Alleviation of Hysteria in Laying Hens with Dietary Tryptophan Stephen R. Laycock and Ronald
ABSTRACT A commercial layer breeder flock that was suffering from hysteria was fed a diet containing 5 grams tryptophan/kg for six days. The incidence of episodes of hysteria declined from five times/hour on day 0 to once/hour on day 6 and none on day 8. Feed consumption increased from 107 g to 145 g/hen/day and egg production increased 23% during the six day feeding period. The tryptophan concentration in plasma doubled (from 95.6 to 188.2 ,umol/mL). Plasma phenylalanine and tyrosine also increased. Birds that were not in lay, by postmortem examination, had significantly higher plasma valine concentrations (476.4 vs 372.7 ,umol/ mL). Tryptophan, serotonin and related metabolites increased in both the hypothalamic region and the remainder of the brain following tryptophan feeding, and subsequently declined. High levels of dietary tryptophan may be useful in alleviating hysteria in poultry.
RESUME
L'effet de l'incorporation du tryptophane a la diete, a raison de 5 g/ kg pendant six jours, fut verifie dans un elevage aviaire dont les animaux presentaient un probleme dhysterie. L'incidence des episodes dhysterie a diminue de 5/h, au jour 0, a 1/h au jour 6, pour disparaitre totalement au jour 8. La consommation d'aliments passa de 107 g a 145 g/poule/jour et la production d'oeufs augmenta de 23% au cours de la periode d'essai de six jours. La concentration plasmatique du tryptophane a double (de 95.6 a
0.
Ball
188.2 ,mol/mL). La phenylalananine et la tyrosine plasmatiques augmenterent egalement. Chez les oiseaux qui
etaient en dehors de leur periode de ponte, on observa, au moment de l'evaluation post-mortem, des concentrations plasmatiques de valine beaucoup plus elevees (476.4 vs 372.7 ,umol/mL). Le tryptophane, la serotonine et leurs metabolites augmenterent au niveau de l'hypothalamus et des autres regions de l'encephale pendant l'alimentation au tryptophane, puis diminuerent par la suite. L'addition a la diete de quantites substantielles de tryptophane semble donc utile dans le traitement du probleme dhysterie chez la poule.
INTRODUCTION Hysteria in poultry is an occasional but significant economic problem. Hysteria has been reported in both caged and floor-raised hens (1-4). The result of severe hysteria is increased mortality and a dramatic reduction in egg production. Hansen (1) suggested that the two most plausible causes of hysteria are high population densities and pain resulting from hens clawing each other. Although many different treatments have been tried to alleviate hysteria, including changes in light intensity, claw removal and addition of niacin to the diet (1) none of these has been successful. Given the sedative-like effect of excess tryptophan in mammals and the data (5,6) which indicate that brain neurotransmitters in chicks could be altered by dietary amino acids, it was hypothesized that dietary tryptophan could be an effective treatment for hysteria.
MATERIALS AND METHODS A commercial layer breeder flock (n was used in this study. The barn consisted of four rooms, each measuring approximately 23 m long and 12 m wide. The floors consisted of wooden slats in the centre (9 m wide) and a sand runway along either wall. Each room contained approximately 2200 hens and at the start of the trial the hens were 47 weeks old. One of the rooms was selected for study. The guidelines of the Guide to the Care and Use of Experimental Animals of the Canadian Council on Animal Care were followed. The mean daytime barn ambient temperature was between 19°C and 21°C and the relative humidity was approximately 60%. Composition of the diet is shown in Table I. Proximate analysis was conducted by standard methods (7). The tryptophan (TRP) treated diet was supplemented with 5 g of TRP per kg of mixed complete feed and provided to hens in the selected room. The original diet was adequate in tryptophan, supplying 2.6 g/ kg by =
10,000)
analysis. The first sampling (Day 0) was performed on eight hens which had been consuming the control diet. Immediately after collection, at 09:00, the feed containing TRP was offered. The quantity of feed offered was recorded each day in order to estimate feed intake. The schedule of feeding and sampling is given in Table II. Observations of hysteria were made on days 0, 2, 4, 6 and 8 to count the number of episodes per hour. The observer stood outside the room for a one hour period, prior to sampling of the birds and recorded the number of spontaneous episodes (hysteria).
