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The Effect of Bovine Growth Hormone, Desoxycorticosterone, and Cortisone on the Weight of the Bursa of Fabricius, Adrenal Glands, Heart, and Body Weight of Young Chickens
VITAMIN B12 IN DEVELOPING EGGS
min Ba, activity assay. Fifteenth edition, pp. 885-889. Robinson, R. A., M. E. Fitzgerald, K. Fehr and J. J. Grimshaw, 1954. Vitamin B ^ in crude liver extracts. Nature, 174: 558-559. Sehiya, S., 1953. Jap. J. Pharmacol. 3 : 1. Quoted by Fischer et al. (1958). Skarzynski, B., Z. Zak, W. Ostrowski, M. K. Davies, M. E. Gregory and S. K. Kon, 1959. Vitamin Bis in the developing chick embryo. Brit. J. Nutrition, 13: 72-78. Snell, E. E., and E. Quarles, 1941. The effect of incubation on the vitamin content of eggs. J. Nutrition, 22: 483-489. Swendseid, M. E., R. Jackson and E. M. Benson, 1958. The formation of citrovorum factor in the developing chick egg and the effect of injecting folic' acid antagonists. Arch. Biochem. Bioph. 78: 221-226. Tastaldi, H., 1959. Ensaio microbiologico da vitamina BM. Praticas de Quimica Biologica, 2nd part, 4th edition, pp. 194-199. Woolley, W., 1954. Synthesis of vitamin Bis by spontaneous cancers. Federation Proc. 13: 482483. Yacowitz, H., C. H. Hill, L. C. Norris and G. F. Heuser, 1952. Distribution of vitamin B« in the organs and tissue of the chick. Proc. Soc. Exptl. Biol. Med. 79: 279-280.
The Effect of Bovine Growth Hormone, Desoxycorticosterone, and Cortisone on the Weight of the Bursa of Fabricius, Adrenal Glands, Heart, and Body Weight of Young Chickens BRUCE G L I C K
Department of Poultry Husbandry, Mississippi State University, State College, Mississippi (Received for publication March 8, 1960)
R
ESEARCH to date has revealed little or no effect of growth hormone in young chickens, Carter et al. (1955) and Libby et al. (1955). Hsieh et al. (1952) have reported a growth promoting and diabetogenic effect of growth hormone in chick embryos. Guinea pigs and man appear to be insensitive to bovine growth hormone, Selye (1955), while the data of Beck et al. Journal Article No. 807
(1957) indicate a slight but significant growth effect of human and monkey growth hormone on a 13-year-old human male. Lack of stimulation by growth hormone may be due to contamination with adrenocorticotropic hormone, to treatment procedure, to species differences, Wilhelmi (1955) and Knobil (1955), or to "conditioning" of tissue to growth hormone by the adrenal glands, Selye (1955) and Selye and Heuser (1954).
Downloaded from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015
cient chick embryos. A.M.A. Arch. Pathol. 60: 393-400. Fischer, N. A., E. M. Benson and M. E. Swendseid, 19S8. The distribution of vitamin Bis in the developing chick egg. Arch. Biochem. Bioph. 74: 458-464. Hayes, M., and T. P. Luckey, 1957. Effect of incubation upon folic acid content of eggs. Proc. Soc. Exptl. Biol. Med. 94: 777-778. Jackson, J. I., L. J. Machlin, E. A. Brandenburger, W. L. Kellog and C. A. Denton, 1953. Retention of Co60 labeled vitamin Bi2 in chickens. Proc. Soc. Exptl. Biol. Med. 83: 221-222. Krieger, C. H., 1952. Report on vitamin Bu,. microbiological method. J. Ass. Off. Agr. Chem. 35: 726-736. Lillie, R. J., M. W. Olsen and H. R. Bird, 1949. Role of vitamin B« in reproduction of poultry. Proc. Soc. Exptl. Biol. Med. 72: 598-602. Mariakulandai, A., and J. McGinnis, 1953. The vitamin B^ requirement for hatchability of chicken eggs. Poultry Sci. 32: 3-7. Nogueira, D. M., 1960. Vitamin Bis activity in chick egg. In press. Olcese, O., J. R. Couch, J. H. Quisenberry and P. B. Pearson, 1950. Congenital anomalies in the chick due to vitamin Bis deficiency. J. Nutrition. 4 1 : 423-431. Pharmacopoeia of the United States, 1955. Vita-
1527
1528
B. GLICK TABLE 1.—The effect of growth hormone on body weight gain of 9-23-day-old chickens Treatments
Age (16 birds per group)
9-day-old (initial wt.), gm. 9-16-day-old, gm.1 16-23-day-old, gm.1 1
No injection
Saline
0
.4 ml./injection
.2 ml./inj. (2 mg.)
Growth hormone .6 ml./inj. (6 mg.)
101 + 2.1 89+3.0 113 + 3.7
107 + 2.1 93 + 3.0 105 + 2.7
106 + 1.6 93 + 3.7 107 + 5.5
107 + 2.0 89 + 2.7 107 + 5.0
Weight gain.
