolactin (ng/ml)

8

4

2

0 Pre/In POSt/It 8 POSt/In b

Time period

Cortisol (ng/ml) 4 0

2O

10

0 Pre/In Post/ In a Post/ In b

Time period

Fig. 1. Average concentrations (with s.e.m. ) ofprolactin and cortisol on the day before (Pre/In) and the day after (Post / In) introduction of gilts to crates (II; n = 16 ) or pens ( [3; n = 17 ). Post/In" is the average concentration for the first 2 h of Post/In and Post/In b the average for the remainder of Post / In.

40 ProlacUn (nglml) farrowing

3O

2O

10

0 P/In - 4 8 - 2 4 - 1 2 - 6 - 3 0

Time period

t F1 F2 F7

Fig. 2. Average concentrations (with s.e.m. ) of prolactin over the peri-parturient period for gilts in crates ( I ; n = 16 ) or pens ( I-1; n = 17 ). Pre/In (P / In ) values are shown for comparison. Time period - 4 8 refers to period - 4 8 / - 2 4 , period - 2 4 refers to - 2 4 / - 12 etc.


a)

50 :'rolectin (ng/ml) Prop. of tim~ loo

/

40 0 .~0

30 0.60

20 ~ 0.40

10 ~ 0.20

. . O . . . . . . . " . . . . . . . . . . . . . . . . . " " ' - ' - ° ' " ' ~ ' * 0 0 0

-48 -42 -36 -30 -24 -18 -12 -6

b) 1.00

0.80

0.60

0.40

. . . . . . . . . • t ~'~ "'" '1 0.00 -48 -42 -36 -90 -24 - 18 - I ~ -6 0

e) 50 1.00

40 0.SO

30 06O

:".. ~4 .": 20 ~ 0.40

lO , " . . . . 0.20 ,.,....)... ili / , , i ! ! ~. J J ,i ,.

-48 42 -36 -30 - 24 - 1 8 -12 -6 0.00

d) 50 1 O0

40 0,80

30 ~ 0.60

2O 04O

10 - 0 2 0

. . . . . . . . . . . . . . • . . . . . . . . . . . .

-48 -42 36 -30 -24 18 - 12 -6 0.00

Time (hre)

50 Prolacti . (ng/ml) Prop. of tim(

40 I-

3 O ~

- 4 8 - 4 2 - 3 6 - 3 0 - 2 4 - 1 8 - 1 2 - 6 0

100

0.80

0.60

0.40

0,20

3.00

40

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~o ig

10

oL . . . . . . / . "... . . . . . - 48 - 4 2 - 3 6 - 3 0 -24 - l a - 12 -6

1 O0

0 8 0

0 60

0.40

0.20

0.00

z~O

30 .~

~o ii il

100

0.80

060

OAO

0.20

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~o

~0

!O

- 48 42 -36 -30 -24 18 -12 -6

Time (IYs)

tOO

O8O

0.6o

o.4o

0.2o

0.00 o

Fig. 3. Representative examples of gilts with differing patterns of prolactin ( - - ) and substrate-directed behaviour (--; floor- and fixture-directed behaviour for gilts in crates; straw-directed behaviour for gilts in pens) over the 48 h before parturition. All gilts showed a rise in substrate-directed behaviour. (a) Gilts in crates with no statistical rise in prolactin; (b) gilts in crates with a statistical rise in prolactin; (c) gilts in pens with no statistical rise in prolactin; (d) gilts in pens with a statistical rise in prolactin.


70 Cortisol (nglml)

6 0

5 0

4O

10

0 p/In - 4 8 - 2 4 - 1 2 -~

f a r row ing

-6 ~3 0

Time period

F I F 2 ~ 7

Fig. 4. Average concentrat ions (with s.e.m. ) o f total plasma cortisol over the peri-parturient period for gilts in crates (11; n = 16) or pens ( [ ] ; n = 17). P r e / In ( P / I n ) values are shown for comparison. Time period - 48 refers to period - 4 8 / - 24, period - 24 refers to - 2 4 / - 12 etc.

having elevated cortisol relative to treatment P over much of the pre-parturient period (Fig. 4; overall effect of treatment: F1,24=20.48, P<0.001; time × treatment interaction: F1o,242 = 2.41, P< 0.01 ). The elevation of cortisol in treatment C gilts, although already apparent at time period - 4 8 / - 2 4 , did not reach significance by comparison with treatment P until period - 2 4 / - 12 (LSD test; P< 0.05). Comparison of means within treatment C gilts showed a strong tendency for cortisol to have increased by period - 4 8 / - 24 relative to the Pre/ In baseline (paired t-test: t~2= 2.06, P=0.062). Treatment C gilts subsequently experienced significant rises in cortisol between periods - 4 8 / - 2 4 to - 2 4 / - 12, - 3 / - 0 . 5 to 0/0.75 and 0/0.75 to 1/1.75 (all LSD: P<0.05). There were also non-significant reductions between periods - 1 2 / - 6 to - 6 / - 3, and 1 / 1.75 to 2/4 (see also Fig. 4 ). Comparison of means within treatment P showed there was no difference between concentrations at - 4 8 / - 24 and those at Pre/In (paired t-test: tl6= --0.75, ns). Treatment P concentrations only increased significantly between periods 0/0.75 to 1/1.75 (LSD: P<0.05 ). There were also non-significant rises between periods - 2 4 / - 12 to - 12 / -6 , and 1/1.75 to 2/4 (Fig. 4). As a result of these within treatment patterns of cortisol response, there was no difference in circulating cortisol between the treatments by period 2/4. There also continued to be no treatment effect on cortisol over days F 1, F2 and F7.