Department of Animal and Poultry Science, University of Guelph, Guelph, Ontario NIG 2W1. Present address of S.R. Laycock: Rhone-Poulenc Canada Inc., Mississauga, Ontario. Reprint requests to R.O. Ball. Submitted August 29, 1989.
Can J Vet Res 1990; 54: 291-295
291
TABLE I. Composition of diet
Ingredient Corn Soybean meal (44%) Meat meal (55%) Limestone Oyster shell Dicalcium phosphate Fat Salt Methionine Ethyacal Mineral/vitamin premix
Analysis Dry matter Protein Crude fiber Calcium Phosphorous Gross energy (Kcal/g) Tryptophan
g/ kg 618.4 220.0 50.0 33.0 50.0 10.0 5.0 3.0 1.0 7.5 1.1 1000.0 885.9 180.6 37.3 29.1 6.8 4.3 2.6
Following the hysteria observations eight birds from the study room were sampled at random for measures of metabolism. Each hen was decapitated within 30 s of being caught and removed from the room. Immediately upon decapitation, 5 mL of mixed arterial and venous blood were collected in heparinized tubes and placed on ice. The brain was then removed and dissected into a hypothalamic portion (approximately 0.5 g) and a portion composed of the remainder of the brain. Each sample was weighed and then immediately frozen in liquid nitrogen and stored in crushed ice during transport to a -80° C freezer for storage until analysis. Hens were palpated for distance between the pubic bones to determine if they were in lay. Carcasses were submitted for postmortem examina-
tion to the Department of Pathology, University of Guelph. Blood, brain and feed samples were analyzed for amino acids by high performance liquid chromatography (HPLC) as described (8) using a Waters Pico-TagT" system (Millipore/ Waters Ltd., Mississauga, Ontario) consisting of WISP 710B autosampler, two M6000A pumps, temperature control module, M440 ultraviolet absorbance detector and 840 data analysis system. Brain neurotransmitters were determined using the following procedure. Approximately 0.5 g of tissue were weighed into a glass test tube containing 2.0 mL H2SO4 solution (60 ,uL concentrated H2SO4, 0.1 g sodium metabisulfite, 100 mL filtered deionized double distilled organic free water) and 50 ,uL of internal standard (10 mM 4hydroxy-3-methoxyphenyl acetic acid). This mixture was vortexed briefly, homogenized for 20 s, and centrifuged at 10,00 x g for 10 min. The supernatant was extracted and filtered through a 0.22 ,ifilter (Millex GS, Millipore/ Waters) into a 1.5 mL microfuge tube containing 25 ,uL Tris buffer. The sample was vortexed and frozen at -80° C for subsequent analysis. The HPLC analysis consisted of an autosampler refrigerated to 40 C (Model AS-100, Biorad, Toronto, Ontario), an M5 10 pump, catecholamine column
and M460 electrochemical detector (all from Millipore/ Waters). The running conditions were 0.8 mL/ min of eluant (0.1 M sodium acetate, 0.1 M citric acid, 0.5 mM sodium octane sulfonate, 0. 15 mM ethylenediamine tetraacetate, 1.0 mM di-N-butylamine, 10% methanol) for 25 min with the electrochemical detector set at a potential of 0.85 volts. The effect of day on measured parameters was assessed using the General Linear Model procedure of the Statistical Analysis System (9). Where an overall significant effect was observed the differences among means were assessed by a Student-NewmanKuels multiple range test at the 0.05 level (9).
RESULTS AND DISCUSSION PRODUCTION PARAMETERS
to
Egg production had decreased, due hysteria, from 1707 eggs/day on
March 23 to 392 eggs/day on March 30 in the room used for the study. This low level of production was maintained until the commencement of the experiment on May 4. No apparent physiological or clinical reason for such a dramatic decrease in production could be determined, other than the obvious hysteria. The production
EGGS/DAY 650 600
550 500
450 400
TABLE II. Schedule of supplemental tryptophan feeding and sampling of hens No. hens No. hens Diet Day sampled in lay 0 8 Control/TRPa 5 TRP 1 2 TRP 8 6 3 TRP 4 TRP 8 5 5 TRP 6 TRP 8 2 7 Control 8 Control 8 6 aTryptophan (TRP) diet was introduced following sampling of hens
292
350 300 250
200
150
100 50 0
0
1
2
3
4
5
6
DAY OF TRIAL Fig. 1. Effects of additional dietary tryptophan on the egg production rate.
