- Email: [email protected]
Steroidal stimulation of mammary glands in prenatally feminized male rats
EUROPEAN JOURNAL OF PHARMACOLOGY 1 (1967) 120-123.
NORTH-HOLLAND PUBL. COMP., AMSTERDAM
STEROIDAL STIMULATION OF MAMMARY GLANDS IN PRENATALLY FEMINIZED MALE RATS F . N E U M A N N a n d W. E L G E R
Schering AG, Abteilung flit Endokrinologie, Berlin, Germany Accepted 12 D e c e m b e r 1966
F. NEUMANN and W. ELGER, Steroidal stimulation of mammary glands in prenatally feminized male rats, European J. P h a r m a c o l . 1 (1967) 120-123. When a d m i n i s t e r e d to gravid r a t s during pregnancy an anti-androgenic steroid (1, 2 ~ - m e t h y l e n e - 6 chloro-A4, 6 - p r e g n a d i e n e - 1 7 ~ - o l - 3 , 20-dione-17C~-acetate) induced development of nipples in male fetuses. These nipples and associated glandular t i s s u e s develop after b i r t h as in n o r m a l female animals. P r o g e s t i n - e s t r o g e n t r e a t m e n t of adult, c a s t r a t e d feminized m a l e s produced stimulation of the glandular tissue s i m i l a r to that seen after t r e a t m e n t of c a s t r a t e d female animals. In c a s t r a t e d male r a t s this t r e a t m e n t p r o d u c e s little glandular proliferation. It is concluded that androgens normally prevent the development of nipples and extensive formation of m a m m a r y tissue in male fetuses. Female sex h o r mones s e e m to be without significance in differentiation of the m a m m a r y glands and t h e i r p r e - p u b e r t a l development. Differentiation of m a m m a r y gland Antiandrogen
Feminized r a t s Weight of m a m m a r y glands
1. I N T R O D U C T I O N T r e a t m e n t of p r e g n a n t r a t s w i t h a n a n t i a n d r o g e n i c s t e r o i d (1, 2 a - m e t h y l e n e - 6 - c h l o r o - A 4, 6 pregnadiene-17a-ol-3, 20-dione-17a-acetate, cyproterone acetate) * produces a feminine organog e n e s i s of t h e m a m m a r y g l a n d i n m a l e f e t u s e s ( N e u m a n n a n d E l g e r , 1966a), i n a d d i t i o n to t h e f e m i n i z a t i o n of o t h e r s o m a t i c - a n d C N S - s e x u a l characteristics (Hamada, Neumann and Junkm a n n , 1963; N e u m a n n , E l g e r a n d K r a m e r , 1966; N e u m a n n a n d E l g e r , 1965). T h i s e f f e c t i s r e s p o n s i b l e f o r t h e d e v e l o p m e n t of t h e n i p p l e s , w h i c h n o r m a l l y do not d e v e l o p i n m a l e r a t s . E v e n a f t e r d i s c o n t i n u a t i o n of t r e a t m e n t n i p p l e s c o n t i n u e to d e v e l o p t h r o u g h o u t p o s t - n a t a l s t a g e s . However, in the feminized male rats, pron o u n c e d d e v e l o p m e n t of g l a n d u l a r t i s s u e c o u l d b e recognized with certainty in the proximal sect i o n s of t h e g l a n d u l a r b u d o n l y f r o m day 19 to 21 of e m b r y o n i c d e v e l o p m e n t . A t t h e t i m e of b i r t h this observation was made difficult by the ext e n s i v e a r e a of g l a n d u l a r t i s s u e ( N e u m a n n a n d E l g e r , 1966a). T h e p r e s e n t p a p e r w i l l c o n s i d e r how t h e d e * This compound was synthesized by Dr. R. Wieehert, Schering AG, Berlin.
Pseudogravidity
v e l o p m e n t of m a m m a r y t i s s u e i n m a l e a n i m a l s i s i n f l u e n c e d b y t r e a t m e n t of t h e i r m o t h e r s w i t h a n a n t i a n d r o g e n d u r i n g p r e g n a n c y a n d how t h e i r mammary t i s s u e s r e a c t to e s t r o g e n - p r o g e s t i n treatment when these feminized animals are adult.
