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The influence of exogenous ACTH on the levels of corticosterone and cortisol in the plasma of young chicks (Gallus domesticus)
GENERAL
AND
COMPARATIVE
ENDOCRINOLOGY
44, 249-251 (1981)
NOTES T
~nflue~~~ oatisol
of Exogenous in the Plasma
ACTH on of Young
The aim of this study was to determine the influence of ACTH on the levels of corticosterone and cortisol in the plasma of posthatching chicks. Chicks were injected with IO 1-U of ACTH daily from 3 to 17 days of age. Plasma, collected on Days 4, IO. and 17, was separated by thin-layer chromatography and analyzed by radioimmunoassay for the presence of cortisol and corticosterone. Corticosterone levels from control chicks were significantly lower at 4 days (5.0 &ml) than at 10 or 17 days. Cortisol levels were very low at first (Less than I @ml) by 4 days and disappeared by Day 17. Under the influence of ACTH, both corticosterone and cortisol increased significantly at 4 and 10 days indicating that AC’FH was not selectively acting on only one pathway. However, by 17 days, the levels of corticosterone and cortisol were similar to those of control values thus indicating an apparent failure of the ACTH response.
The predominance of corticosterone and the absence or relatively low levels of cortisol in the plasma and adrenals of adult birds (see review of Kalliecharan and Hall, 1974) resulted in later studies being directed primarily toward determining the presence of corticosterone and its intermediates. oos and deRoos (1964) and demonstrated that both TH and an acid extract of enohypophysis can significantly increase both corticosterone and alosterone ‘on in vitro. Recently, owever, ura et al. (1978) have at the 17-hydroxylase activity that is necessary for cortisol formation in chick embryos (Kahiecharan and Hall, 1977; Idler ek al., 1976) disappears from the adrenal lands of 7- to 14”day-old posthatching hicks. In view of this, porcine ACTH was tested thatching chicks to determine it can influence the biosynthetic s of corticosterone and cortisol in the same manner as reported in the in vitro ryonic chick adrenals (KalHall, 1977) or whether prot can cause cortisol ing steroid as demass et al., 1954). y-old posthatching and onward were injected intramuscularly
perimental
chicks recei
d in the abode gelatin s were wea
minute cortisol ple. Samples
water (98:lQ: 1) mixture
of [3H]coreach were
we
w
249 0016-6480/81/060249-03So1.OO/O Copyright @ 1981 by Academic Press, Inc. AIi rights of mprodwtion in any form resend
250
NOTES
costerone antiserum B21-42 (Endocrine Sciences, Tarzana, Calif.) was used for the determination of corticosterone, whereas a radioimmunoassay kit for cortisol was obtained from New England Nuclear (Boston, Mass.). The concentration of corticosteroids in the plasma of 4-, lo-, and 17-day-old chicks is presented in Table 1, and the difference in weight between Days 3 and 4, 3 and 10, and 3 and 17 is shown in Table 2. The results from Table 1 indicate that the corticosterone level in the plasma of control chicks is low (5 &ml) at 4 days posthatching, yet significantly less than the levels at 10 (8.0 &ml) and 17 (8.1 &ml) days posthatching (PC 0.001). These values correlate well with the increasing activity of the enzyme, 21-hydroxylase that is necessary for corticosterone formation, in 3-, 7-, and 14day-old chick adrenals (Nakamura et d., 1978). The decreasing pattern and total disappearance of cortisol from the plasma of control chicks -(see Table 1) agree with the adrenal gland data from posthatching chicks (Nakamura et al., 1978). The fact that corticosterone and cortisol increased significantly in the plasma of 4and IO-day-old chicks following ACTH treatment (see Table 1) indicates that ACTH is not selectively acting on only one corticosteroid pathway (Bush, 1953; Kalliecharan and Hall, 1977). However, the