7
8
13
indicated emaciation, bacterial hepatitis, and septicemia, probably caused by the severe state of hysteria. In the week prior to the TRP supplementation, feed consumption was approximately 107 g/hen/day. The TRP supplementation to the diet resulted in an increase in feed consumption to approximately 145 g/ hen/day and an increase in egg production by 23% (Fig. 1) during the six day feeding period. Egg production declined thereafter as the hysteria returned.
EPISODES/HOUR 6
5-
4-
3-
2-
HYSTERIA OBSERVATIONS
Hysteria became a serious problem in the entire barn over a one week period from March 23 to March 30, 1988. Hens were observed to be calmly 1 walking about, intermingling with 0 2 4 6 8 roosters, when the entire room would DAY OF TRIAL suddenly be affected for no apparent Fig. 2. Effects of additional dietary tryptophan on the degree of hysteria (episodes/h). reason. Normally, hysteria would originate from one point in the room and spread out in all directions. Hens would run madly in circles of approxipattern was similar in the other three March 30 to May 4 the owner tried a mately 6 m diameter. Roosters were rooms but varied in timing and number of treatments, all unsuccess- never observed to be involved in the severity according to the onset and fully. Postmortem examination of hysteria. degree of hysteria. During the period birds prior to and during the study Hens exploded into hysteria five times/ hour at the commencement of the trial. By day 2 of the trial, the hens were observed to be much calmer, with many laying around the watering umol/ml bowls. This had not been previously 550 observed. By day 2 of the trial the number of episodes had been reduced 500 to three/hour, and hysteria com450 pletely disappeared by day 8 of the trial. 400 Hysteria gradually returned during the week after the hens were again 350 placed on the control diet (Fig. 2). 300 Repeating the TRP treatment with the whole barn, rather thanjust one room, 250 again eliminated hysteria, which did not return thereafter. This suggests 200 that the return of hysteria to the first 150 room was a result of the psychological pressure caused by the audible activity 100 in the other rooms of the barn. 50 Although detailed observations were not taken when the treatment was repeated, the fact that the treatment 2 4 6 was effective twice is strong supportDAY OF TRIAL ing evidence for an effect of TRP.
OL
~
_-
--
~
--
--
0
0
Fig. 3. Effects of additional dietary tryptophan on plasma amino acid levels in the hen. The lines represent: tryptophan ( ), phenylalanine (---), tyrosine (--- -), and valine (- -)The standard error for each amino acid was 19.3, 26.8, 12.5 and 25.2, respectively. Treatment significantly affected the concentration of tryptophan (p < 0.01), phenylalanine (p < 0.01) and valine (p < 0.05) compared to control values (day 0).
PLASMA AMINO ACIDS
Plasma free TRP levels increased twofold (p < 0.01) following introduc-
293
nmol/ml 12 11
10 9
8 7
6 5 4
3 2
0
2
4
6
8 8
DAY OF TRIAL Fig. 4. Effects of additional dietary tryptophan on related metabolites in the hypothalamic regiion. ), serotonin (--- -), 5-hydroxyindole acetic acid (---) an(d 5The lines represent: tryptophan ( hydroxytryptophan (- .-). The standard error for each compound was 0.15, 0.56, 0.11 and 0 .10, respectively. Values on days 2, 4 and 6 were significantly different (p < 0.01) from control (day 0) for all compounds.
tion of the TRP supplemented diet (Fig. 3) from 95.6 ,tmol/mL on day 0 to 188.2 ,umol/mL on day 2. Plasma
TRP concentration remained high until day 6 when the diet was withdrawn and subsequently de-
nmol/ml 8 7
6 5 4
3 2
0
2
4
6
DAY OF TRIAL Effects of additional dietary tryptophan on related metabolites in the brain. The lines represent: tryptophan ( ), serotonin (--- -), 5-hydroxyindole acetic acid (---) and 5hydroxytryptophan (- .-). The standard error for each compound was 0.25, 0.35, 0.20 and 0.10, respectively. Values on days 2, 4 and 6 were significantly different (p < 0.01) from control (day 0) for all compounds.