2. M A T E R I A L S AND M E T H O D S Adult female Sprague-Dawley rats were placed with males during pro-estrus. The day sperm was found in the vaginal smears was c o n s i d e r e d a s day 1 of p r e g n a n c y o r of e m b r y o n ic d e v e l o p m e n t . T h e p r e g n a n t f e m a l e s r e c e i v e d c y p r o t e r o n e a c e t a t e , 10 m g / d a y s u b c u t a n e o u s l y , f r o m day 13 to day 22 i n 0.2 m l of a b e n z y l b e n z o a t e - c a s t o r o i l m i x t u r e ( 1 : 5 ) . On day 23 of pregnancy the fetuses were removed and transf e r r e d to f o s t e r m o t h e r s . T h e a d u l t r a t s , c a s t r a t e d on d a y 150 of l i f e , w e r e t r e a t e d f o r a 2 2 - d a y p e r i o d w i t h 10 ~ g e s t r a d i o l b e n z o a t e a n d 30 m g p r o g e s t e r o n e / d a y by intramuscular administration. Both drugs w e r e g i v e n t o g e t h e r i n a s i n g l e i n j e c t i o n of 0.2 ml benzyl benzoate - castor oil mixture (1:5). Castrated female and male animals, subjected to t h e s a m e t r e a t m e n t , w e r e u s e d a s c o n t r o l s .
MAMMARY GLANDS IN FEMINIZED RATS
121
4. DISCUSSION
Fig. 1. Nipple of an adult feminized male rat. The m o t h e r s were t r e a t e d with 10 mg cyproterone a c e t a t e / day from the 13th to the 22nd day of pregnancy. The animals were killed on the 23rd day and the mammary glands were excised and weighed. A f t e r f i x a t i o n i n 10% f o r m a l i n , t h e m a t e r i a l w a s e m b e d d e d i n p a r a f f i n , cut, a n d s t a i n e d w i t h hematoxylin-eosine.
3. R E S U L T S T h e e x i s t i n g n i p p l e s of f e m i n i z e d m a l e a n i mals and female controls developed under the i n f l u e n c e of e s t r o g e n - p r o g e s t i n t r e a t m e n t ( m a l e c o n t r o l s h a v e no n i p p l e s ) . On n e c r o p s y , a l l a n i mals exhibited a more or less developed mamm a r y g l a n d . Fig. 2 s h o w s t h e m a m m a r y g l a n d w e i g h t of f e m i n i z e d m a l e a n i m a l s ( g r o u p II), f e m a l e c o n t r o l s ( g r o u p I) a n d m a l e c o n t r o l s ( g r o u p III), following estrogen-progestin treatment. While in the male controls only a small amount of m a m m a r y t i s s u e w a s found, t h e r e w a s a p r o nounced development in feminized male animals and in female animals. Pectoral and inguinal glandular tissues showed equivalent development. T h e m a m m a r y g l a n d s of t h e f e m i n i z e d m a l e a n i m a l s r e a c t e d to e s t r o g e n - p r o g e s t i n t r e a t m e n t i n a m a n n e r s i m i l a r to t h o s e of f e m a l e a n i m a l s , while the male controls showed less reaction. H i s t o l o g i c a l e x a m i n a t i o n r e v e a l e d no e v i d e n t qualitative differences in the glandular struct u r e s of t h e v a r i o u s g r o u p s ; i n a l l c a s e s c o n s i d erable glandular proliferation was noticed. Q u a l i t a t i v e l y t h e s e g l a n d s c o r r e s p o n d to t h o s e of p r e g n a n t f e m a l e r a t s s h o r t l y b e f o r e t h e o n s e t of lactation. Some alveoli contained glandular sec r e t i o n ( s e e fig. 3).
