Potential role of maternal progesterone in the sexual differentiation of the brain.

In rats, fetal testosterone directs sexual differentiation of the brain. However, fetuses are also exposed to maternal progesterone. Here we report that progestin receptor immunoreactivity in the medial preoptic nucleus (MPN) of fetal and neonatal rats is high in males but virtually absent in females. The MPN is one of the most sexually dimorphic structures in the rat brain and mediates several sexually differentiated behaviors. This suggests that progesterone may play a previously overlooked role in the development of sex differences in the brain and behavior. Henceforth, a novel function of the mother in the sexual differentiation of the CNS must be considered.

Neurophysin-containing pathway from the paraventricular nucleus of the hypothalamus to a sexually dimorphic motor nucleus in lumbar spinal cord.

A model that has been widely used in the study of steroid sensitive neurons, the spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic motor nucleus in the lower lumbar spinal cord that innervates the striated bulbocavernosus (BC) muscle. The BC is responsible for penile reflexes in the male rat, which are important in ensuring pregnancy in females. The characterization of afferents to the SNB aids in the understanding of the neural circuitry involved in reproductive behavior. We have recently identified the paraventricular nucleus (PVN) as a possible source of afferents to the SNB. Because the PVN is the major source of oxytocin/vasopressin within the central nervous system (CNS), the purpose of the present study was to examine and characterize a neurophysin (NP)-containing pathway from the PVN to the SNB. The results demonstrate that neurons of the lateral parvicellular subnucleus of PVN, which project to levels of spinal cord containing SNB motoneurons, contain NP, the coproduct of oxytocin and vasopressin. NP-containing fibers and putative terminals were found in the region of the SNB and appear to contact the soma and proximal dendrites of SNB motoneurons which were retrogradely identified as BC-innervating. Electrolytic lesions, which destroy the lateral parvicellular subnucleus of PVN, abolish NP-containing fibers in the region of the SNB, suggesting that the PVN is the source of these NP fibers. The results of this study indicate a NP-containing projection from the hypothalamus directly to SNB motoneurons. It is suggested that this pathway may play a role in the integration of penile reflexes with other aspects of male copulatory behavior that are under hypothalamic control.

Distribution and steroid hormone regulation of aromatase mRNA expression in the forebrain of adult male and female rats: a cellular-level analysis using in situ hybridization.

Many of the effects of gonadal steroid hormones in the male brain are due to the actions of the testosterone metabolite estradiol, which is synthesized by the actions of the P450 enzyme aromatase. Aromatase activity is present in regions of the preoptic area, hypothalamus, and limbic system. Levels of aromatase activity in the brain are highly dependent on gonadal steroid hormones in many brain regions, but not all. We examined the distribution of aromatase mRNA in adult male and female rat brains as well as the regulation of the levels of aromatase mRNA in the brains of males by gonadal steroid hormones using in situ hybridization. This method was performed using a 35S-labelled cRNA probe, transcribed in vitro from the rat ovarian aromatase cDNA. In the adult male, many heavily labelled cells were found in the encapsulated bed nucleus of the stria terminalis (BNST), the medial preoptic nucleus (MPN), the ventromedial nucleus (VMN), the medial amygdala (mAMY), and the cortical amygdala (CoAMY). The regional distribution of aromatase mRNA was similar in females, but females tended to have a lower number of aromatase mRNA expressing cells in each region compared to males. Aromatase mRNA levels in the BNST, MPN, VMN, and mAMY tended to be lower in castrated males than in intact males, whereas aromatase mRNA levels were unaltered by castration in the CoAMY. The degree of reduction in mean levels of aromatase mRNA following castration does not simply account for the large changes measured in activity following castration. Examination of the entire population of individual cells expressing aromatase mRNA in castrated males suggests that aromatase mRNA may be regulated by steroid hormones differentially in specific populations of neurons within regions where activity is known to decrease following castration.

Evidence for estrogen receptor in cell nuclei and axon terminals within the lateral habenula of the rat: regulation during pregnancy.