Fig.
294
5.
creased to 140.3 ,imol/mL on day 8. Plasma phenylalanine increased from 136.9 to 171 ,umol/ mL (p < 0.05) between day 0 and 6. Tyrosine showed a similar trend, increasing from 158.6 to 205.5 ,umol/mL, but the difference was not significant. Plasma valine concentration declined, but not significantly, during the first four days of the trial from 450.4 to 360.1 ,.tmol/mL then increased sharply to 524 ,umol/mL (p < 0.05) on day 6 when the TRP diet was withdrawn, followed by a decline to 394.7 ,imol/mL on day 8 (Fig. 3). Analysis of the data for all birds showed that birds that were not in lay, as determined by postmortem examination, had significantly (p < 0.01) higher plasma valine concentrations: 476.4 vs 372.7 ,tmol/mL (SEM 15.9). Inspection of the data for day 6, when mean plasma valine was highest, revealed that
none
of the birds
sampled that day was in lay. There was no relationship to laying for any of the other amino acids. The reason for the effect on valine is unknown. HYPOTHALAMUS AND BRAIN
Tryptophan concentration in the hypothalamus increased (p < 0.01) following introduction of the TRP supplemented diet from 1.66 nmol/g on day 0 to 2.73 nmol/ g on day 4 (Fig. 4). The lower value observed on day 6 may have been a result of the high plasma valine concentration since plasma valine and TRP compete for transport into the brain (10). Alternatively, the values may have reflected adaptation to the diet in amino acid transport and metabolism. The lower values on day 8 probably reflected the lower plasma TRP concentration as a result of withdrawal of the supplemented feed. Concentration of TRP metabolites in the hypothalamus, serotonin (5HT), 5-hydroxyindole acetic acid (5HIAA) and 5-hydroxy tryptophan (5OHTRP), generally reflected the concentration of TRP except on day 2. We cannot explain this anomalous value on day 2, although it may relate to some other factor beyond our measurements, similar to the observed relationship between plasma valine and state of lay. These data indicate that further research should be conducted into the factors influencing serotonin concentration in poultry.
Analysis of the remaining portion of the brain showed trends (Fig. 5) similar to the hypothalamus but with generally lower concentrations. In conclusion, these data show that in a commercial environment, diet supplementation with TRP at 5 g per kg of complete feed appeared to be an effective treatment for hysteria in poultry. The reduction in hysteria appeared to follow a similar trend to the changes in plasma and brain amino acids and in hypothalamic neurotransmitters. Since only one level of tryptophan was used, a number of questions remain regarding possible optimum concentrations and feeding periods.
ACKNOWLEDGMENTS Special thanks to Halchemix Canada/ Heartland Lysine for a donation of tryptophan and D. Price-Jones for expert assistance. Supported by the Ontario Ministry of Agriculture and Food. REFERENCES 1. HANSEN RS. Nervousness and hysteria of mature female chickens. Poult Sci 1976; 55: 531-543. 2. JONES RB. Fear response of individually caged laying hens as a function of cage level and aisle. Appl Anim Behav Sci 1985; 14: 63-74. 3. NICOL CJ. Effect of cage height and area on the behaviour of hens housed in battery cages. Br Poult Sci 1987; 28: 327-335. 4. OKPOKHO NA, CRAIG JV. Fear-related behavior of hens in cages. Effects of hearing
5.
6.
7.
8.
9.