apparent failure of prolonged ACTH treatment to significantly alter corticosterone and cortisol levels in the plasma of 17-dayold chicks may be due to an inactivation of the ACTH molecule by antibodies formed as a result of the daily injections of ACTH over a prolonged period. The low levels of corticosterone found may reflect basal levels from the adrenal glands in the absence of ACTH stimulation (Woods et al., 1971) and this may account for the significant decrease in body weight (see Table 2). This decrease in body weight and the inability of corticosterone to respond to prolonged ACTH has also been demonstrated by Kass et al., (1954). These authors have
NOTES
BODY
WEIGHT
INCREASES
251
TABLE 2 3 AND 4, 3 AND 10, AND 3 ACTH-TREATED POSTHATCHING CHICKS BETWEEN
DAYS
AND
17 IN CONTROL .\XD
Body weight increase Age of chicks (davs)
Control
4 10 17
5.4 t 0.7 (6)” 45 f 1.1 (6) 130.9 +- 3.1 (5)
3.8 + 0.6 (6) 38.2 k 2.2 (6) 101 i- 5.9 (5)
Note. Values are means + SEM. u Number of animals shown in parentheses. * Not significant at the 5% level. V Statistical comparison of weight differences between control and ACTH-treated
shown that the corticosteroid pathways of rabbits can be altered to secrete primarily cortisol instead of corticosterone following prolonged ACTH treatment. In addition, they reported that the higher cortisol level compensated for the loss in plasma corticosterone. Since, cortisol is present in minute quantities in the plasma of control rabbits (Rass eb al., 1954), the total absence of this steroid from the plasma of 17-day-old contro1 chicks indicates that a genetic block on the enzymes of the C-17 hydroxylating pathway is effective somewhere between and 17. It appears that once this genetic mechanism is activated, possibly would have nb effect on this pathway as shown in Table 1. If there is an alteration of corticosteroid production, as indicated by ass et al. (1954), then, some other steroid may have to fulfil this role, possibly aldosterone which increases in irds following ACTH stimulation (deRoos, 969). CKNQWLEDGMENT Supported by National Sciences and Engineering Research Council of Canada grant (A0460) to R. Kalliecharan.
NS” NS P < O.QI’
animais.
deRoos, R., and deRoos, C. C. (1964). Effects of mammalian corticotropin and chicken adenohypophyseal extracts on steroidogenesis by chicken adrenal tissue in vitro. Gen. Camp. Endocrinol. 4, 602, 607. Idler, D. R., Walsh. J. M.. Kalliecharan, R., and Hall, B. K. (1976). Identification of authentic cortisol in plasma of the embryonic chick. GePz. Comg. Ep?docrinol.
30, 539-540.
Kalliecharan, R., and Hail, B. K. (1974). A developmental study of the levels of progesterone, COB’ticosterone, cortisol, and cortisone circulating in plasma of chick embryos. Gen. Camp. Endocrinol.
24, 364-372.
Kalliecharan, R., and Hall, B. K. (1977). The in vitro biosynthesis of steroids from pregnenolone and cholesterol and the effects of bovine ACTH on corticoid production by adrenal glands of em”bryonic chicks. Gen. Camp. Ertdocritioi. 33, 147-159. Kass, E. H., Hechter, O., Mac&i, 1. A., and Mou, T. W. (1954). Changes in the pattern of secretion of corticosteroids in rabbits after nrolonsed treatment with ACTH. Proc. Sot. Exp. Bioi. k&d. 85, 583-587. Nakamura, T., Tanabe. Y., and Hirano, H. (1978). Evidence of the in vitro formation of cortisol by the adrenai gland of embryonic and young chickens (Gallas domesricus). Gen. Camp. Endocvinot 3.5, 302-308. Woods, J. E., Devries, V. W., and Thommes, R. C. (1971). Ontogenesis of the ~ituita~-adrenal axis in chick embryos. Gcn. Comp. Endocrine!. 17, 407-415.
EFERENCES Bush, I. E. (1953), Species differences in adrenocortiretion. J. Endocrinol. 9, 95- 101. cal deRoos, . (1969). Effects of mammalian corticotropin and progesterone on corticoid production by chicken adrenal tissues in vifro. Gen. Camp. Endocrinol.
13, 455-459.