METHOD The effect of growth hormone, desoxycorticosterone acetate, and a combination of growth hormone (GH), desoxycorticosterone acetate (DOCA), and cortisone on the growth of the body, bursa of Fabricius, adrenals, and heart was studied in 1-3-weekold New Hampshire chickens. All birds were raised in batteries and fed a basal ration. Growth hormone in the form of a dry powder was taken up in physiological
saline, allowing each milliliter of solution to contain 10 mg. of the hormone. The desoxycorticosterone acetate was suspended in sesame oil to allow each milliliter of solution to contain 10 mg. of the hormone. Each milliliter of the saline suspension of cortone contained 50 mg. of cortisone. Growth hormone and saline and desoxycorticosterone acetate and oil were administered intramuscularly from the ninth to the twenty-third day of age, respectively. All treatments were replicated. Growth hormone (.4 ml.), desoxycorticosterone (.2 ml.), cortisone (.05 ml.), saline (.4 ml.), and sesame oil (.2 ml.) were administered intramuscularly from the twelfth to the eighteenth day of age. Equal numbers of males and females were used in all trials. Body and gland weights were analyzed by the analysis of variance and covariance, respectively, Snedecor (1946) Duncan's (1955) new multiple range and F test was utilized when determining which means differed significantly. Blood chlorides were de-
TABLE 2.—The eject of growth hormone on the weight of the bursa of Fabricius, heart, and adrenal glands of 23-day-old chickens1 Treatments Glands, grams (12 birds per treat.)
Bursa of Fabricius Heart Adrenal 1
No injection
Saline
Growth hormone
0
.2 ml./injection
.2 ml./inj. (2 mg.) .6 ml./inj. (6 mg.)
.654+ .067 2.089+.012 .040+ .002
.591 +.084 2.199+.081 .044+ .002
.831 +.139 2.250+.109 .045+.003
.603+.066 2.234+.098 .046+ .002
The birds in all trials were sacrificed by injecting intravenously an overdose of Nembutal.
Downloaded from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015
The present experiments were designed to test a bovine growth hormone with no adrenocorticotrophic activity detectable at doses up to 4 mg. per 100 gm. of body weight and to determine the interaction of bovine growth hormone with desoxycorticosterone acetate and cortisone acetate on growth of body and certain glands. All references to growth hormone in this paper, unless otherwise specified will refer to bovine growth hormone.
HORMONE INJECTIONS AND GLAND WEIGHTS
1529
TABLE 3.—The effect of desoxycorticosterone acetate on body weight gain of 9-23-day-old chickens1 Treatments Age (12 birds per treatment)
Sesame oil
9-16-day-old, gm. 16-23-day-old, gm.
2
DOCA
DOCA
.2 ml./injection
0
96 + 2.7
106+2.0
98 + 2.5
100 + 2.0
101 + 1.7
104+3.2
115 + 3.7
91 + 5.2
98 + 7.7
86±4.5
137 + 3.0
131 + 3.7
124 + 5.4
105 + 8.2
89 + 8.7
.15 ml./inj. (1.5 mg.)
.25 ml./inj. (2.5 mg.)
All means not underscored by the same line are significantly different at the 5% level, Duncan (1955). Weight gain.
termined by the method of Whitehorn (1921) and Schales and Schales (1941). Salmon color was used as the end point for the Whitehorn method since the color dictionary of Maerz and Paul (1950) did not show salmon-red. The use of salmon in place of salmon-red might explain the relatively high chloride content reported in Table 6 for the 23-day-old birds. RESULTS Growth hormone did not significantly affect gain in body weight or size of the bursa of Fabricius, heart, and adrenal glands (Tables 1 and 2). The largest adrenals, heart, and bursae were found in the growth hormone groups. The 1.5 and 2.5 mg. levels of desoxycorticosterone acetate (DOCA) significantly reduced body weight
gains from 16-23 days of age (Table 3). The weight of the bursa and adrenal glands were reduced by DOCA. However, DOCA at the 1.5 and 2.5 mg. levels significantly increased the size of the heart (Table 4). Growth hormone and DOCA alone or in combination did not significantly affect gain in body weight while cortisone significantly reduced gains (Table 5). The catabolic influence of cortisone on body weight appeared to be modified by the addition of growth hormone. It is apparent from the data of Table 5 that the inhibitory action of cortisone and DOCA on the bursa appear to be slightly modified by the presence of growth hormone. Cortisone and DOCA alone or in combination significantly reduced the size of the adrenal gland when compared with the adrenals of birds re-
TABLE 4.—The effect of desoxycorticosterone acetate on the weight of the bursa of Fabricius, heart, and adrenal glands in 23-day-old chickens1Treatments Gland (12 birds/treat.)
Bursa of Fabricius, gm. Heart, gm. Adrenal, gm. 1
S e s a m e oil
No injection
D 0 C A
DOCA
DOCA
.2 ml./injection
0
.05 ml./inj. (.5 mg.)
.15 ml./inj. (1.5 mg.)
.25 ml./inj. (2.5 mg.)
.938+.069
1.153+.092
. 8 7 6 + . 087
.757+.073
.684+ .005
2.051 + .074
2.305+.144
2.179+.140
2.982+.576
2.682+.233
.043+.007
.043±.009
.040+.008
.034+.009
.042+.010
All means not underscored by the same line are significantly different at the 5 % level, Duncan (1955).
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1 2
DOCA .05 ml./inj. (.5 mg.)
9-day-old (initial wt.), gm. 2
No injection
3
2
1
.029+.006
.034+.005
.029+.001
.028+.005
Oil
GHand DOCA
64+10
Saline
.408+.03
GHand Cortisone
.028+.007
Control
1.852+ .17
GHand DOCA
GHand DOCA and Cortisone 1.906+.47
.024+.072
GHand Cortisone
.329+.10
DOCA
55±4
GHand Cortisone
.026+ .014
GHand DOCA and Cortisone
Treatments
.533+.09
GHand DOCA
Treatments
70 + 9
Control
All means not underscored by the same line are significantly different at the 5 % level, Duncan (1955). Ml. per injection: oil, .2 ml.; saline, .4 ml.; GH, .4 ml.; DOCA, .2 ml.; cortisone, .05 ml. Six birds per treatment.