10.
environment, age and habituation. Poult Sci 1987; 66: 376-377. HARRISON L, D'MELLO JPF. Large neutral amino acids in the diet and neurotransmitter concentrations in the chick brain. Proc Nutr Soc 1986; 45: 72A. HARRISON L, D'MELLO JPF. Zinc deficiency, amino acid imbalance and brain catecholamine concentration in the chick. Proc Nutr Soc 1987; 46: 58A. ASSOCIATION OF OFFICIAL ANALYTICAL CHEMISTS. Official Methods of Analysis (12th ed). Washington, DC, 1975. EARLY RJ, BALL RO. Amino acid analysis of physiological fluids and some applications in biological research. J Anal Purification 1987; 2: 47-51. STATISTICAL ANALYSIS SYSTEM. SAS User's Guide: Statistics. Cary, North Carolina: Statistical Analysis System Institute, 1985. LEATHWOOD PD. Tryptophan availability and serotonin synthesis. Proc Nutr Soc 1987; 46: 143-156.
295
ABSTRACT A commercial layer breeder flock that was suffering from hysteria was fed a diet containing 5 grams tryptophan/kg for six days. The incidence of episodes of hysteria declined from five times/hour on day 0 to once/hour on day 6 and none on day 8. Feed consumption increased from 107 g to 145 g/hen/day and egg production increased 23% during the six day feeding period. The tryptophan concentration in plasma doubled (from 95.6 to 188.2 ,umol/mL). Plasma phenylalanine and tyrosine also increased. Birds that were not in lay, by postmortem examination, had significantly higher plasma valine concentrations (476.4 vs 372.7 ,umol/ mL). Tryptophan, serotonin and related metabolites increased in both the hypothalamic region and the remainder of the brain following tryptophan feeding, and subsequently declined. High levels of dietary tryptophan may be useful in alleviating hysteria in poultry.
RESUME
L'effet de l'incorporation du tryptophane a la diete, a raison de 5 g/ kg pendant six jours, fut verifie dans un elevage aviaire dont les animaux presentaient un probleme dhysterie. L'incidence des episodes dhysterie a diminue de 5/h, au jour 0, a 1/h au jour 6, pour disparaitre totalement au jour 8. La consommation d'aliments passa de 107 g a 145 g/poule/jour et la production d'oeufs augmenta de 23% au cours de la periode d'essai de six jours. La concentration plasmatique du tryptophane a double (de 95.6 a
0.
Ball
188.2 ,mol/mL). La phenylalananine et la tyrosine plasmatiques augmenterent egalement. Chez les oiseaux qui
etaient en dehors de leur periode de ponte, on observa, au moment de l'evaluation post-mortem, des concentrations plasmatiques de valine beaucoup plus elevees (476.4 vs 372.7 ,umol/mL). Le tryptophane, la serotonine et leurs metabolites augmenterent au niveau de l'hypothalamus et des autres regions de l'encephale pendant l'alimentation au tryptophane, puis diminuerent par la suite. L'addition a la diete de quantites substantielles de tryptophane semble donc utile dans le traitement du probleme dhysterie chez la poule.
INTRODUCTION Hysteria in poultry is an occasional but significant economic problem. Hysteria has been reported in both caged and floor-raised hens (1-4). The result of severe hysteria is increased mortality and a dramatic reduction in egg production. Hansen (1) suggested that the two most plausible causes of hysteria are high population densities and pain resulting from hens clawing each other. Although many different treatments have been tried to alleviate hysteria, including changes in light intensity, claw removal and addition of niacin to the diet (1) none of these has been successful. Given the sedative-like effect of excess tryptophan in mammals and the data (5,6) which indicate that brain neurotransmitters in chicks could be altered by dietary amino acids, it was hypothesized that dietary tryptophan could be an effective treatment for hysteria.
MATERIALS AND METHODS A commercial layer breeder flock (n was used in this study. The barn consisted of four rooms, each measuring approximately 23 m long and 12 m wide. The floors consisted of wooden slats in the centre (9 m wide) and a sand runway along either wall. Each room contained approximately 2200 hens and at the start of the trial the hens were 47 weeks old. One of the rooms was selected for study. The guidelines of the Guide to the Care and Use of Experimental Animals of the Canadian Council on Animal Care were followed. The mean daytime barn ambient temperature was between 19°C and 21°C and the relative humidity was approximately 60%. Composition of the diet is shown in Table I. Proximate analysis was conducted by standard methods (7). The tryptophan (TRP) treated diet was supplemented with 5 g of TRP per kg of mixed complete feed and provided to hens in the selected room. The original diet was adequate in tryptophan, supplying 2.6 g/ kg by =
10,000)
analysis. The first sampling (Day 0) was performed on eight hens which had been consuming the control diet. Immediately after collection, at 09:00, the feed containing TRP was offered. The quantity of feed offered was recorded each day in order to estimate feed intake. The schedule of feeding and sampling is given in Table II. Observations of hysteria were made on days 0, 2, 4, 6 and 8 to count the number of episodes per hour. The observer stood outside the room for a one hour period, prior to sampling of the birds and recorded the number of spontaneous episodes (hysteria).
