Estrogen Receptor Null Mice - Endocrine Reviews

June, 1999 ESTROGEN RECEPTOR NULL MICE 391
gonadotropin promoter sequences of the construct. Furthermore,
the postovariectomy rise in promoter activity could be
prevented by administration of a GnRH antagonist to the
transgenic animal, thereby inhibiting GnRH action at the
gonadotrope. Therefore, a loss of estradiol action via ovariectomy
appeared to result in increased GnRH release from
the hypothalamus, which in turn stimulated increased transcription
of the reporter constructs. However, the possibility
of estradiol actions at the level of the gonadotrope that may
directly effect gene transcription or alter GnRH responsiveness
can not be ruled out.
Regardless of the precise site at which estrogens negatively
regulate gonadotropin expression and release, genetic disruption
of the ER� gene was expected to have effects in the
female hypothalamic-pituitary axis that mimic ovariectomy.
Northern blot analysis of RNA from pituitaries of intact
wild-type and �ERKO females indicates this to be true.
Scully et al. (282) demonstrated that in the �ERKO female, the
levels of the �GSU transcript are elevated 4-fold, whereas
FSH� and LH� mRNAs are as high as 7-fold compared with
wild-type. Ovariectomy in wild-type female littermates produced
elevated gonadotropin subunit mRNAs that approximated
the levels observed in the intact �ERKO, indicating
that the effects of an acute loss of estrogen action are similar
to those produced by a hereditary loss of ER� (282). Therefore,
despite the fact that the hypothalamic-pituitary axis of
the �ERKO female is chronically exposed to elevated levels
of estradiol, the significantly increased level of all three gonadotropin
subunit transcripts resemble those of an ovariectomized
female. These data provide strong support for a
critical role of ER�, rather than ER�, in the negative regulation
of transcription of the gonadotropin subunit genes.
However, at this time, studies of the �ERKO have not provided
data to further elucidate the precise mechanism or site
at which a loss in ER� action has resulted in this effect.
Although the transcript levels of both the LH�, FSH�, and
�GSU are significantly elevated in the �ERKO pituitary, this
effect does not extend to the serum levels of the gonadotropins.
Whereas serum LH is elevated 4- to 7-fold in the adult
�ERKO female, levels of FSH appear to be within the normal
range (Table 2) (252). A similar effect is reported in the PRKO
female mice, although the serum LH levels in this mutant
female are not nearly as elevated as those in the �ERKO
female (387). In addition, ovariectomy in the PRKO female
results in a further elevation of serum LH (387), most likely
due to the loss of serum estrogens. This effect is not observed
in the �ERKO female (252), indicating that estradiol is the
predominant steroid hormone maintaining tonic levels of LH
in the female.
As shown in Table 2, the serum gonadotropin levels in the
�ERKO female indicate that only LH is significantly elevated,
whereas FSH is within the wild-type range. This is in
contrast to the levels of FSH� mRNA in the anterior pituitary
of the �ERKO, which are elevated 7-fold and equal to those
exhibited by an ovariectomized wild-type female. Furthermore,
assays of pituitary homogenates from intact �ERKO
females for FSH protein indicate levels within the wild-type
range, suggesting that the divergence between gene expression
and serum levels for FSH does not appear to be due to
a decreased secretory rate of the hormone but rather at the
level of translation. This is in contrast to the ArKO female
mouse, in which serum levels of both gonadotropins are
reportedly elevated 3-fold compared with the wild-type
(257).
There are a number of possible explanations for this observation
in the �ERKO female. Whereas estradiol treatment
of ovariectomized rats has been reported to completely block
the expected increases in LH� mRNA and LH secretion, it
appears to be only partially effective in reducing transcription
of the FSH� gene and secretion of FSH (376). It is now
known that FSH� gene expression and FSH secretion is selectively
regulated in a positive or negative nature by the
peptides activin and inhibin, respectively (reviewed in Ref.
388). This is illustrated in the knockout mouse model of the
activin receptor type II gene, which exhibits suppressed FSH
levels in the adult of both sexes, supporting a role for activin
in the positive regulation of FSH secretion (248). Although it
is believed that the reciprocal effects of the activin/inhibin
peptides are mediated at the level of the anterior pituitary,
the precise mechanisms of action remain unclear. Inhibin has
been shown to alter the levels of GnRH receptor (389) and
decrease FSH� mRNA levels (390–392), as well as inhibit
translation of FSH� mRNA (391, 393). Activin appears to
utilize similar mechanisms to exert opposite effects on FSH�
mRNA transcription and translation and ultimate secretion
of the hormone (394, 395). Therefore, the normal levels of
FSH in the pituitary and serum of female �ERKO mice may
indicate that disruption of the ER� gene has no effect on the
pattern of activin and inhibin secretion. This is supported by
the apparent negative regulation of FSH� at the translational
level in the �ERKO female, suggesting the presence of an
active inhibin-signaling pathway. Further support is provided
by studies indicating that ovariectomy of the �ERKO
female, and therefore a loss of ovarian inhibin secretion,
results in elevated levels of serum FSH similar to those seen
in ovariectomized wild types (252). Estradiol replacement
was partially effective in reducing the serum FSH in the
ovariectomized wild-type but completely ineffective in the
ovariectomized �ERKO (252). These studies provide for at
least two conclusions: 1) the partial effectiveness of estradiol
in reducing serum FSH levels in the wild-type was likely via
functional ER� in the hypothalamus that may complement
the actions of inhibin in the intact female; and 2) ovarian
factors other than estradiol, most likely inhibin, are maintaining
normal serum FSH levels in the �ERKO female, possibly
by mechanisms that override the loss of ER� action. It
is also possible that both activin and inhibin synthesis and
secretion may be altered in the �ERKO female, but do not
result in a net difference in serum FSH levels. Future investigations
to determine the serum levels of the activin/inhibin
subunit peptides and their functional dimers in the �ERKO
female are warranted.
Another possible explanation for the selective increase in
serum LH in the �ERKO female may be that a lack of ER�
action has resulted in aberrations in the GnRH pulse frequency
and amplitude that are more conducive to LH secretion
from the anterior pituitary. Normal levels of �GSU
transcripts can be maintained with constant GnRH stimulation
of the anterior pituitary (376). However, normal expression
of both gonadotropin �-subunit genes and gonadotro-