Heart
Adrenal, gm.
Saline
GH
.551+.23
.581 + .16
Saline
Control
GH
Oil
. 6 9 6 + . 13
78 + 20
DOCA
79±4
GH
82 + 19
GHand DOCA
82 + 3
Bursa of Fabricius, gm. . 8 2 3 + . 17
Gain in body weight, gm.
Oil
Treatments
1.579+.37
DOCA and Cortisone
Treatments
. 0 2 3 + . 003
DOCA
.244+.08
Cortisone
52+4
GHand DOCA and Cortisone
1.338+.13
GH and Cortisone
.022+.003
DOCA and Cortisone
.220+.03
GHand DOCA and Cortisone
42 + 10
DOCA and Cortisone
TABLE 5.—The effect of growth hormone (GH), desoxycorticosterone acetate (DOCA), and cortisone on gain in body weight, and the weight of the bursa of Fabricius, adrenal and heart of 18-day-old chickens1 •*>*
from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015 1.289+.21
Control
.O17+.O02
Cortisone
.215+.08
DOCA and Cortisone
38+18
Cortisone
o
o
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HORMONE INJECTIONS AND GLAND WEIGHTS TABLE 6.—The effect of growth hormone, desoxycorticosterone, and cortisone on the blood chlorides of young chickens1 Treatments No injection
23-day-old
2
Growth hormone
0
.2 ml./inj.
.6 ml./inj.
.4 ml./inj.
130.1 + 3.6
131.1 + 3.7
134.0 + 12.0
1131.1 + 2.8
No injection
Sesame oil
Desoxycorticosterone
0
.05 ml./inj.
.15 ml./inj.
.25 ml./inj.
.20 ml./inj.
128.0 + 37
136.5+.10
135.0+ .27
137.8+.80
135.0+ .27
No injection Oil
Saline
0 18-day-old 1 2 3
3
118.0 + 5.8
118.5 + 4.4
110.8 + 9
Growth hormone
Desoxycorti costerone
Cortisone
112.1 + 10.0 124.6 + 5.5 119.3 + 6.6
All values represent milliequivalents of chloride per liter. Method of Whitehorn (1921). Method of Schales and Schales (1941).
ceiving growth hormone, saline, growth hormone plus DOCA, oil, and no injections (Table 5). In addition to a significantly larger adrenal in the presence of growth hormone, growth hormone appeared to counteract the depressive effect on the adrenal gland of both cortisone and DOCA. Heart size was significantly increased when growth hormone and DOCA were administered together (Table 5). There was a slight but consistent increase in blood chloride following DOCA injections (Table 6). This is similar to the findings of Selye (1942). DISCUSSION
The inability of bovine growth hormone in this study to stimulate body growth of young chickens might indicate a species resistance as exists in rats to fish growth hormone, Pickford (1954), and monkeys to beef and hog growth hormone, Knobil and Greep (1956). It also might indicate that in chickens the true growth hormone is a hormone other than the growth or somatotrophic hormone. According to Huble (1956), prolactin might play the role of
growth hormone in chickens. However, the results of this paper indicate that bovine growth hormone is not without its effect in chickens. For example, the adrenals, heart, and bursa of Fabricius appeared to be stimulated by the growth hormone injections. The enlarged adrenals in the presence of growth hormone might suggest an increased release of adrenocorticotrophic hormone by the anterior pituitary. The action of growth hormone then would be that of a stressor agent. However, it has been demonstrated by Garren and Shaffner (1956) that in chickens enlarged adrenals brought about by stressor agents will result in a reduction in the weight of the bursa of Fabricius. The bursa is known to regress in the presence of exogenous injections of adrenocorticotrophic hormone and cortisone, Huble (1958) and Glick (1957). The limited data, then, indicate that growth hormone did not act by way of the pituitary, but may have acted directly on the adrenal glands without causing an increased release of glucocorticoids. According to Selye (1942), an overdosage of desoxycorticosterone in chicks pro-
Downloaded from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015
23-day-old
2
Saline
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B. GLICK
SUMMARY
Bovine growth hormone and desoxycorticosterone acetate were administered intramuscularly in two separate trials from the ninth to the twenty-third day of age. In a third trial, bovine growth hormone, desoxycorticosterone, and cortisone were administered intramuscularly alone and in combi-
nation from the twelfth to the eighteenth day of age. Growth hormone alone did not influence body weight. However, the catabolic influence of cortisone on body weight appeared to be modified by the addition of bovine growth hormone. The inhibitory action of desoxycorticosterone and cortisone on the bursa of Fabricius was slightly modified by the presence of growth hormone. Significantly larger adrenals were found in birds receiving growth hormone. Growth hormone appeared to counter-act the depressive effect on the adrenal glands from both cortisone and desoxycorticosterone. The data suggest that bovine growth hormone in chickens acts directly on the adrenal gland rather than by way of the anterior pituitary. A significant increase in heart size resulted from the presence of desoxycorticosterone and desoxycorticosterone plus growth hormone. Also, the effects of growth hormone and cortisone in the chicken appeared to be antagonistic. ACKNOWLEDGMENT