The findings indicate that under the influence of a n t i a n d r o g e n s , n o t o n l y c a n t h e d e v e l o p m e n t of n i p p l e s b e i n d u c e d i n m a l e r a t f e t u s e s ( t h e y are nonexistent in normal male rats), but the g l a n d i t s e l f i s s u b j e c t e d to t h e s a m e d e v e l o p m e n t as is seen in female fetuses. The fact that estrog e n - p r o g e s t i n t r e a t m e n t of a d u l t f e m i n i z e d r a t s a n d f e m a l e a n i m a l s c a u s e s t h e s a m e d e g r e e of p r o l i f e r a t i o n of g l a n d u l a r t i s s u e of t h e m a m m a r y g l a n d , w h i l e t h e d e v e l o p m e n t of g l a n d u l a r t i s s u e i s q u i t e l i m i t e d i n t h e m a l e c o n t r o l s , s e r v e s to p r o v e t h a t in f e m i n i z e d m a l e r a t s m o r e g l a n d u l a r t i s s u e e x i s t e d a t t h e b e g i n n i n g of t h e t r e a t m e n t t h a n in n o r m a l m a l e r a t s . T h e d i f f e r e n c e s in t h e d e v e l o p m e n t of g l a n d u lar tissue in female controls and in feminized m a l e a n i m a l s a n d in m a l e c o n t r o l s a r e m e r e l y q u a n t i t a t i v e . T h e h i s t o l o g i c a l p i c t u r e s s h o w no differences. C y p r o t e r o n e a c e t a t e i n h i b i t s t h e e f f e c t of a n drogens in a competitive manner at the recept o r s of t h e t a r g e t o r g a n s . T h i s w a s c o n f i r m e d in earlier studies (Neumann, Richter and Giinzel, 1965; N e u m a n n a n d V o n B e r s w o r d t - W a l l r a b e , 1966; N e u m a n n a n d E l g e r , 1966b). S i n c e t h i s compound does not possess estrogenic proper( 10- g
) = Number of animals
I =S£ ,
t-Test p< 0,001 : I v.TIT ]I v.11T p<0,01 : I v . 1T ,4.
5
11( 11T
]I
I
Fig. 2. Weight of m a m m a r y glands of c a s t r a t e d female controls (group I), c a s t r a t e d feminized male r a t s (group II) and c a s t r a t e d male controls (group I I I ) f o l lowing 22-day t r e a t m e n t with 10 ~lg e s t r a d i o l - b e n z o a t e and 30 mg p r o g e s t e r o n e / d a y i.m. The m o t h e r s were t r e a t e d with 10 mg cyproterone a c e t a t e / d a y from the 13th to the 22nd day of pregnancy.
122
F. NEUMANN and W. E L G E R
Fig. 3. M a m m a r y gland of a c a s t r a t e d feminized m a l e r a t following 22-day t r e a t m e n t with 10 //g e s t r a d i o l - b e n z o a t e and 30 m g p r o g e s t e r o n e / d a y i.m. (x 35). The m o t h e r s w e r e t r e a t e d with 10 m g c y p r o t e r o n e a c e t a t e / d a y f r o m the 13th to the 22nd day of preg~aaney.
Fig. 4. M a m m a r y gland of a c a s t r a t e d feminized m a l e r a t following 22-day t r e a t m e n t with 10 //g e s t r a d i o l - b e n z o a t e and 30 m g p r o g e s t e r o n e / d a y i.m. (x 140). The m o t h e r s w e r e t r e a t e d with 10 m g c y p r o t e r o n e a c e t a t e / d a y f r o m the 13th to the 22rid day of p r e g n a n c y .
