Normal chick embryos swallow and utilize the albumen in the egg between the 11th and 15th day of incubation (Witschi, 1949). They use approximately two-thirds of the yolk by the end of incubation (Romanoff & Romanoff, 1949). Thus the embryo has access to 4 g of protein in the albumen and 3 g of protein in the yolk (Romanoff & Romanoff, 1949). Embryos which have been hypo-physectomized by partial decapitation, which is the only practical way of hypophysectomizing chick embryos (Betz, 1965 a, see also the reviews of Benoit (1962) and of Hinni & Watterson (1963), do not develop an upper beak and do not swallow much of the albumen since most of it remains in the egg (Witschi, 1959; Betz, 1965b).

The rate of growth of hypophyseoprivic embryos between the 14th and 18th day of incubation is reduced to 53% of the normal rate (Vogel, 1965). At 18-5 (Thommes, 1967) and 20 days of incubation (Fugo, 1940; Case, 1952; Betz, 1967) these embryos are only one-half as heavy as normal embryos. These results contradict those of Wolff & Stoll (1937) and Stoll (1939). Witschi (1959) said, in comparing the body weight of normal and hypophysectomized embryos, ‘… it is evident that obstruction of alimentary intake must dwarf the fetus from the 11th day on. On the other hand, if the albumen is swallowed the chick must gain weight even if the albumen were not assimilated. Therefore, the question of somatotrophic hormone (STH) deficiency remains undecided’. Thommes (1967) reported that pars distalis grafts restored the body water content of hypophysectomized chick embryos to normal by 18·5 days of incubation if the data were expressed as the ratio of grams of water in the body to grams of dry body weight; however, both the wet and dry body weights were subnormal. Betz (1967) reported that most of the characteristic defects of hypophysectomized chick embryos are prevented by pars distalis grafts. Even though the growth of these embryos is improved it is still subnormal. Thus, the limited growth of these embryos may be due either to reduced albumen ingestion or failure of the pars distalis grafts to produce normal amounts of STH and other hormones.

This paper reports the results of experiments which distinguish between the effects of hormones and of albumen ingestion on the growth of chick embryos.

The experimental design is summarized in Table 1. White Leghorn eggs from the University of Illinois poultry farm were used in this study. The embryos in groups one, four and six were treated as described by Betz (1967). The rudiment of the upper beak in group two was resected at stage 12 (Hamburger & Hamilton, 1951) of incubation. The albumen in group three was removed with a pipette through a window in the shell on the fourth day of incubation. The embryos in groups five and seven were hypophysectomized at stage 12 by resecting the head at the posterior end of the mesencephalon. This results in a pituitaryless embryo with a non-patent esophagus. The amount of albumen ingested by groups one, two, four and six was estimated by marking the albumen with two drops of sterile carmine suspension (0·1 g/ml of Ringer’s solution) which were added to each egg on the fourth day of incubation through a window in the shell. The eggs were placed in egg cartons and rotated five times. This, it was found, was enough to insure an even distribution of the dye in the albumen.

Table 1

Experimental groups for the effects of graft hormones and albumen on the growth of chick embryos

Experimental groups for the effects of graft hormones and albumen on the growth of chick embryos
Experimental groups for the effects of graft hormones and albumen on the growth of chick embryos

At the end of 20 days of incubation the embryos were killed and the amount of dyed meconium in the ventriculus, the amount of albumen remaining in the egg and the amount of depot fat in the embryos were scored by visual estimation. Then the embryos were fixed according to the method of Betz (1967). The length of the third toe was measured twice to the nearest 0·1 mm according to the criteria of Hamburger & Hamilton (1951). The duplicate measurements were within 1% of each other. The fixed bodies of the normal and hypophysectomized embryos (in which parts of the head are missing) were made comparable by severing the necks between the fifth and sixth cervical vertebrae. Then the headless bodies were weighed. The amount of albumen ingested by the different groups was variable. Therefore the gut was removed and the bodies were weighed in order to see if the gut contents significantly affected the differences between the groups. The headless, gutless bodies were dried to a constant weight at 60°C. All weights were determined twice to the nearest 0·1 g. The duplicate weights did not vary more than 1%. The mean values of the third toe lengths and the body weights of the groups were calculated and in order to compare the groups the mean values of the experimental groups were expressed as a percentage of the normal mean value.