Department of Animal and Poultry Science, University of Guelph, Guelph, Ontario NIG 2W1. Present address of S.R. Laycock: Rhone-Poulenc Canada Inc., Mississauga, Ontario. Reprint requests to R.O. Ball. Submitted August 29, 1989.
Can J Vet Res 1990; 54: 291-295
291
TABLE I. Composition of diet
Ingredient Corn Soybean meal (44%) Meat meal (55%) Limestone Oyster shell Dicalcium phosphate Fat Salt Methionine Ethyacal Mineral/vitamin premix
Analysis Dry matter Protein Crude fiber Calcium Phosphorous Gross energy (Kcal/g) Tryptophan
g/ kg 618.4 220.0 50.0 33.0 50.0 10.0 5.0 3.0 1.0 7.5 1.1 1000.0 885.9 180.6 37.3 29.1 6.8 4.3 2.6
Following the hysteria observations eight birds from the study room were sampled at random for measures of metabolism. Each hen was decapitated within 30 s of being caught and removed from the room. Immediately upon decapitation, 5 mL of mixed arterial and venous blood were collected in heparinized tubes and placed on ice. The brain was then removed and dissected into a hypothalamic portion (approximately 0.5 g) and a portion composed of the remainder of the brain. Each sample was weighed and then immediately frozen in liquid nitrogen and stored in crushed ice during transport to a -80° C freezer for storage until analysis. Hens were palpated for distance between the pubic bones to determine if they were in lay. Carcasses were submitted for postmortem examina-
tion to the Department of Pathology, University of Guelph. Blood, brain and feed samples were analyzed for amino acids by high performance liquid chromatography (HPLC) as described (8) using a Waters Pico-TagT" system (Millipore/ Waters Ltd., Mississauga, Ontario) consisting of WISP 710B autosampler, two M6000A pumps, temperature control module, M440 ultraviolet absorbance detector and 840 data analysis system. Brain neurotransmitters were determined using the following procedure. Approximately 0.5 g of tissue were weighed into a glass test tube containing 2.0 mL H2SO4 solution (60 ,uL concentrated H2SO4, 0.1 g sodium metabisulfite, 100 mL filtered deionized double distilled organic free water) and 50 ,uL of internal standard (10 mM 4hydroxy-3-methoxyphenyl acetic acid). This mixture was vortexed briefly, homogenized for 20 s, and centrifuged at 10,00 x g for 10 min. The supernatant was extracted and filtered through a 0.22 ,ifilter (Millex GS, Millipore/ Waters) into a 1.5 mL microfuge tube containing 25 ,uL Tris buffer. The sample was vortexed and frozen at -80° C for subsequent analysis. The HPLC analysis consisted of an autosampler refrigerated to 40 C (Model AS-100, Biorad, Toronto, Ontario), an M5 10 pump, catecholamine column
and M460 electrochemical detector (all from Millipore/ Waters). The running conditions were 0.8 mL/ min of eluant (0.1 M sodium acetate, 0.1 M citric acid, 0.5 mM sodium octane sulfonate, 0. 15 mM ethylenediamine tetraacetate, 1.0 mM di-N-butylamine, 10% methanol) for 25 min with the electrochemical detector set at a potential of 0.85 volts. The effect of day on measured parameters was assessed using the General Linear Model procedure of the Statistical Analysis System (9). Where an overall significant effect was observed the differences among means were assessed by a Student-NewmanKuels multiple range test at the 0.05 level (9).
