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Notes: Summary Chick embryos were treated with the cytostatic substance triethylenemelamine (TEM) on the 3 1/2th, 4th or 4 1/2th day of incubation (one injection per egg). One group of embryos was fixed in Bouin's solution on the 7th or between the 9th and 11th day of incubation for the comparative study of alterations of the external morphology. The majority of embryos was investigated for alterations in the cartilaginous skeleton after staining with methylene blue and subsequent clearing. This paper deals with the experimental technique, a general survey of the experimental material and the results obtained from the Bouin-fixed embryos. The analysis of the cartilaginous skeleton will be subject of later publications (parts II and III). The teratogenic dose of 7 μg TEM (administration into the air chamber, = „standard treatment“, fixation between 9th and 11th day) was-under the given experimental conditions — equivalent to the embryonal LD 50 when the treatment occurred on the 4th day. For the various stages of treatment (standard treatment at the 3 1/2th, 4th or 4 1/2th day, then fixation between 9th and 11th day of incubation) lethality decreases with increasing age of treatment. Injection of TEM into the yolk sac results in more marked malformations than does injection of the same dose into the air chamber. Treatment with 6–10 μg TEM (injection into the air chamber) or with 5–7 μg TEM (injection into the yolk sac) led to reduction of all dimensions of the body in all experimental embryos (including those with normal outer appearance). In general, the body dimensions were more markedly reduced, the more malformed an embryo was. After standard treatment of the three experimental stages and fixation on the 7th day of incubation the lengths of the head, of the trunk and of the hind legs respectively were more reduced, the earlier the treatment occurred. The outer malformations characteristic for the present experimental material are the following: oedema in various forms, enlarged and anteriorly transposed umbilicus, umbilical hernia, bending, torsion and reduction of extremities (in most cases combined with oligodactylism), reduction and bending downwards of the upper beak, clefts between nasal opening and border of the beak, cleft palate of various degrees, strong reduction of the lower beak, reduction of size of the eyes, suppression of scleral papillae, reduction of eyelids, retardation in the development of embryonal pterylosis, suppression of feather anlagen. Apart from metric criteria, the development of nasal clefts is treatment-phase dependent. Such a phase specifity is recognizable in the outer appearance of strongly malformed hind legs too. The alterations due to TEM treatment between the 3 1/2the and 4 1/2th day of incubation on both sides of the sagittal plane are mirrorimages of a high degree.

Notes: Summary The effects of TEM on the formation of the cartilaginous skeleton of limbs and trunk were investigated in cleared methylene blue preparations of 9–11 day chick embryos. Embryos were treated at 3 1/2, 4 or 4 1/2 days incubation (treatment groups I, II and III respectively). Details of the techniques, dosage effects, lethality and changes in external features have been given in a previous paper (Part I). Effects on the cartilaginous skeleton of the head will be considered in Part III. The total pattern of skeletal alterations shows a marked correlation with the age at which the embryos were treated (treatment-phase specificity). The most distinct range of effects is seen in the foot skeleton. Within each treatment group, the cases can be arranged in a series according to their degree of malformation. The degree of malformation in other parts of the skeleton usually shows some correspondence whit that in the foot. Skeletal changes within each of the three treatment groups may be characterized as follows: in thehind legs, the marginal rays of the autopodium are most sensitive to TEM treatment. In treatment group I, mainly the proximal part of the metatarsus and the marginal rays of the acropodium are reduced. The most severe case shows distal remainders of metatarsals II and III, with reduced numbers of phalanges, and a malformed remainder of the tarsus. In treatment group II, there is more reduction of the acropodium with heavier loss of the marginal rays; in severely affected cases, the reduction covers the whole metatarsus as well. In the most extreme case, there is only a remainder of the acropodial part of the Illrd ray immediately following the tarsus. In treatment group III, the zone of damage is further distal: with increasing degree of malformation phalanges together with the distal metatarsus tend to disappear, while the tarsus is less affected. Within all three treatment groups, characteristic bendings of the metatarsus occur. Reduction of the digits occurs by proximo-distal fusion of adjacent phalanges, in a certain sequence. In the 1st ray, the basal phalanx fuses with the metatarsal. The IIIrd ray most rarely shows any reduction. The long skeletal elements of the limbs also show alterations which are typical for each treatment group. In the fibula not only reductions occur following TEM-treatment, but also relative increases of its length (most frequently in treatment group III). In thewing skeleton, which is less sensitive than that of the leg to TEM, the zones of damage again shift from proximal to distal with increasing age of treatment. Severely malformed embryos of group I show a bending forward and fusion of humerus and ulna, aplasia of the radius, and only slight alterations in the autopodium. In group II, there is more marked damage to the autopodium. In group III, the wing ends with remainders of metacarpals III and IV. In thepelvic girdle the main alterations are: partial or total loss of the os pubis, and reduction of ischium and ilium. In thepectoral girdle, there is reduction or loss of coracoid and scapula. In thethorax, the sternum is reduced (its two anlagen may not having been fused) and there is loss or reduction of the sternal parts of the ribs, terminal reduction of vertebral parts of the ribs, great reduction or loss of the ribs of cervical 15 and thoracic 5, and loss of the uncinate processes. In thevertebral column, some vertebrae of the tail are missing, and in severely affected cases there are fusions of vertebral arches and other anomalies. Apart from a minor abnormality of the foramen arteriae ophthalmicae (see Part III), the os pubis is the most sensitive cartilaginous skeletal element to TEM treatment between the 3rd and 4th days of incubation. At 4 1/2 days, the skeletons of the foot and wing are the most sensitive parts.