Department St. Francis Antigonish, Accepted
ofBio!ogy Xavier University B2G ICO, Nova Scotia October 24. 1980
AND
COMPARATIVE
ENDOCRINOLOGY
44, 249-251 (1981)
NOTES T
~nflue~~~ oatisol
of Exogenous in the Plasma
ACTH on of Young
The aim of this study was to determine the influence of ACTH on the levels of corticosterone and cortisol in the plasma of posthatching chicks. Chicks were injected with IO 1-U of ACTH daily from 3 to 17 days of age. Plasma, collected on Days 4, IO. and 17, was separated by thin-layer chromatography and analyzed by radioimmunoassay for the presence of cortisol and corticosterone. Corticosterone levels from control chicks were significantly lower at 4 days (5.0 &ml) than at 10 or 17 days. Cortisol levels were very low at first (Less than I @ml) by 4 days and disappeared by Day 17. Under the influence of ACTH, both corticosterone and cortisol increased significantly at 4 and 10 days indicating that AC’FH was not selectively acting on only one pathway. However, by 17 days, the levels of corticosterone and cortisol were similar to those of control values thus indicating an apparent failure of the ACTH response.
The predominance of corticosterone and the absence or relatively low levels of cortisol in the plasma and adrenals of adult birds (see review of Kalliecharan and Hall, 1974) resulted in later studies being directed primarily toward determining the presence of corticosterone and its intermediates. oos and deRoos (1964) and demonstrated that both TH and an acid extract of enohypophysis can significantly increase both corticosterone and alosterone ‘on in vitro. Recently, owever, ura et al. (1978) have at the 17-hydroxylase activity that is necessary for cortisol formation in chick embryos (Kahiecharan and Hall, 1977; Idler ek al., 1976) disappears from the adrenal lands of 7- to 14”day-old posthatching hicks. In view of this, porcine ACTH was tested thatching chicks to determine it can influence the biosynthetic s of corticosterone and cortisol in the same manner as reported in the in vitro ryonic chick adrenals (KalHall, 1977) or whether prot can cause cortisol ing steroid as demass et al., 1954). y-old posthatching and onward were injected intramuscularly
perimental
chicks recei
d in the abode gelatin s were wea
minute cortisol ple. Samples
water (98:lQ: 1) mixture
of [3H]coreach were
we
w
249 0016-6480/81/060249-03So1.OO/O Copyright @ 1981 by Academic Press, Inc. AIi rights of mprodwtion in any form resend
250
NOTES
costerone antiserum B21-42 (Endocrine Sciences, Tarzana, Calif.) was used for the determination of corticosterone, whereas a radioimmunoassay kit for cortisol was obtained from New England Nuclear (Boston, Mass.). The concentration of corticosteroids in the plasma of 4-, lo-, and 17-day-old chicks is presented in Table 1, and the difference in weight between Days 3 and 4, 3 and 10, and 3 and 17 is shown in Table 2. The results from Table 1 indicate that the corticosterone level in the plasma of control chicks is low (5 &ml) at 4 days posthatching, yet significantly less than the levels at 10 (8.0 &ml) and 17 (8.1 &ml) days posthatching (PC 0.001). These values correlate well with the increasing activity of the enzyme, 21-hydroxylase that is necessary for corticosterone formation, in 3-, 7-, and 14day-old chick adrenals (Nakamura et d., 1978). The decreasing pattern and total disappearance of cortisol from the plasma of control chicks -(see Table 1) agree with the adrenal gland data from posthatching chicks (Nakamura et al., 1978). The fact that corticosterone and cortisol increased significantly in the plasma of 4and IO-day-old chicks following ACTH treatment (see Table 1) indicates that ACTH is not selectively acting on only one corticosteroid pathway (Bush, 1953; Kalliecharan and Hall, 1977). However, the apparent failure of prolonged ACTH treatment to significantly alter corticosterone and cortisol levels in the plasma of 17-dayold chicks may be due to an inactivation of the ACTH molecule by antibodies formed as a result of the daily injections of ACTH over a prolonged period. The low levels of corticosterone found may reflect basal levels from the adrenal glands in the absence of ACTH stimulation (Woods et al., 1971) and this may account for the significant decrease in body weight (see Table 2). This decrease in body weight and the inability of corticosterone to respond to prolonged ACTH has also been demonstrated by Kass et al., (1954). These authors have