The bovine growth hormone used in this work was a generous gift from the Endocrinology Study Section, National Institute' of Health. The Cortone, a saline suspension of cortisone acetate, was kindly donated by Dr. Laurent Michaud of Merck and Co. The assistance of Mrs. Rosemary Harris and Mrs. Bobbie Brashear, laboratory technicians, in the collection of the data presented in this paper was sincerely appreciated. REFERENCES Beck, J. C , E. E. McGarry, I. Dyrenfurth and E. H. Vanning, 1957. Metabolic effects of human and monkey growth hormone in man. Science, 125: 884-885. Carter, R. D., R. N. Risner and J. H. Yacowitz, 1955. Some effects of growth hormone preparations in pullets and mature hens. Poultry Sci. 34: 1407-1415. Duncan, D. B., 1955. Multiple range and multiple F test. Biometrics, 11: 1-42. Garren, H. W., and C. S. Shaffner, 1956. How
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duced an enlarged heart. Heart size was also increased in this experiment in the presence of desoxycorticosterone. In addition, growth hormone increased heart size. Growth hormone in combination with desoxycorticosterone and in combination with both desoxycorticosterone and cortisone significantly increased heart size above the controls. On the other hand, when cortisone was injected with either desoxycorticosterone or growth hormone, heart size was not significantly different from the controls. Unpublished data in this laboratory revealed that heart size was reduced in the presence of cortisone. The data, then, indicate that the effects of desoxycorticosterone and growth hormone on the heart are in opposition to the effects of cortisone. According to Glick (1957), cortisone has an inhibitory effect on the weight of the body, bursa, and adrenal glands of chickens. The data of the present experiments indicate that the adverse effects of cortisone on growth, the bursa of Fabricius, and adrenal glands can be partially counter-acted by simultaneous administration with growth hormone. Therefore, under the conditions of this experiment, bovine growth hormone and cortisone appeared to be antagonistic in the young chicken. Sobel (1958) has reported that growth hormone and cortisone are antagonistic in the chick embryo. Before a definite statement can be made concerning the effect of growth hormone on young chickens, the growth hormone should be administered to hypophysectomized chickens. Such a study will be undertaken.
HORMONE INJECTIONS AND GLAND WEIGHTS
tomized male killifish to purified fish growth hormone as compared with the response to purified beef growth hormone. Endocrinology, SS: 274. Schales, O., and S. S. Schales, 1941. Simple and accurate method for the determination of chloride in biological fluid. J. Biol. Chem. 140: 879884. Selye, H., 1942. Production of nephrosclerosis by overdosage with desoxycorticosterone acetate. Canad. Med. Assoc. J. 47: 515-519. Selye, H., 19S5. The hypophyseal growth hormone: Nature and Actions. R. W. Smith, Jr., O. H. Gaebler, C. N. H. Long, Eds. Blakiston, New York. p. 123. Selye, H., and G. Heuser, 19S4. Fourth Annual Report on Stress. Acta, Inc., Montreal, pp. S7-63. Snedecor, G. W., 1946. Statistical Methods. Iowa State College Press, Ames, Iowa. Sobel, H., 19S8. Antagonistic effects of cortisone and growth hormone on the developing chick embryo. Proc. Soc. Expt. Biol. Med. 97: 495498. Whitehorn, J. C , 1921. Simplified method for the determination of chlorides in blood or plasma. A system of blood analysis. Supplement II. J. Biol. Chem. 45: 449-461. Wilhelmi, A. E., 19SS. The hypophyseal growth hormone: Nature and Actions. R. W. Smith, Jr., O. H. Gaebler, C. N. H. Long, Eds. Blakiston, New York. p. S9.
Estimation of Genetic Diversity by Skin-graft Reactions in Young Chicks1'2 J. V. CRAIG, C. R. POLLEY AND S. WEARDEN3 Kansas State University, Manhattan, Kansas (Received for publication March 8. 1960)
INTRODUCTION
S
EVERAL methods of estimating genetic relationships between donor and host groups by means of reactions to transplanted tissue have met with limited sucJ
This investigation was part of the Kansas contribution to the NC-47 Regional Poultry Breeding Project. 2 Contribution No. 2S4, Department of Poultry Husbandry and Contribution No. SO, Department
cess. Among these, the relative-lymphocyteincrease technique, following tissue grafting, was shown to be useful where known relationship coefficients were involved within of Statistics, Kansas Agricultural Experiment Station, Manhattan. 3 Associate Professor and Graduate Research Assistant in Poultry Genetics, Department of Poultry Husbandry, and Associate Professor of Statistics, respectively.