3.5. Relationships among behaviour, prolactin and cortisol

In general, there was little evidence of consistent correlations among behaviour, prolactin and cortisol. Cortisol and prolactin were correlated in treatment C only at period - 4 8 / - 2 4 (r=0.590, n= 16, P<0.05), but not thereafter (e.g. r= -0.26, ns at period - 1 2 / - 6). Cortisol and prolactin were not correlated at any stage in treatment P.

There were large individual differences in the pattern of prolactin and behaviour in the pre-parturient period. Certain individuals, although showing no ap-

326 A.B. Lawrence et al. / Appl. Anirn. Behav. Sci. 39 (1994) 313-330

parent rise in prolactin over the 48 and 24 h before parturition, still expressed increased substrate-directed behaviour (see above and Fig. 3 ). These individual differences were reflected in a general lack of correlation between prolactin and substrate-directed behaviour. In treatment P gilts, there were no significant correlations between prolactin and either time spent standing or straw-directed behaviour at any time period before parturition commenced. Floor-directed behaviour in treatment P was correlated to prolactin (r=0.593, n= 17, P<0.05) at time period - 4 8 / - 24, but not at other times. Behaviour was most correlated to prolactin at time period 1/1.75 when ventral lying, standing and alert were neg- atively related to prolactin ( r= - 0.554, r= - 0.554, r= - 0.630, respectively; all n = 17, P<0.05) , whilst lateral lying and inactive were positively correlated (r=0.603 and r=0.532, respectively; both P<0.05) . In treatment C there were no correlations among prolactin, standing and floor-directed behaviour at any time period before parturition. Fixture-directed behaviour was correlated with prolactin (r=0.552, P<0.05 ) at period - 1 2 / - 6 , but not at other times.

There were also few correlations between cortisol and behaviour in either treatment. For example, neither straw-directed behaviour in treatment P gilts, nor floor- and fixture-directed behaviour in treatment C were correlated with cortisol at any point before parturition.

4. Discussion

As with previous studies of peri-parturient behaviour (Hansen and Curtis, 1981; Vestergaard and Hansen, 1984; Lammers and De Lange, 1986; Heckt et al., 1988; Meunier-Salaun et al., 1991; Haskell, 1992 ), the present work found pigs in the pre-parturient period to be more active both in terms of the rate and amount of time spent standing. Again, in common with this previous work, from about 24 h before parturition there was an increase in substrate-directed behaviour. There is now considerable evidence to show that this pre-parturient behaviour is strongly modified by the environment and by the availability of substrates; in the present study gilts given the opportunity directed increasing amounts of behaviour at straw from 24 h pre-partum, although they still directed a small amount ofbehav- iour at the floor and walls of the pen. Gilts in crates during the same period spent increasing amounts of time in oral activities and pawing directed at the floor and the bars of the crate. Similarly, other studies of peri-parturient behaviour have reported increases in behaviour directed at straw (Widowski and Curtis, 1990; Arey et al., 1991; Castren et al., 1994), wood shavings (Cronin et al., 1993), cloth tassels (Widowski and Curtis, 1990) and, in the absence of manipulable substrates, at the floor, walls (Heckt et al., 1988 ) and fixtures such as the bars of the crate (Lammers and De Lange, 1986 ).

It has been proposed (e.g. Baxter, 1982; Vestergaard and Hansen, 1984; Lam- mers and De Lange, 1986) that this pre-parturient behaviour observed in con- fined sows is an expression of the nest-building activity observed in pigs under wild (Gundlach, 1968) and semi-natural conditions (Stolba and Wood-Gush,

A.B. Lawrence et al. / Appl. Anirn. Behav. Sci. 39 (1994) 313-330 327

1984; Jensen, 1986). A number of anthors (e.g. Vestergaard and Hansen, 1984; Widowski and Curtis, 1990) have suggested nest-building activities in the pig may be largely controlled by internal (physiological) stimuli, a view supported in this study by the spontaneous appearance of the behaviour in primiparous pigs in both environments.

Prolactin has been implicated in the initiation of nest-building in a number of species including mice (Voci and Carlson, 1973) and rabbits (Zarrow et al., 1971 ). The present study and others on pigs (Taverne et al., 1979; Dusza and Krymowska, 1981; Vale and Wagner, 1981; Meunier-Salaun et al., 1991 ) have shown prolactin to rise as parturition approaches. The present results may indicate that pre-parturient prolactin concentrations increase with parity in pigs, as the gilts in this study had concentrations of prolactin lower than those reported in other work that used multiparous sows of similar genotype (e.g. Meunier- Salaun et al., 1991 ).