MAMMARY GLANDS IN FEMINIZED RATS t i e s (which w o u l d l e a d to e n t i r e l y d i f f e r e n t m a m m a r y m a l f o r m a t i o n ( R a y n a u d a n d R a y n a u d , 1956, 1957)), t h e f o l l o w i n g c o n c l u s i o n s m a y b e d r a w n r e g a r d i n g t h e s i g n i f i c a n c e of t h e m a l e a n d f e m a l e s e x u a l h o r m o n e s f o r t h e d i f f e r e n t i a t i o n of t h e m a m m a r y g l a n d : T h e d i s p l a c e m e n t of the f e tal a n d r o g e n s ( t e s t o s t e r o n e ? ) f r o m the r e c e p t o r s of t h e m a m m a r y g l a n d in the m a l e f e t u s e s a t the t i m e of t h e d i f f e r e n t i a t i o n of t h i s o r g a n i s s u f f i c i e n t to i n d u c e t h e d e v e l o p m e n t of b o t h n i p p l e s a n d g l a n d u l a r t i s s u e in t h e s a m e m a n n e r a s w o u l d n o r m a l l y t a k e p l a c e only in f e m a l e a n i m a l s . On t h e b a s i s of o u r s t u d i e s , it a p p e a r s to b e e x t r e m e l y d o u b t f u l t h a t any d e c i d e d s i g n i f i c a n c e c a n b e a s c r i b e d to t h e e s t r o g e n s a s t a k i n g p a r t in any o n e p h a s e of d e v e l o p m e n t of t h e m a m m a r y g l a n d s up to p u b e r t y . It m a y t h u s b e s t a t e d w i t h c e r t a i n t y t h a t only t h e a n d r o g e n s of t h e f e t a l t e s t e s p r e v e n t a m o r e p r o n o u n c e d d e v e l o p m e n t of m a m m a r y t i s s u e in male animals.
A C K N O W LE DGE M E N T We wish to thank Miss Boehnisch, Miss F l e i s c h e r and Miss Blank for t h e i r a s s i s t a n c e in the experiments.
REFERENCES Hamada, H., F. Neumann and K. Junkmann, 1963, Intrauterine antimaskuline Beeinflussung yon Rattenfeten dutch ein stark gestagen w i r k s a m e s Steroid, Acta endocr. (Kbh.) 44, 380.
123
Neumann, F. and W. Elger, 1965, Physiological and psychical intersexuality of male r a t s by early t r e a t m e n t with an anti-androgenic agent (1,2ormethylene- 6-chloro- A 6 - h y d r o x y - p r o g e s t e r o n e - a c e tare), 5th Acta Endocr. Congress, Hamburg, Acta endocr. (Kbh.) Suppl. 100, 174 (Abstract No. 142). Neumann, F. and W. Elger, 1966a, The effect of the antiandrogen 1, 2Ot-methylene-6-chloro-A4, 6 - p r e g nadiene- 17ot-ol-3, 20-dione- 17Ol-acetate (cyprotetone acetate) on the development of the m a m m a r y glands of male fetal r a t s , J. Endocr., 36, 347. Neumann, F. and W. Elger, 1966b, Proof of the activity of androgenic agents on the differentiation of the external genitalia, the m a m m a r y gland and the hypothalamus-pituitary-gland s y s t e m in rats, Proc. 2nd Symp. on Steroid Hormones "Androgens - S e cretion and Metabolism in Normal and Pathological Conditions", Ghent 1965, Excerpta Med. Int. Congress Series No. 101, 168. Neumann, F., W. Elger and M . K r a m e r , 1966, The development of a vagina in male r a t s by inhibiting the androgen r e c e p t o r s with an antiandrogen during the critical phase of organogenesis, Endocrinology 78, 628. Neumann, F. and R. Von Berswordt-Wallrabe, 1966, Effects of the antagonist cyproterone acetate on the testicular structure, s p e r m a t o g e n e s i s and a c c e s sory sexual glands of t e s t o s t e r o n e - t r e a t e d adult hypophysectomized rats, J. Endocr. 35, 363. Neumann, F., K.-D. Richter and P. Gtinzel, 1965, Wirkungen von Antiandrogenen, Zbl. Vet. Med., Reihe A, 12, 171. Raynaud, A. and J. Raynaud, 1956, La production exp~rimentale de malformations m a m m a i r e s chez le foetus de souris, par l'aetion des hormones sexuelles, Ann. Inst. P a s t e u r 90, 187. Raynaud, A. and J. Raynaud, 1957, Sur les modifications pr~coces apport~es par l'hormone oestrog~ne dans la structure des ~bauches m a m m a i r e s des foetus de souris, Bull. Soe. Zool. (France) 82, 204.