Downloaded from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015
the period of exposure to different stress stimuli affects the endocrine and lymphatic gland weights of young chickens. Poultry Sci. 3 5 : 266-273. Glick, B., 1957. Experimental modification of the growth of the bursa of Fabricius. Poultry Sci. 36: 18-23. Hsieh, K.-M., T.-Y. Wang and H. Blumenthal, 1952. The diabetogenic and growth promoting activities of growth hormone (somatotropin) in the developing chick embryo. Endocrinology, S i : 298-302. Huble, J., 19S6. Gonadal and hypophyseal interactions on the young fowl. Acta Endocrinologica, 23: 101-104. Huble, J., 1958. Effects of hormones on endocrine and lympho-epithelial glands in young fowl. Poultry Sci. 37: 297-301. Knobil, E., 1955. The hypophyseal growth hormone. Nature and Actions, R. W. Smith, Jr., O. H. Gaebler, C. N. H. Long, Eds. Blakiston, New York, p. 2SS. Knobil, E. and R. O. Greep, 19S6. Physiological effects of growth hormone of primate origin in the hypophysectomized rhesus monkey. Fed. Proc. I S : 111-112. Libby, D. A., J. Meites and P. J. Schaible, 19SS. Growth hormone effects in chickens. Poultry Sci. 34: 1329-1331. Maerz, A., and M. R. Paul, 19S0. Dictionary of Color. McGraw-Hill Book Co., Inc. Pickford, G. E., 19S4. The response of hyphosec-
1533
min Ba, activity assay. Fifteenth edition, pp. 885-889. Robinson, R. A., M. E. Fitzgerald, K. Fehr and J. J. Grimshaw, 1954. Vitamin B ^ in crude liver extracts. Nature, 174: 558-559. Sehiya, S., 1953. Jap. J. Pharmacol. 3 : 1. Quoted by Fischer et al. (1958). Skarzynski, B., Z. Zak, W. Ostrowski, M. K. Davies, M. E. Gregory and S. K. Kon, 1959. Vitamin Bis in the developing chick embryo. Brit. J. Nutrition, 13: 72-78. Snell, E. E., and E. Quarles, 1941. The effect of incubation on the vitamin content of eggs. J. Nutrition, 22: 483-489. Swendseid, M. E., R. Jackson and E. M. Benson, 1958. The formation of citrovorum factor in the developing chick egg and the effect of injecting folic' acid antagonists. Arch. Biochem. Bioph. 78: 221-226. Tastaldi, H., 1959. Ensaio microbiologico da vitamina BM. Praticas de Quimica Biologica, 2nd part, 4th edition, pp. 194-199. Woolley, W., 1954. Synthesis of vitamin Bis by spontaneous cancers. Federation Proc. 13: 482483. Yacowitz, H., C. H. Hill, L. C. Norris and G. F. Heuser, 1952. Distribution of vitamin B« in the organs and tissue of the chick. Proc. Soc. Exptl. Biol. Med. 79: 279-280.
The Effect of Bovine Growth Hormone, Desoxycorticosterone, and Cortisone on the Weight of the Bursa of Fabricius, Adrenal Glands, Heart, and Body Weight of Young Chickens BRUCE G L I C K
Department of Poultry Husbandry, Mississippi State University, State College, Mississippi (Received for publication March 8, 1960)
R
ESEARCH to date has revealed little or no effect of growth hormone in young chickens, Carter et al. (1955) and Libby et al. (1955). Hsieh et al. (1952) have reported a growth promoting and diabetogenic effect of growth hormone in chick embryos. Guinea pigs and man appear to be insensitive to bovine growth hormone, Selye (1955), while the data of Beck et al. Journal Article No. 807
(1957) indicate a slight but significant growth effect of human and monkey growth hormone on a 13-year-old human male. Lack of stimulation by growth hormone may be due to contamination with adrenocorticotropic hormone, to treatment procedure, to species differences, Wilhelmi (1955) and Knobil (1955), or to "conditioning" of tissue to growth hormone by the adrenal glands, Selye (1955) and Selye and Heuser (1954).
Downloaded from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015
cient chick embryos. A.M.A. Arch. Pathol. 60: 393-400. Fischer, N. A., E. M. Benson and M. E. Swendseid, 19S8. The distribution of vitamin Bis in the developing chick egg. Arch. Biochem. Bioph. 74: 458-464. Hayes, M., and T. P. Luckey, 1957. Effect of incubation upon folic acid content of eggs. Proc. Soc. Exptl. Biol. Med. 94: 777-778. Jackson, J. I., L. J. Machlin, E. A. Brandenburger, W. L. Kellog and C. A. Denton, 1953. Retention of Co60 labeled vitamin Bi2 in chickens. Proc. Soc. Exptl. Biol. Med. 83: 221-222. Krieger, C. H., 1952. Report on vitamin Bu,. microbiological method. J. Ass. Off. Agr. Chem. 35: 726-736. Lillie, R. J., M. W. Olsen and H. R. Bird, 1949. Role of vitamin B« in reproduction of poultry. Proc. Soc. Exptl. Biol. Med. 72: 598-602. Mariakulandai, A., and J. McGinnis, 1953. The vitamin B^ requirement for hatchability of chicken eggs. Poultry Sci. 32: 3-7. Nogueira, D. M., 1960. Vitamin Bis activity in chick egg. In press. Olcese, O., J. R. Couch, J. H. Quisenberry and P. B. Pearson, 1950. Congenital anomalies in the chick due to vitamin Bis deficiency. J. Nutrition. 4 1 : 423-431. Pharmacopoeia of the United States, 1955. Vita-
1527
1528
B. GLICK TABLE 1.—The effect of growth hormone on body weight gain of 9-23-day-old chickens Treatments
Age (16 birds per group)
9-day-old (initial wt.), gm. 9-16-day-old, gm.1 16-23-day-old, gm.1 1
No injection
Saline
0
.4 ml./injection
.2 ml./inj. (2 mg.)
Growth hormone .6 ml./inj. (6 mg.)
101 + 2.1 89+3.0 113 + 3.7
107 + 2.1 93 + 3.0 105 + 2.7
106 + 1.6 93 + 3.7 107 + 5.5
107 + 2.0 89 + 2.7 107 + 5.0
Weight gain.