In this study prolactin concentrations were unaffected by environment, and in both crates and pens broadly followed the same temporal pattern as the increase in substrate-directed behaviour. Taverne et al. (1979) also found similarity in the temporal patterning of prolactin and nest-building, and suggested that nest- building coincided with the pre-parturient prolactin peak. Furthermore, injection of prostaglandin (PGF2 alpha) has been shown to lead to an immediate increase in prolactin concentrations and a rapid commencement of nest-building (Blackshaw, 1983; Widowski et al., 1990). More recently, Castren et al. (1994) found prolactin concentrations at 48 h before parturition to be correlated with the onset of nest-building. They also found prolactin to increase before nest- building and to reach maximum values only after it, suggesting that the pre-parturient rise in prolactin concentration initiates nest-building. In this study we were unable to measure the onset of nest-building. However, we observed a number of individuals that despite lacking a significant rise in prolactin over the pre- parturient period, still expressed an increase in substrate-directed behaviour. This suggests that nest-building in gilts is not dependent on a rise in prolactin in the immediate pre-parturient period, and calls into question the supposed causal relationship between prolactin and nest-building in the pig. It may also- be that the close temporal relationship between the two is strengthened with increased parity, perhaps as a result of a more pronounced pre-parturition prolactin rise (see above).

Cortisol showed a similar temporal pattern to prolactin and substrate-directed behaviour in crated, but not penned gilts over the pre-parturient period. In- creased concentrations of porcine maternal cortisol in the pre-parturient phase have been reported in a number of studies (Killian et al., 1973; Molokwu and Wagner, 1973; Randall, 1983; Meunier-Salaun et al., 1991 ). Similarly, increases in maternal cortisol have been reported in sheep (Challis and Brooks, 1989) and humans (Carr et al., 1981 ).

Interest in glucocorticoids at parturition largely results from the central role of the fetal hypothalamic-pituitary-adrenal axis in the initiation of parturition in sheep (see Challis and Brooks, 1989, for a review), although in pigs it may play

328 A.B. Lawrence et al. / Appl. Anita. Behav. Sci. 39 (1994) 313-330

a less dominant role (Randall et al., 1990). The timing of the pre-parturient increase in maternal cortisol led a number of authors to conclude that it is a result rather than a cause of parturition (e.g. Molokwu and Wagner, 1973 ), perhaps as a response to parturition 'stress' (e.g. Genazzani et al., 1975 ). In the present work the increased concentrations of maternal cortisol that occurred largely after the onset of parturition in the penned gilts, could have been a response to the intrin- sically stress-inducing aspects of birth such as pain or novelty (e.g. neonate movements). However, the increased concentrations of maternal cortisol that occurred only in the crated gilts from 24 h before parturition, cannot be related to the parturition process itself but must, it seems, be a response to the crated environment.

The most striking difference between the crated and penned environments was the degree of behavioural restriction, and the modification of pre-parturient behaviour this brought about (see above). It has been suggested (e.g. Vestergaard and Hansen, 1984) that prevention of nest-building activity by close confinement is frustrating and stress-inducing. In this study, in addition to performing behaviour directed at floors and bars, crated gilts were scored as being more alert over much of the pre-farrowing period. In contrast, it was very apparent that gilts in pens were largely impervious to distracting stimuli when involved in straw- directed behaviour. These differences may indicate that the substrate-directed behaviour observed in the crated animals is not the motivational equivalent of the apparent nest-building observed in pens. The elevated cortisol in crated animals may then have reflected the physical interference of the crate in nest-building activity. However, the relative difference in cortisol between treatments reached a maximal level some time before the peak of substrate-directed behaviour was reached. Furthermore, there was no evidence of a correlational relationship between the level of substrate-directed behaviour and cortisol concentrations in crated animals at any time. An alternative possibility is that the elevation in cortisol reflected general discomfort in the crated animals that spent longer periods of time in ventral lying, tended to have higher rates of standing, and took longer to move between standing and lying down. However, there was no evidence of consistent correlations between any of these measures and cortisol concentrations.

5. Conclusions

In conclusion, the present work has confirmed the increase in activity and substrate-directed behaviour in sows during the pre-parturient period, and the considerable effect that parturition environment has on the expression of this pre- parturient behaviour. Prolactin followed broadly the same pattern as substrate- directed behaviour. However, the causal role of prolactin in nest-building must now be questioned given that a number of gilts, which failed to show a rise in prolactin in the pre-farrowing period, still showed increased substrate-directed behaviour. It may be that parity has a strong influence on pre-parturient concen-


trations of prolactin in the pig. There was evidence on the basis of cortisol concentrations that close confinement may be stress-inducing in the pre-parturient period, but it was not possible to be specific about the exact mechanism through which close confinement gave rise to elevated cortisol.

Acknowledgements

This work was supported by the Ministry for Agriculture, Fisheries and Food and the Scottish Office, Agriculture and Fisheries Department. We are grateful to Graham Horgan for assistance with the statistical analysis and Peter Finnie for assistance with care of the animals.

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The effect of environment on behaviour, plasma cortisol and prolactin in parturient sows