NORTH-HOLLAND PUBL. COMP., AMSTERDAM
STEROIDAL STIMULATION OF MAMMARY GLANDS IN PRENATALLY FEMINIZED MALE RATS F . N E U M A N N a n d W. E L G E R
Schering AG, Abteilung flit Endokrinologie, Berlin, Germany Accepted 12 D e c e m b e r 1966
F. NEUMANN and W. ELGER, Steroidal stimulation of mammary glands in prenatally feminized male rats, European J. P h a r m a c o l . 1 (1967) 120-123. When a d m i n i s t e r e d to gravid r a t s during pregnancy an anti-androgenic steroid (1, 2 ~ - m e t h y l e n e - 6 chloro-A4, 6 - p r e g n a d i e n e - 1 7 ~ - o l - 3 , 20-dione-17C~-acetate) induced development of nipples in male fetuses. These nipples and associated glandular t i s s u e s develop after b i r t h as in n o r m a l female animals. P r o g e s t i n - e s t r o g e n t r e a t m e n t of adult, c a s t r a t e d feminized m a l e s produced stimulation of the glandular tissue s i m i l a r to that seen after t r e a t m e n t of c a s t r a t e d female animals. In c a s t r a t e d male r a t s this t r e a t m e n t p r o d u c e s little glandular proliferation. It is concluded that androgens normally prevent the development of nipples and extensive formation of m a m m a r y tissue in male fetuses. Female sex h o r mones s e e m to be without significance in differentiation of the m a m m a r y glands and t h e i r p r e - p u b e r t a l development. Differentiation of m a m m a r y gland Antiandrogen
Feminized r a t s Weight of m a m m a r y glands
1. I N T R O D U C T I O N T r e a t m e n t of p r e g n a n t r a t s w i t h a n a n t i a n d r o g e n i c s t e r o i d (1, 2 a - m e t h y l e n e - 6 - c h l o r o - A 4, 6 pregnadiene-17a-ol-3, 20-dione-17a-acetate, cyproterone acetate) * produces a feminine organog e n e s i s of t h e m a m m a r y g l a n d i n m a l e f e t u s e s ( N e u m a n n a n d E l g e r , 1966a), i n a d d i t i o n to t h e f e m i n i z a t i o n of o t h e r s o m a t i c - a n d C N S - s e x u a l characteristics (Hamada, Neumann and Junkm a n n , 1963; N e u m a n n , E l g e r a n d K r a m e r , 1966; N e u m a n n a n d E l g e r , 1965). T h i s e f f e c t i s r e s p o n s i b l e f o r t h e d e v e l o p m e n t of t h e n i p p l e s , w h i c h n o r m a l l y do not d e v e l o p i n m a l e r a t s . E v e n a f t e r d i s c o n t i n u a t i o n of t r e a t m e n t n i p p l e s c o n t i n u e to d e v e l o p t h r o u g h o u t p o s t - n a t a l s t a g e s . However, in the feminized male rats, pron o u n c e d d e v e l o p m e n t of g l a n d u l a r t i s s u e c o u l d b e recognized with certainty in the proximal sect i o n s of t h e g l a n d u l a r b u d o n l y f r o m day 19 to 21 of e m b r y o n i c d e v e l o p m e n t . A t t h e t i m e of b i r t h this observation was made difficult by the ext e n s i v e a r e a of g l a n d u l a r t i s s u e ( N e u m a n n a n d E l g e r , 1966a). T h e p r e s e n t p a p e r w i l l c o n s i d e r how t h e d e * This compound was synthesized by Dr. R. Wieehert, Schering AG, Berlin.
Pseudogravidity
v e l o p m e n t of m a m m a r y t i s s u e i n m a l e a n i m a l s i s i n f l u e n c e d b y t r e a t m e n t of t h e i r m o t h e r s w i t h a n a n t i a n d r o g e n d u r i n g p r e g n a n c y a n d how t h e i r mammary t i s s u e s r e a c t to e s t r o g e n - p r o g e s t i n treatment when these feminized animals are adult.