NOTES
BODY
WEIGHT
INCREASES
251
TABLE 2 3 AND 4, 3 AND 10, AND 3 ACTH-TREATED POSTHATCHING CHICKS BETWEEN
DAYS
AND
17 IN CONTROL .\XD
Body weight increase Age of chicks (davs)
Control
4 10 17
5.4 t 0.7 (6)” 45 f 1.1 (6) 130.9 +- 3.1 (5)
3.8 + 0.6 (6) 38.2 k 2.2 (6) 101 i- 5.9 (5)
Note. Values are means + SEM. u Number of animals shown in parentheses. * Not significant at the 5% level. V Statistical comparison of weight differences between control and ACTH-treated
shown that the corticosteroid pathways of rabbits can be altered to secrete primarily cortisol instead of corticosterone following prolonged ACTH treatment. In addition, they reported that the higher cortisol level compensated for the loss in plasma corticosterone. Since, cortisol is present in minute quantities in the plasma of control rabbits (Rass eb al., 1954), the total absence of this steroid from the plasma of 17-day-old contro1 chicks indicates that a genetic block on the enzymes of the C-17 hydroxylating pathway is effective somewhere between and 17. It appears that once this genetic mechanism is activated, possibly would have nb effect on this pathway as shown in Table 1. If there is an alteration of corticosteroid production, as indicated by ass et al. (1954), then, some other steroid may have to fulfil this role, possibly aldosterone which increases in irds following ACTH stimulation (deRoos, 969). CKNQWLEDGMENT Supported by National Sciences and Engineering Research Council of Canada grant (A0460) to R. Kalliecharan.
NS” NS P < O.QI’
animais.
deRoos, R., and deRoos, C. C. (1964). Effects of mammalian corticotropin and chicken adenohypophyseal extracts on steroidogenesis by chicken adrenal tissue in vitro. Gen. Camp. Endocrinol. 4, 602, 607. Idler, D. R., Walsh. J. M.. Kalliecharan, R., and Hall, B. K. (1976). Identification of authentic cortisol in plasma of the embryonic chick. GePz. Comg. Ep?docrinol.
30, 539-540.
Kalliecharan, R., and Hail, B. K. (1974). A developmental study of the levels of progesterone, COB’ticosterone, cortisol, and cortisone circulating in plasma of chick embryos. Gen. Camp. Endocrinol.
24, 364-372.
Kalliecharan, R., and Hall, B. K. (1977). The in vitro biosynthesis of steroids from pregnenolone and cholesterol and the effects of bovine ACTH on corticoid production by adrenal glands of em”bryonic chicks. Gen. Camp. Ertdocritioi. 33, 147-159. Kass, E. H., Hechter, O., Mac&i, 1. A., and Mou, T. W. (1954). Changes in the pattern of secretion of corticosteroids in rabbits after nrolonsed treatment with ACTH. Proc. Sot. Exp. Bioi. k&d. 85, 583-587. Nakamura, T., Tanabe. Y., and Hirano, H. (1978). Evidence of the in vitro formation of cortisol by the adrenai gland of embryonic and young chickens (Gallas domesricus). Gen. Camp. Endocvinot 3.5, 302-308. Woods, J. E., Devries, V. W., and Thommes, R. C. (1971). Ontogenesis of the ~ituita~-adrenal axis in chick embryos. Gcn. Comp. Endocrine!. 17, 407-415.
EFERENCES Bush, I. E. (1953), Species differences in adrenocortiretion. J. Endocrinol. 9, 95- 101. cal deRoos, . (1969). Effects of mammalian corticotropin and progesterone on corticoid production by chicken adrenal tissues in vifro. Gen. Camp. Endocrinol.
13, 455-459.
Department St. Francis Antigonish, Accepted
ofBio!ogy Xavier University B2G ICO, Nova Scotia October 24. 1980