METHOD The effect of growth hormone, desoxycorticosterone acetate, and a combination of growth hormone (GH), desoxycorticosterone acetate (DOCA), and cortisone on the growth of the body, bursa of Fabricius, adrenals, and heart was studied in 1-3-weekold New Hampshire chickens. All birds were raised in batteries and fed a basal ration. Growth hormone in the form of a dry powder was taken up in physiological
saline, allowing each milliliter of solution to contain 10 mg. of the hormone. The desoxycorticosterone acetate was suspended in sesame oil to allow each milliliter of solution to contain 10 mg. of the hormone. Each milliliter of the saline suspension of cortone contained 50 mg. of cortisone. Growth hormone and saline and desoxycorticosterone acetate and oil were administered intramuscularly from the ninth to the twenty-third day of age, respectively. All treatments were replicated. Growth hormone (.4 ml.), desoxycorticosterone (.2 ml.), cortisone (.05 ml.), saline (.4 ml.), and sesame oil (.2 ml.) were administered intramuscularly from the twelfth to the eighteenth day of age. Equal numbers of males and females were used in all trials. Body and gland weights were analyzed by the analysis of variance and covariance, respectively, Snedecor (1946) Duncan's (1955) new multiple range and F test was utilized when determining which means differed significantly. Blood chlorides were de-
TABLE 2.—The eject of growth hormone on the weight of the bursa of Fabricius, heart, and adrenal glands of 23-day-old chickens1 Treatments Glands, grams (12 birds per treat.)
Bursa of Fabricius Heart Adrenal 1
No injection
Saline
Growth hormone
0
.2 ml./injection
.2 ml./inj. (2 mg.) .6 ml./inj. (6 mg.)
.654+ .067 2.089+.012 .040+ .002
.591 +.084 2.199+.081 .044+ .002
.831 +.139 2.250+.109 .045+.003
.603+.066 2.234+.098 .046+ .002
The birds in all trials were sacrificed by injecting intravenously an overdose of Nembutal.
Downloaded from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015
The present experiments were designed to test a bovine growth hormone with no adrenocorticotrophic activity detectable at doses up to 4 mg. per 100 gm. of body weight and to determine the interaction of bovine growth hormone with desoxycorticosterone acetate and cortisone acetate on growth of body and certain glands. All references to growth hormone in this paper, unless otherwise specified will refer to bovine growth hormone.
HORMONE INJECTIONS AND GLAND WEIGHTS
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TABLE 3.—The effect of desoxycorticosterone acetate on body weight gain of 9-23-day-old chickens1 Treatments Age (12 birds per treatment)
Sesame oil
9-16-day-old, gm. 16-23-day-old, gm.
2
DOCA
DOCA
.2 ml./injection
0
96 + 2.7
106+2.0
98 + 2.5
100 + 2.0
101 + 1.7
104+3.2
115 + 3.7
91 + 5.2
98 + 7.7
86±4.5
137 + 3.0
131 + 3.7
124 + 5.4
105 + 8.2
89 + 8.7
.15 ml./inj. (1.5 mg.)
.25 ml./inj. (2.5 mg.)
All means not underscored by the same line are significantly different at the 5% level, Duncan (1955). Weight gain.
termined by the method of Whitehorn (1921) and Schales and Schales (1941). Salmon color was used as the end point for the Whitehorn method since the color dictionary of Maerz and Paul (1950) did not show salmon-red. The use of salmon in place of salmon-red might explain the relatively high chloride content reported in Table 6 for the 23-day-old birds. RESULTS Growth hormone did not significantly affect gain in body weight or size of the bursa of Fabricius, heart, and adrenal glands (Tables 1 and 2). The largest adrenals, heart, and bursae were found in the growth hormone groups. The 1.5 and 2.5 mg. levels of desoxycorticosterone acetate (DOCA) significantly reduced body weight
gains from 16-23 days of age (Table 3). The weight of the bursa and adrenal glands were reduced by DOCA. However, DOCA at the 1.5 and 2.5 mg. levels significantly increased the size of the heart (Table 4). Growth hormone and DOCA alone or in combination did not significantly affect gain in body weight while cortisone significantly reduced gains (Table 5). The catabolic influence of cortisone on body weight appeared to be modified by the addition of growth hormone. It is apparent from the data of Table 5 that the inhibitory action of cortisone and DOCA on the bursa appear to be slightly modified by the presence of growth hormone. Cortisone and DOCA alone or in combination significantly reduced the size of the adrenal gland when compared with the adrenals of birds re-
TABLE 4.—The effect of desoxycorticosterone acetate on the weight of the bursa of Fabricius, heart, and adrenal glands in 23-day-old chickens1Treatments Gland (12 birds/treat.)
Bursa of Fabricius, gm. Heart, gm. Adrenal, gm. 1
S e s a m e oil
No injection
D 0 C A
DOCA
DOCA
.2 ml./injection
0
.05 ml./inj. (.5 mg.)
.15 ml./inj. (1.5 mg.)
.25 ml./inj. (2.5 mg.)
.938+.069
1.153+.092
. 8 7 6 + . 087
.757+.073
.684+ .005
2.051 + .074
2.305+.144
2.179+.140
2.982+.576
2.682+.233
.043+.007
.043±.009
.040+.008
.034+.009
.042+.010
All means not underscored by the same line are significantly different at the 5 % level, Duncan (1955).
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1 2
DOCA .05 ml./inj. (.5 mg.)
9-day-old (initial wt.), gm. 2
No injection
3
2
1
.029+.006
.034+.005
.029+.001
.028+.005
Oil
GHand DOCA
64+10
Saline
.408+.03
GHand Cortisone
.028+.007
Control
1.852+ .17
GHand DOCA
GHand DOCA and Cortisone 1.906+.47
.024+.072
GHand Cortisone
.329+.10
DOCA
55±4
GHand Cortisone
.026+ .014
GHand DOCA and Cortisone
Treatments
.533+.09
GHand DOCA
Treatments
70 + 9
Control
All means not underscored by the same line are significantly different at the 5 % level, Duncan (1955). Ml. per injection: oil, .2 ml.; saline, .4 ml.; GH, .4 ml.; DOCA, .2 ml.; cortisone, .05 ml. Six birds per treatment.
