or 1000 g zearalenone. (Zl000) on Days. 1-10 of life and were castrated on Day. 21. On Day ... in SDN size among the male groups. These data show that exposure ... be detected in the urine of all herbivores, including hu- mans. [9]. In addition to leafy ... [12]. Phytoestrogens have been implicated as the cause of subfertility.
BIOLOGY
OF REPRODUCTION
45, 649-653
(1991)
The Effect of Neonatal Exposure to Diethylstilbestrol, Pituitary Responsiveness and Sexually Dimorphic Adult Rat1 KENNETh
Divisions
of Reproductive
Biology
Gynecology,
and
A. FABER2
and
Duke
and
ClAUDE
Reproductive University
Genistein, and Zearalenone on Nucleus Volume in the Castrated
L. HUGHES,
Endocrinology
Medical
JR.
Infertility,
and
Durham,
Center,
North
Department Carolina
of Obstetrics
27710
ABSTRACT The
neonatal
intracerebral
may
environment
hormone
also
encounter
estrogens
was to assess the effects natal
rats
received
(Gl000),
Day
42,
15,
and
30
siveness
right min.
of either
zearalenone
(Zl0O),
catheters
SDN
of the volume;
exposure
corn or
the
area
and
had
to environmental
no
zearalenone (50 G1000,
of the
administered,
Z100,
was
and
by
groups
were
alters
Sex
and
Zl000
size
Day
1000
were
for
decreased
showed
decreased
Volumes DES,
G1000,
and
male
groups.
These
to GnRH
response
21.
or attendimorphic
Zl000
increased
data
and
respon-
UI
sexually
among pituitary
Day
increased
tonic
of the
In females, the
on
LH at 0, 5, 10,
pituitary
also
Neo-
j.g genistein
castrated
to GlOO
either 49.
study
of this
and pituitary.
sampled
significantly
exhibited
compared.
and was
neonatally
on
postpubertal
blood
from animals
purpose
(GlOO),
of life
showed
of males
The
genistein
1-10
exposed
in SDN
show
that
“androgenizes”
5DN-POA.
INTRODUCTION hormones
differentiated ology [1]. androgens tem (CNS)
g and
decapitation
no difference
in development
Days
Males
groups
killed
exposed
There
early
on
was
Developing
of the hypothalamus
100
derived
Estrogens,
development.
estrogens).
(environmental
(DES),
growth.
(CNS)
and morphology
LH secretion.
were
system
(Zl000)
ng/kg)
estrogen-exposed
animals
effect.
estrogens
g
to DES,
(SDN-POA)
G100
sources
0.1
of brain
pattern
nervous
central
xenobiotic
GnRH-induced
The
differentiated
sexual
on the physiology tg diethylstilbestrol
GnRH
remaining
LH secretion.
preoptic
1000
placed,
increased
and
and
oil,
neonatally
G100
to GnRJI,
Z100
fungal,
estrogens
were
exposed
whereas
GnRH-stimulated
nucleus
plant,
injections
Females
response
uated
from
sexually
the
male
promote
of environmental
heart
to GnRH,
pituitary
the
jg
100
On
determines
of testosterone,
aromatization
determine
the
A complete development
of sexually
behaviors, anatomy, and reproductive physiFemales exposed to estrogens or aromatizable during the critical period of central nervous syssexual development are anovulatory as adults
exhibit
male
patterns
of
behavior
and
assessment
of the
hormonal
reproductive physiology requires an impact of exogenous estrogen exposure. be
detected
in the
urine
mans [9]. In addition moderately potent
of all
to leafy estrogens
influences
on
the
appreciation of Plant estrogens
herbivores,
can
including
plants, some (zearalenone
hu-
fungi produce and related
compounds [10]). Mold on improperly stored corn has been implicated in causing hyperestrogenism in livestock [11], and a recent review suggests that ingestion of fungal estro-
gonadotropin
secretion [2-4]. A morphologic marker of CNS sexual differentiation lies in the intensely staining region of the anterior hypothalamus within the preoptic area (POA)-the sexually dimorphic nucleus (SDN-POA)-which is 3-5-fold larger in males [5]. Castration of neonatal males decrease SDN-POA size [5, 6], whereas androgenization of female rat
gen may have contributed significantly to the decline in rural 19th- and early 20th-century European populations [12]. Phytoestrogens have been implicated as the cause of subfertility in animals grazing on estrogenic pasture [13, 14].
pups with aromatizable creased SDN-POA size
Exposure to estrogenic results in permanent
An estrogenic sponsiveness.