2. M A T E R I A L S AND M E T H O D S Adult female Sprague-Dawley rats were placed with males during pro-estrus. The day sperm was found in the vaginal smears was c o n s i d e r e d a s day 1 of p r e g n a n c y o r of e m b r y o n ic d e v e l o p m e n t . T h e p r e g n a n t f e m a l e s r e c e i v e d c y p r o t e r o n e a c e t a t e , 10 m g / d a y s u b c u t a n e o u s l y , f r o m day 13 to day 22 i n 0.2 m l of a b e n z y l b e n z o a t e - c a s t o r o i l m i x t u r e ( 1 : 5 ) . On day 23 of pregnancy the fetuses were removed and transf e r r e d to f o s t e r m o t h e r s . T h e a d u l t r a t s , c a s t r a t e d on d a y 150 of l i f e , w e r e t r e a t e d f o r a 2 2 - d a y p e r i o d w i t h 10 ~ g e s t r a d i o l b e n z o a t e a n d 30 m g p r o g e s t e r o n e / d a y by intramuscular administration. Both drugs w e r e g i v e n t o g e t h e r i n a s i n g l e i n j e c t i o n of 0.2 ml benzyl benzoate - castor oil mixture (1:5). Castrated female and male animals, subjected to t h e s a m e t r e a t m e n t , w e r e u s e d a s c o n t r o l s .
MAMMARY GLANDS IN FEMINIZED RATS
121
4. DISCUSSION
Fig. 1. Nipple of an adult feminized male rat. The m o t h e r s were t r e a t e d with 10 mg cyproterone a c e t a t e / day from the 13th to the 22nd day of pregnancy. The animals were killed on the 23rd day and the mammary glands were excised and weighed. A f t e r f i x a t i o n i n 10% f o r m a l i n , t h e m a t e r i a l w a s e m b e d d e d i n p a r a f f i n , cut, a n d s t a i n e d w i t h hematoxylin-eosine.
3. R E S U L T S T h e e x i s t i n g n i p p l e s of f e m i n i z e d m a l e a n i mals and female controls developed under the i n f l u e n c e of e s t r o g e n - p r o g e s t i n t r e a t m e n t ( m a l e c o n t r o l s h a v e no n i p p l e s ) . On n e c r o p s y , a l l a n i mals exhibited a more or less developed mamm a r y g l a n d . Fig. 2 s h o w s t h e m a m m a r y g l a n d w e i g h t of f e m i n i z e d m a l e a n i m a l s ( g r o u p II), f e m a l e c o n t r o l s ( g r o u p I) a n d m a l e c o n t r o l s ( g r o u p III), following estrogen-progestin treatment. While in the male controls only a small amount of m a m m a r y t i s s u e w a s found, t h e r e w a s a p r o nounced development in feminized male animals and in female animals. Pectoral and inguinal glandular tissues showed equivalent development. T h e m a m m a r y g l a n d s of t h e f e m i n i z e d m a l e a n i m a l s r e a c t e d to e s t r o g e n - p r o g e s t i n t r e a t m e n t i n a m a n n e r s i m i l a r to t h o s e of f e m a l e a n i m a l s , while the male controls showed less reaction. H i s t o l o g i c a l e x a m i n a t i o n r e v e a l e d no e v i d e n t qualitative differences in the glandular struct u r e s of t h e v a r i o u s g r o u p s ; i n a l l c a s e s c o n s i d erable glandular proliferation was noticed. Q u a l i t a t i v e l y t h e s e g l a n d s c o r r e s p o n d to t h o s e of p r e g n a n t f e m a l e r a t s s h o r t l y b e f o r e t h e o n s e t of lactation. Some alveoli contained glandular sec r e t i o n ( s e e fig. 3).