Heart
Adrenal, gm.
Saline
GH
.551+.23
.581 + .16
Saline
Control
GH
Oil
. 6 9 6 + . 13
78 + 20
DOCA
79±4
GH
82 + 19
GHand DOCA
82 + 3
Bursa of Fabricius, gm. . 8 2 3 + . 17
Gain in body weight, gm.
Oil
Treatments
1.579+.37
DOCA and Cortisone
Treatments
. 0 2 3 + . 003
DOCA
.244+.08
Cortisone
52+4
GHand DOCA and Cortisone
1.338+.13
GH and Cortisone
.022+.003
DOCA and Cortisone
.220+.03
GHand DOCA and Cortisone
42 + 10
DOCA and Cortisone
TABLE 5.—The effect of growth hormone (GH), desoxycorticosterone acetate (DOCA), and cortisone on gain in body weight, and the weight of the bursa of Fabricius, adrenal and heart of 18-day-old chickens1 •*>*
from http://ps.oxfordjournals.org/ at Grand Valley State University on June 9, 2015 1.289+.21
Control
.O17+.O02
Cortisone
.215+.08
DOCA and Cortisone
38+18
Cortisone
o
o
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HORMONE INJECTIONS AND GLAND WEIGHTS TABLE 6.—The effect of growth hormone, desoxycorticosterone, and cortisone on the blood chlorides of young chickens1 Treatments No injection
23-day-old
2
Growth hormone
0
.2 ml./inj.
.6 ml./inj.
.4 ml./inj.
130.1 + 3.6
131.1 + 3.7
134.0 + 12.0
1131.1 + 2.8
No injection
Sesame oil
Desoxycorticosterone
0
.05 ml./inj.
.15 ml./inj.
.25 ml./inj.
.20 ml./inj.
128.0 + 37
136.5+.10
135.0+ .27
137.8+.80
135.0+ .27
No injection Oil
Saline
0 18-day-old 1 2 3
3
118.0 + 5.8
118.5 + 4.4
110.8 + 9
Growth hormone
Desoxycorti costerone
Cortisone
112.1 + 10.0 124.6 + 5.5 119.3 + 6.6
All values represent milliequivalents of chloride per liter. Method of Whitehorn (1921). Method of Schales and Schales (1941).
ceiving growth hormone, saline, growth hormone plus DOCA, oil, and no injections (Table 5). In addition to a significantly larger adrenal in the presence of growth hormone, growth hormone appeared to counteract the depressive effect on the adrenal gland of both cortisone and DOCA. Heart size was significantly increased when growth hormone and DOCA were administered together (Table 5). There was a slight but consistent increase in blood chloride following DOCA injections (Table 6). This is similar to the findings of Selye (1942). DISCUSSION
The inability of bovine growth hormone in this study to stimulate body growth of young chickens might indicate a species resistance as exists in rats to fish growth hormone, Pickford (1954), and monkeys to beef and hog growth hormone, Knobil and Greep (1956). It also might indicate that in chickens the true growth hormone is a hormone other than the growth or somatotrophic hormone. According to Huble (1956), prolactin might play the role of
growth hormone in chickens. However, the results of this paper indicate that bovine growth hormone is not without its effect in chickens. For example, the adrenals, heart, and bursa of Fabricius appeared to be stimulated by the growth hormone injections. The enlarged adrenals in the presence of growth hormone might suggest an increased release of adrenocorticotrophic hormone by the anterior pituitary. The action of growth hormone then would be that of a stressor agent. However, it has been demonstrated by Garren and Shaffner (1956) that in chickens enlarged adrenals brought about by stressor agents will result in a reduction in the weight of the bursa of Fabricius. The bursa is known to regress in the presence of exogenous injections of adrenocorticotrophic hormone and cortisone, Huble (1958) and Glick (1957). The limited data, then, indicate that growth hormone did not act by way of the pituitary, but may have acted directly on the adrenal glands without causing an increased release of glucocorticoids. According to Selye (1942), an overdosage of desoxycorticosterone in chicks pro-
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23-day-old
2
Saline
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B. GLICK
SUMMARY
Bovine growth hormone and desoxycorticosterone acetate were administered intramuscularly in two separate trials from the ninth to the twenty-third day of age. In a third trial, bovine growth hormone, desoxycorticosterone, and cortisone were administered intramuscularly alone and in combi-
nation from the twelfth to the eighteenth day of age. Growth hormone alone did not influence body weight. However, the catabolic influence of cortisone on body weight appeared to be modified by the addition of bovine growth hormone. The inhibitory action of desoxycorticosterone and cortisone on the bursa of Fabricius was slightly modified by the presence of growth hormone. Significantly larger adrenals were found in birds receiving growth hormone. Growth hormone appeared to counter-act the depressive effect on the adrenal glands from both cortisone and desoxycorticosterone. The data suggest that bovine growth hormone in chickens acts directly on the adrenal gland rather than by way of the anterior pituitary. A significant increase in heart size resulted from the presence of desoxycorticosterone and desoxycorticosterone plus growth hormone. Also, the effects of growth hormone and cortisone in the chicken appeared to be antagonistic. ACKNOWLEDGMENT
The bovine growth hormone used in this work was a generous gift from the Endocrinology Study Section, National Institute' of Health. The Cortone, a saline suspension of cortisone acetate, was kindly donated by Dr. Laurent Michaud of Merck and Co. The assistance of Mrs. Rosemary Harris and Mrs. Bobbie Brashear, laboratory technicians, in the collection of the data presented in this paper was sincerely appreciated. REFERENCES Beck, J. C , E. E. McGarry, I. Dyrenfurth and E. H. Vanning, 1957. Metabolic effects of human and monkey growth hormone in man. Science, 125: 884-885. Carter, R. D., R. N. Risner and J. H. Yacowitz, 1955. Some effects of growth hormone preparations in pullets and mature hens. Poultry Sci. 34: 1407-1415. Duncan, D. B., 1955. Multiple range and multiple F test. Biometrics, 11: 1-42. Garren, H. W., and C. S. Shaffner, 1956. How