RESULTS AND DISCUSSION PRODUCTION PARAMETERS
to
Egg production had decreased, due hysteria, from 1707 eggs/day on
March 23 to 392 eggs/day on March 30 in the room used for the study. This low level of production was maintained until the commencement of the experiment on May 4. No apparent physiological or clinical reason for such a dramatic decrease in production could be determined, other than the obvious hysteria. The production
EGGS/DAY 650 600
550 500
450 400
TABLE II. Schedule of supplemental tryptophan feeding and sampling of hens No. hens No. hens Diet Day sampled in lay 0 8 Control/TRPa 5 TRP 1 2 TRP 8 6 3 TRP 4 TRP 8 5 5 TRP 6 TRP 8 2 7 Control 8 Control 8 6 aTryptophan (TRP) diet was introduced following sampling of hens
292
350 300 250
200
150
100 50 0
0
1
2
3
4
5
6
DAY OF TRIAL Fig. 1. Effects of additional dietary tryptophan on the egg production rate.
7
8
13
indicated emaciation, bacterial hepatitis, and septicemia, probably caused by the severe state of hysteria. In the week prior to the TRP supplementation, feed consumption was approximately 107 g/hen/day. The TRP supplementation to the diet resulted in an increase in feed consumption to approximately 145 g/ hen/day and an increase in egg production by 23% (Fig. 1) during the six day feeding period. Egg production declined thereafter as the hysteria returned.
EPISODES/HOUR 6
5-
4-
3-
2-
HYSTERIA OBSERVATIONS
Hysteria became a serious problem in the entire barn over a one week period from March 23 to March 30, 1988. Hens were observed to be calmly 1 walking about, intermingling with 0 2 4 6 8 roosters, when the entire room would DAY OF TRIAL suddenly be affected for no apparent Fig. 2. Effects of additional dietary tryptophan on the degree of hysteria (episodes/h). reason. Normally, hysteria would originate from one point in the room and spread out in all directions. Hens would run madly in circles of approxipattern was similar in the other three March 30 to May 4 the owner tried a mately 6 m diameter. Roosters were rooms but varied in timing and number of treatments, all unsuccess- never observed to be involved in the severity according to the onset and fully. Postmortem examination of hysteria. degree of hysteria. During the period birds prior to and during the study Hens exploded into hysteria five times/ hour at the commencement of the trial. By day 2 of the trial, the hens were observed to be much calmer, with many laying around the watering umol/ml bowls. This had not been previously 550 observed. By day 2 of the trial the number of episodes had been reduced 500 to three/hour, and hysteria com450 pletely disappeared by day 8 of the trial. 400 Hysteria gradually returned during the week after the hens were again 350 placed on the control diet (Fig. 2). 300 Repeating the TRP treatment with the whole barn, rather thanjust one room, 250 again eliminated hysteria, which did not return thereafter. This suggests 200 that the return of hysteria to the first 150 room was a result of the psychological pressure caused by the audible activity 100 in the other rooms of the barn. 50 Although detailed observations were not taken when the treatment was repeated, the fact that the treatment 2 4 6 was effective twice is strong supportDAY OF TRIAL ing evidence for an effect of TRP.
OL
~
_-
--
~
--
--
0
0
Fig. 3. Effects of additional dietary tryptophan on plasma amino acid levels in the hen. The lines represent: tryptophan ( ), phenylalanine (---), tyrosine (--- -), and valine (- -)The standard error for each amino acid was 19.3, 26.8, 12.5 and 25.2, respectively. Treatment significantly affected the concentration of tryptophan (p < 0.01), phenylalanine (p < 0.01) and valine (p < 0.05) compared to control values (day 0).