The findings indicate that under the influence of a n t i a n d r o g e n s , n o t o n l y c a n t h e d e v e l o p m e n t of n i p p l e s b e i n d u c e d i n m a l e r a t f e t u s e s ( t h e y are nonexistent in normal male rats), but the g l a n d i t s e l f i s s u b j e c t e d to t h e s a m e d e v e l o p m e n t as is seen in female fetuses. The fact that estrog e n - p r o g e s t i n t r e a t m e n t of a d u l t f e m i n i z e d r a t s a n d f e m a l e a n i m a l s c a u s e s t h e s a m e d e g r e e of p r o l i f e r a t i o n of g l a n d u l a r t i s s u e of t h e m a m m a r y g l a n d , w h i l e t h e d e v e l o p m e n t of g l a n d u l a r t i s s u e i s q u i t e l i m i t e d i n t h e m a l e c o n t r o l s , s e r v e s to p r o v e t h a t in f e m i n i z e d m a l e r a t s m o r e g l a n d u l a r t i s s u e e x i s t e d a t t h e b e g i n n i n g of t h e t r e a t m e n t t h a n in n o r m a l m a l e r a t s . T h e d i f f e r e n c e s in t h e d e v e l o p m e n t of g l a n d u lar tissue in female controls and in feminized m a l e a n i m a l s a n d in m a l e c o n t r o l s a r e m e r e l y q u a n t i t a t i v e . T h e h i s t o l o g i c a l p i c t u r e s s h o w no differences. C y p r o t e r o n e a c e t a t e i n h i b i t s t h e e f f e c t of a n drogens in a competitive manner at the recept o r s of t h e t a r g e t o r g a n s . T h i s w a s c o n f i r m e d in earlier studies (Neumann, Richter and Giinzel, 1965; N e u m a n n a n d V o n B e r s w o r d t - W a l l r a b e , 1966; N e u m a n n a n d E l g e r , 1966b). S i n c e t h i s compound does not possess estrogenic proper( 10- g
) = Number of animals
I =S£ ,
t-Test p< 0,001 : I v.TIT ]I v.11T p<0,01 : I v . 1T ,4.
5
11( 11T
]I
I
Fig. 2. Weight of m a m m a r y glands of c a s t r a t e d female controls (group I), c a s t r a t e d feminized male r a t s (group II) and c a s t r a t e d male controls (group I I I ) f o l lowing 22-day t r e a t m e n t with 10 ~lg e s t r a d i o l - b e n z o a t e and 30 mg p r o g e s t e r o n e / d a y i.m. The m o t h e r s were t r e a t e d with 10 mg cyproterone a c e t a t e / d a y from the 13th to the 22nd day of pregnancy.
122
F. NEUMANN and W. E L G E R
Fig. 3. M a m m a r y gland of a c a s t r a t e d feminized m a l e r a t following 22-day t r e a t m e n t with 10 //g e s t r a d i o l - b e n z o a t e and 30 m g p r o g e s t e r o n e / d a y i.m. (x 35). The m o t h e r s w e r e t r e a t e d with 10 m g c y p r o t e r o n e a c e t a t e / d a y f r o m the 13th to the 22nd day of preg~aaney.
Fig. 4. M a m m a r y gland of a c a s t r a t e d feminized m a l e r a t following 22-day t r e a t m e n t with 10 //g e s t r a d i o l - b e n z o a t e and 30 m g p r o g e s t e r o n e / d a y i.m. (x 140). The m o t h e r s w e r e t r e a t e d with 10 m g c y p r o t e r o n e a c e t a t e / d a y f r o m the 13th to the 22rid day of p r e g n a n c y .