The albumenless embryos in group three were Hyline strain 934F. This group was compared to normal Hyline embryos. Therefore, the mean body weight of group three expressed as a percentage of the normal value is comparable to those of the other groups. Only fresh and dry body weights were determined.

The embryos in group three were small but otherwise normal in their development. These embryos even retracted their yolk sacs and reduced their vitelline and chorioallantoic circulations as did the embryos in groups one, two, four and five. This was not accomplished by the embryos in groups six and seven. None of the embryos were achondroplastic, nor were there any apparent differences in the relative amounts of depot fat or any dimorphic somatic differences. The data concerning the amount of albumen ingested, the length of the third toe and the body weights are summarized in Table 2.

Table 2

The mean length (mm) of the 3rd toe, weight (g) of the modified bodies and amount of albumen ingested by the embryos in the normal and experimental groups

The mean length (mm) of the 3rd toe, weight (g) of the modified bodies and amount of albumen ingested by the embryos in the normal and experimental groups
The mean length (mm) of the 3rd toe, weight (g) of the modified bodies and amount of albumen ingested by the embryos in the normal and experimental groups

The amount of albumen swallowed by groups two and four was the same, two-fifths of the normal amount, but it is twice the amount swallowed by group six. The lengths of the third toe of the embryos in groups two, four and five were reduced to a similar extent, but were still longer than those of groups six and seven. The differences in the length of the third toe indicate that the skeletal growth of groups two, four and five was greater than that in groups six and seven. The body weights of groups two and four were not different from each other although they were abnormal. Group five was actually lighter than two and four but they were all significantly heavier than groups six and seven. On the basis of the percentage of normal value group three was slightly heavier than groups two and four.

For all the groups, the contents of the gut and the water content of the body did not appear to make an appreciable contribution to the differences among them.

Apparently the growth of the embryos in groups two, three, four and five was retarded because of dietary restriction. For some unknown reason these embryos were unable to use the protein in the yolk to attain normal growth. The additional dwarfing of group five, even though the amount of skeletal growth was the same as groups two and four, was apparently due to the absence of albumen. However, the embryos of group three which also did not have any albumen were slightly heavier than those of groups two and four. Perhaps the Hyline embryos normally use more of the protein in the yolk for growth than do the White Leghorn embryos.

The pars distalis grafts in group four apparently produced normal amounts of hormones because these embryos grew the same amount as the embryos in group two which had intact partes distales.

The embryos in groups six and seven were significantly smaller than those in the other groups. The small amount of albumen swallowed by the hypophysectomized embryos in group six had no effect on their growth.

Simpson, Asling & Evans (1950) and Nalbandov (1963) argue that the growth of vertebrates depends upon growth hormone and that other hormones and nutrition are only permissive in the process. These data do not show the extent to which the thyroid and adrenocortical hormones may be involved in the growth of chick embryos.

The data confirm and extend the work of Betz (1967) and they support the conclusion of Hadorn (1961), Herrmann, Clark & Landauer (1963) and Allenspach (1966) that the lethality of the syndrome of the crooked neck dwarf mutant in chick embryos is not due to chronic starvation caused by esophageal atresia as proposed by Pun (1954). However, these results would explain the reduced body size which is a characteristic of these mutant embryos.

The growth which occurs in hypophysectomized chick embryos is independent of pars distales hormones and it is not affected by albumen ingestion. However, in embryos with intact partes distales and in hypophysectomized embryos with pars distalis grafts the additional growth which occurs depends on hormones. If the dietary intake of these embryos is variously restricted, the amount of growth is proportionately limited.

Les effets de la greffe de pars distalis embryonnaire et de I’albumen, sur la croissance d’embryons de poulet

La croissance d’embryons de poulet hypophysectomisés est indépendante des hormones de la adénohypophyse et n’est pas affectée par l’ingestion d’albumen. Néammoins, chez les embryons à adénohypophyse intacte et chez es embryons hypophysectomisés porteurs de adénohypophyse greffée, la croissance additionnelle observée est sous la dépendance d’hormones. Si l’absorption alimentaire de ces embryons est réduite de manière variable, le degré de croissance est proportion nellement limité.

I wish to acknowledge the expert technical assistance of Mr James Bell and particularly of Miss Blanche Vachon. I am especially indebted to Professor A. E. Wilhelmi for a critical review of the manuscript.

This research was supported in part by a grant to Dr Ray L. Watterson from the Graduate School of the University of Illinois and by a grant from the National Research Council of Canada and the Ontario Department of University Affairs.

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