PLASMA AMINO ACIDS
Plasma free TRP levels increased twofold (p < 0.01) following introduc-
293
nmol/ml 12 11
10 9
8 7
6 5 4
3 2
0
2
4
6
8 8
DAY OF TRIAL Fig. 4. Effects of additional dietary tryptophan on related metabolites in the hypothalamic regiion. ), serotonin (--- -), 5-hydroxyindole acetic acid (---) an(d 5The lines represent: tryptophan ( hydroxytryptophan (- .-). The standard error for each compound was 0.15, 0.56, 0.11 and 0 .10, respectively. Values on days 2, 4 and 6 were significantly different (p < 0.01) from control (day 0) for all compounds.
tion of the TRP supplemented diet (Fig. 3) from 95.6 ,tmol/mL on day 0 to 188.2 ,umol/mL on day 2. Plasma
TRP concentration remained high until day 6 when the diet was withdrawn and subsequently de-
nmol/ml 8 7
6 5 4
3 2
0
2
4
6
DAY OF TRIAL Effects of additional dietary tryptophan on related metabolites in the brain. The lines represent: tryptophan ( ), serotonin (--- -), 5-hydroxyindole acetic acid (---) and 5hydroxytryptophan (- .-). The standard error for each compound was 0.25, 0.35, 0.20 and 0.10, respectively. Values on days 2, 4 and 6 were significantly different (p < 0.01) from control (day 0) for all compounds.
Fig.
294
5.
creased to 140.3 ,imol/mL on day 8. Plasma phenylalanine increased from 136.9 to 171 ,umol/ mL (p < 0.05) between day 0 and 6. Tyrosine showed a similar trend, increasing from 158.6 to 205.5 ,umol/mL, but the difference was not significant. Plasma valine concentration declined, but not significantly, during the first four days of the trial from 450.4 to 360.1 ,.tmol/mL then increased sharply to 524 ,umol/mL (p < 0.05) on day 6 when the TRP diet was withdrawn, followed by a decline to 394.7 ,imol/mL on day 8 (Fig. 3). Analysis of the data for all birds showed that birds that were not in lay, as determined by postmortem examination, had significantly (p < 0.01) higher plasma valine concentrations: 476.4 vs 372.7 ,tmol/mL (SEM 15.9). Inspection of the data for day 6, when mean plasma valine was highest, revealed that
none
of the birds
sampled that day was in lay. There was no relationship to laying for any of the other amino acids. The reason for the effect on valine is unknown. HYPOTHALAMUS AND BRAIN
Tryptophan concentration in the hypothalamus increased (p < 0.01) following introduction of the TRP supplemented diet from 1.66 nmol/g on day 0 to 2.73 nmol/ g on day 4 (Fig. 4). The lower value observed on day 6 may have been a result of the high plasma valine concentration since plasma valine and TRP compete for transport into the brain (10). Alternatively, the values may have reflected adaptation to the diet in amino acid transport and metabolism. The lower values on day 8 probably reflected the lower plasma TRP concentration as a result of withdrawal of the supplemented feed. Concentration of TRP metabolites in the hypothalamus, serotonin (5HT), 5-hydroxyindole acetic acid (5HIAA) and 5-hydroxy tryptophan (5OHTRP), generally reflected the concentration of TRP except on day 2. We cannot explain this anomalous value on day 2, although it may relate to some other factor beyond our measurements, similar to the observed relationship between plasma valine and state of lay. These data indicate that further research should be conducted into the factors influencing serotonin concentration in poultry.
Analysis of the remaining portion of the brain showed trends (Fig. 5) similar to the hypothalamus but with generally lower concentrations. In conclusion, these data show that in a commercial environment, diet supplementation with TRP at 5 g per kg of complete feed appeared to be an effective treatment for hysteria in poultry. The reduction in hysteria appeared to follow a similar trend to the changes in plasma and brain amino acids and in hypothalamic neurotransmitters. Since only one level of tryptophan was used, a number of questions remain regarding possible optimum concentrations and feeding periods.
ACKNOWLEDGMENTS Special thanks to Halchemix Canada/ Heartland Lysine for a donation of tryptophan and D. Price-Jones for expert assistance. Supported by the Ontario Ministry of Agriculture and Food. REFERENCES 1. HANSEN RS. Nervousness and hysteria of mature female chickens. Poult Sci 1976; 55: 531-543. 2. JONES RB. Fear response of individually caged laying hens as a function of cage level and aisle. Appl Anim Behav Sci 1985; 14: 63-74. 3. NICOL CJ. Effect of cage height and area on the behaviour of hens housed in battery cages. Br Poult Sci 1987; 28: 327-335. 4. OKPOKHO NA, CRAIG JV. Fear-related behavior of hens in cages. Effects of hearing
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