MAMMARY GLANDS IN FEMINIZED RATS t i e s (which w o u l d l e a d to e n t i r e l y d i f f e r e n t m a m m a r y m a l f o r m a t i o n ( R a y n a u d a n d R a y n a u d , 1956, 1957)), t h e f o l l o w i n g c o n c l u s i o n s m a y b e d r a w n r e g a r d i n g t h e s i g n i f i c a n c e of t h e m a l e a n d f e m a l e s e x u a l h o r m o n e s f o r t h e d i f f e r e n t i a t i o n of t h e m a m m a r y g l a n d : T h e d i s p l a c e m e n t of the f e tal a n d r o g e n s ( t e s t o s t e r o n e ? ) f r o m the r e c e p t o r s of t h e m a m m a r y g l a n d in the m a l e f e t u s e s a t the t i m e of t h e d i f f e r e n t i a t i o n of t h i s o r g a n i s s u f f i c i e n t to i n d u c e t h e d e v e l o p m e n t of b o t h n i p p l e s a n d g l a n d u l a r t i s s u e in t h e s a m e m a n n e r a s w o u l d n o r m a l l y t a k e p l a c e only in f e m a l e a n i m a l s . On t h e b a s i s of o u r s t u d i e s , it a p p e a r s to b e e x t r e m e l y d o u b t f u l t h a t any d e c i d e d s i g n i f i c a n c e c a n b e a s c r i b e d to t h e e s t r o g e n s a s t a k i n g p a r t in any o n e p h a s e of d e v e l o p m e n t of t h e m a m m a r y g l a n d s up to p u b e r t y . It m a y t h u s b e s t a t e d w i t h c e r t a i n t y t h a t only t h e a n d r o g e n s of t h e f e t a l t e s t e s p r e v e n t a m o r e p r o n o u n c e d d e v e l o p m e n t of m a m m a r y t i s s u e in male animals.
A C K N O W LE DGE M E N T We wish to thank Miss Boehnisch, Miss F l e i s c h e r and Miss Blank for t h e i r a s s i s t a n c e in the experiments.
REFERENCES Hamada, H., F. Neumann and K. Junkmann, 1963, Intrauterine antimaskuline Beeinflussung yon Rattenfeten dutch ein stark gestagen w i r k s a m e s Steroid, Acta endocr. (Kbh.) 44, 380.
123
Neumann, F. and W. Elger, 1965, Physiological and psychical intersexuality of male r a t s by early t r e a t m e n t with an anti-androgenic agent (1,2ormethylene- 6-chloro- A 6 - h y d r o x y - p r o g e s t e r o n e - a c e tare), 5th Acta Endocr. Congress, Hamburg, Acta endocr. (Kbh.) Suppl. 100, 174 (Abstract No. 142). Neumann, F. and W. Elger, 1966a, The effect of the antiandrogen 1, 2Ot-methylene-6-chloro-A4, 6 - p r e g nadiene- 17ot-ol-3, 20-dione- 17Ol-acetate (cyprotetone acetate) on the development of the m a m m a r y glands of male fetal r a t s , J. Endocr., 36, 347. Neumann, F. and W. Elger, 1966b, Proof of the activity of androgenic agents on the differentiation of the external genitalia, the m a m m a r y gland and the hypothalamus-pituitary-gland s y s t e m in rats, Proc. 2nd Symp. on Steroid Hormones "Androgens - S e cretion and Metabolism in Normal and Pathological Conditions", Ghent 1965, Excerpta Med. Int. Congress Series No. 101, 168. Neumann, F., W. Elger and M . K r a m e r , 1966, The development of a vagina in male r a t s by inhibiting the androgen r e c e p t o r s with an antiandrogen during the critical phase of organogenesis, Endocrinology 78, 628. Neumann, F. and R. Von Berswordt-Wallrabe, 1966, Effects of the antagonist cyproterone acetate on the testicular structure, s p e r m a t o g e n e s i s and a c c e s sory sexual glands of t e s t o s t e r o n e - t r e a t e d adult hypophysectomized rats, J. Endocr. 35, 363. Neumann, F., K.-D. Richter and P. Gtinzel, 1965, Wirkungen von Antiandrogenen, Zbl. Vet. Med., Reihe A, 12, 171. Raynaud, A. and J. Raynaud, 1956, La production exp~rimentale de malformations m a m m a i r e s chez le foetus de souris, par l'aetion des hormones sexuelles, Ann. Inst. P a s t e u r 90, 187. Raynaud, A. and J. Raynaud, 1957, Sur les modifications pr~coces apport~es par l'hormone oestrog~ne dans la structure des ~bauches m a m m a i r e s des foetus de souris, Bull. Soe. Zool. (France) 82, 204.