We
androgens [6, 7].
hormonal recently
or estrogens
milieu also demonstrated
tion of DES (1 p.g) to rats on Days 1-10 their subsequent pituitary responsiveness trated adults [8]. Additionally, o,p’-DDT 1-10), an estrogenic isomer creased basal and GnRH-induced not in females.
of
affects that
leads
to in-
pituitary readministra-
as clover tract and
of life decreased to GnRH as cas(0.5 mg, Days
the pesticide LH secretion
DDT, in males,
tems
for with
more than two defeminization,
disease [15]. Although changes the hypothalamic-pituitaiy-ovarian
clover disease phytoestrogens
debut
pasture
infertility
are
not
formation
have on well
regarding
genize the It is our
brain. hypothesis
been reported development understood. the
in the reproductive axis in ewes with [16-19], the neuroendocrine
In addition,
ability that
of
seasons known
there
of phytoestrogens
environmental
effects is no
June
Received
March
1This Research
project
posure
19, 1991. was
26,
1991.
supported
by funding
from
the
Duke
University
DUMC,
Durham,
Medical
School
Fund.
2Correspondence:
Kenneth
A. Faber,
Box
3143
may
developmental we determined marker
NC 27710.
649
of CNS
make
assessment
of
estrogens
its
size
in-
to androandro-
genize the brain and alter CNS regulation of pituitary tion. The sensitivity of the SDN-POA to early estrogen Accepted
of sys-
a
funcex-
marker
of
hormonal toxicity. In the study reported here, the volume of the SDN-POA as an anatomic androgenization
in rats
exposed
neonatally
FABER
650 to either
corn
oil (control),
the estrogenic or
the
estrogenic
none
(100
fungal
or
sponsiveness functional
diethylstilbestrol
plant isoflavone 1 000
resorcyclic
jig).
(DES,
genistein
We
HUGHES Intra-
0.1 g),
(100 or 1000 rig),
acid
also
AND
lactone
assessed
the
to GnRH in the same exposure indicator of hypothalamic-hypophysial
interassay
coefficients
of LH in three
serum
respectively.
zearale-
pituitary
and
surement
re-
groups as a develop-
ment.
and
The
values
Pregnant were
rats
of the
purchased
AND
Charles
at 15-17
air-conditioned ston-Purina,
MO)
schedule
was
CD strain
gestation
with
St. Louis,
The lighting 1900 h EST.
River
days
quarters
METHODS
Purina and
(Raleigh,
and
maintained
Laboratory
Chow
water
available
1 4L: 1OD,
with
ad
NC) in (Ral0500-
maximum possible On Day 49 the animals were promptly removed
ig
Wilton, NH, Lot #8836; 0.1 p.g in corn oil), Biomedicals, Costa Mesa, CA, Lot #224383;
[G100]
(Sigma
or
1 000
Chemical
.tg
[G1000]
Co.,
in corn
St. Louis,
MO,
oil),
gen100
Lot #119F4003;
100
tg
[Z100] or 1000 tg [Zi 000] in corn oil). All pups within a litter received the same treatment. On Day 21 of life, the animals were castrated under ketamine anesthesia (100 mg/kg) were housed in each plexiglass heart kg)
catheterization was
under
performed.
To
and cage,
weaned. and on
ketamine control
anesthesia
for
in saline (50 Blood samples
ng/kg; (0.3
(100
nonspecific
cannulation and fluid injection, the animals ized to receive either saline alone (1 mI/kg) (Factrel, Ayerst Laboratories, Inc., New York, dissolved nulation.
Six siblings Day 42 right
1 mI/kg) ml) were
mg/
effects
and
with
GnRH
Blood ture
and
g for for
antibody and the sity ond
were
10 mm.
later Serum
injection
samples then Sera
RIA. LH
in those
were
that
allowed
centrifuged were
then
concentrations
RIA with a rat National Hormone
LH
aspirated
and
were
measured
1 500
frozen by
of Maryland School of Medicine, Baltimore, antibody (sheep, anti-rabbit) was graciously
X
doubleNIDDKD (Univer-
MD). Secsupplied
University Medical Center, Durham, (50 p.l or less) were assayed in du-
plicate
expressed
RP-3.
the
means
in terms
of NIDDKD-rLH-
the
was
right-sided were
traced area
volumes
were
compared
of which
was
confirmed
SDN
brains in the and of 2
violet acedid not
with
Corte area
the
Student’s falling
by parametric
use
compared
of
CA). of
among
the
groups
by
t-test for norbelow the assay
using the Kruskal-Wallis the Wilcoxon sign-rank with
by Goras il-
Madera, thickness
X
among
and values
sowere
was computed by computed software
was
compared
of variance data. LH
were compared of variance and
and
size,
ratio
sex-specific
cantly There
one-way test. SDN
analysis,
the validity
Kolmogorov-Smirnov
testing.
lated
of males castrated
LH levels
between
GnRH-Stimulated