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duced an enlarged heart. Heart size was also increased in this experiment in the presence of desoxycorticosterone. In addition, growth hormone increased heart size. Growth hormone in combination with desoxycorticosterone and in combination with both desoxycorticosterone and cortisone significantly increased heart size above the controls. On the other hand, when cortisone was injected with either desoxycorticosterone or growth hormone, heart size was not significantly different from the controls. Unpublished data in this laboratory revealed that heart size was reduced in the presence of cortisone. The data, then, indicate that the effects of desoxycorticosterone and growth hormone on the heart are in opposition to the effects of cortisone. According to Glick (1957), cortisone has an inhibitory effect on the weight of the body, bursa, and adrenal glands of chickens. The data of the present experiments indicate that the adverse effects of cortisone on growth, the bursa of Fabricius, and adrenal glands can be partially counter-acted by simultaneous administration with growth hormone. Therefore, under the conditions of this experiment, bovine growth hormone and cortisone appeared to be antagonistic in the young chicken. Sobel (1958) has reported that growth hormone and cortisone are antagonistic in the chick embryo. Before a definite statement can be made concerning the effect of growth hormone on young chickens, the growth hormone should be administered to hypophysectomized chickens. Such a study will be undertaken.
HORMONE INJECTIONS AND GLAND WEIGHTS
tomized male killifish to purified fish growth hormone as compared with the response to purified beef growth hormone. Endocrinology, SS: 274. Schales, O., and S. S. Schales, 1941. Simple and accurate method for the determination of chloride in biological fluid. J. Biol. Chem. 140: 879884. Selye, H., 1942. Production of nephrosclerosis by overdosage with desoxycorticosterone acetate. Canad. Med. Assoc. J. 47: 515-519. Selye, H., 19S5. The hypophyseal growth hormone: Nature and Actions. R. W. Smith, Jr., O. H. Gaebler, C. N. H. Long, Eds. Blakiston, New York. p. 123. Selye, H., and G. Heuser, 19S4. Fourth Annual Report on Stress. Acta, Inc., Montreal, pp. S7-63. Snedecor, G. W., 1946. Statistical Methods. Iowa State College Press, Ames, Iowa. Sobel, H., 19S8. Antagonistic effects of cortisone and growth hormone on the developing chick embryo. Proc. Soc. Expt. Biol. Med. 97: 495498. Whitehorn, J. C , 1921. Simplified method for the determination of chlorides in blood or plasma. A system of blood analysis. Supplement II. J. Biol. Chem. 45: 449-461. Wilhelmi, A. E., 19SS. The hypophyseal growth hormone: Nature and Actions. R. W. Smith, Jr., O. H. Gaebler, C. N. H. Long, Eds. Blakiston, New York. p. S9.
Estimation of Genetic Diversity by Skin-graft Reactions in Young Chicks1'2 J. V. CRAIG, C. R. POLLEY AND S. WEARDEN3 Kansas State University, Manhattan, Kansas (Received for publication March 8. 1960)
INTRODUCTION
S
EVERAL methods of estimating genetic relationships between donor and host groups by means of reactions to transplanted tissue have met with limited sucJ
This investigation was part of the Kansas contribution to the NC-47 Regional Poultry Breeding Project. 2 Contribution No. 2S4, Department of Poultry Husbandry and Contribution No. SO, Department
cess. Among these, the relative-lymphocyteincrease technique, following tissue grafting, was shown to be useful where known relationship coefficients were involved within of Statistics, Kansas Agricultural Experiment Station, Manhattan. 3 Associate Professor and Graduate Research Assistant in Poultry Genetics, Department of Poultry Husbandry, and Associate Professor of Statistics, respectively.
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the period of exposure to different stress stimuli affects the endocrine and lymphatic gland weights of young chickens. Poultry Sci. 3 5 : 266-273. Glick, B., 1957. Experimental modification of the growth of the bursa of Fabricius. Poultry Sci. 36: 18-23. Hsieh, K.-M., T.-Y. Wang and H. Blumenthal, 1952. The diabetogenic and growth promoting activities of growth hormone (somatotropin) in the developing chick embryo. Endocrinology, S i : 298-302. Huble, J., 19S6. Gonadal and hypophyseal interactions on the young fowl. Acta Endocrinologica, 23: 101-104. Huble, J., 1958. Effects of hormones on endocrine and lympho-epithelial glands in young fowl. Poultry Sci. 37: 297-301. Knobil, E., 1955. The hypophyseal growth hormone. Nature and Actions, R. W. Smith, Jr., O. H. Gaebler, C. N. H. Long, Eds. Blakiston, New York, p. 2SS. Knobil, E. and R. O. Greep, 19S6. Physiological effects of growth hormone of primate origin in the hypophysectomized rhesus monkey. Fed. Proc. I S : 111-112. Libby, D. A., J. Meites and P. J. Schaible, 19SS. Growth hormone effects in chickens. Poultry Sci. 34: 1329-1331. Maerz, A., and M. R. Paul, 19S0. Dictionary of Color. McGraw-Hill Book Co., Inc. Pickford, G. E., 19S4. The response of hyphosec-
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