The habenular complex is involved in several estrogen-dependent reproductive behaviors in female rats, namely, sexual behavior, maternal behavior, and postpartum sexual behavior. Although it is known that estrogen acts in other brain regions to mediate these behaviors, it is not known whether estrogen may also act directly on the habenular complex. To address this possibility, we examined this region for the presence of estrogen receptor (ER). This analysis was carried out in separate experiments by using in situ hybridization, immunocytochemistry at the light and electron microscopic levels, and steroid autoradiography. Neurons within the lateral habenula (LHb), but not the medial habenula, express ER mRNA, contain ER immunoreactivity (ER-ir) in their nuclei, and concentrate radiolabelled estradiol, providing strong evidence for the presence of functional ER in the lateral habenula. There were also ER-ir containing punctate fibers within the LHb, which, at the electron microscopic level, in part, proved to be axons and presynaptic axonal terminals. Both the level of ER-ir in cell nuclei and the density of ER-ir fibers within the LHb were regulated during the course of pregnancy and the postpartum period, suggesting that the sensitivity of the LHb to estrogen may be altered during this time. Taken together, these results demonstrate that the LHb is likely a more estrogen-sensitive region than was previously considered, and they suggest alternative mechanisms of action for ER. ER within the LHb may play a critical role in the involvement of the LHb in estrogen-dependent female reproductive behaviors.

Hormonal priming and triggering of maternal behavior in the rat with special reference to the relations between estrogen receptor binding and ER mRNA in specific brain regions.

Estrogen stimulation of maternal behavior during pregnancy in the rat has been studied at several levels of analysis. These include (a) changes in maternal responsiveness during pregnancy; (b) hormonal stimulation of maternal behavior; and (c) correlation between nuclear binding of estradiol in the medial preoptic area and the stimulation of maternal behavior (i.e., in pregnancy-terminated, ovariectomized females treated with estradiol benzoate). These studies have given rise to the concepts of hormonal priming and triggering of maternal behavior during pregnancy and at parturition. More recently, using in situ hybridization, ER mRNA was measured during pregnancy (also diestrus and postpartum) in brain regions in which binding previously had been studied, to investigate further the regulation of hormonal priming. Steady state levels of ER mRNA per cell and cell densities of ER mRNA produced a measure of total ER mRNA per brain region which was then compared to nuclear estrogen receptor binding. The relation between binding and ER mRNA is presented for one of the brain regions, the rostral medial preoptic nucleus. The results indicate that ER transcription is regulated during pregnancy, but regulation is specific to each brain region and there is no simple relation between ER mRNA and nuclear estrogen receptor binding.

In situ analysis of estrogen receptor mRNA expression in the brain of female rats during pregnancy.

Estrogen's action in specific brain regions, particularly the medial preoptic nucleus (MPN), is necessary for the onset of maternal behavior in the pregnant female rat. There is an increase in estrogen binding in the MPN during pregnancy, and it has been hypothesized that this increase is part of the mechanism by which the brain is readied to support estrogen-dependent maternal behavior. This experiment determines whether an alteration in the levels of estrogen receptor mRNA precedes the increase in estrogen binding to its receptor. Using in situ hybridization, estrogen receptor (ER) mRNA levels were measured in specific brain regions in females on day 8, 16 or 22 of pregnancy or on postpartum day 1 or in non-pregnant females. ER mRNA levels are significantly higher in the MPN in females on day 8 of pregnancy compared with non-pregnant females or with females on day 16. In the ventromedial nucleus, which is important for estrogen's role in postpartum sexual receptivity, there was an increase in ER mRNA levels on day 22 of pregnancy compared with day 16 of pregnancy. These results suggest that ER levels may increase in specific, behaviorally relevant brain regions at critical times during pregnancy through regulation of ER mRNA levels.

Levels of estrogen receptor immunoreactivity are altered in behaviorally-relevant brain regions in female rats during pregnancy.

Pregnancy and parturition are accompanied by unique behavioral changes. Only some of the neural mechanisms behind the dramatic changes in behavior are understood. Estrogen's action within the medial preoptic nucleus (MPN) is necessary for the induction of maternal behavior around the time of parturition, and estrogen acts within the ventromedial nucleus (VMN) to trigger postpartum sexual receptivity shortly after parturition. We have hypothesized that the sensitivity of various brain regions to estrogen may be altered by pregnancy to support these unique behavioral patterns. Using immunocytochemistry, this study examined whether the levels of estrogen receptor (ER) protein, within behaviorally relevant brain regions, differ among females on day 8, day 16, and day 22 of pregnancy, or on postpartum day 1. On day 16 and day 22 of pregnancy, the MPN contained a significantly greater number of cells expressing high levels of ER-ir compared to day 8 or postpartum day 1. In the VMN, the mean amount of ER-ir per cell was significantly higher on day 22 of pregnancy than on day 16 or postpartum day 1. In the bed nucleus of the stria terminalis, ER-ir levels were significantly increased on postpartum day 1 compared to day 22 of pregnancy. There were no significant changes in ER-ir in the medial amygdala. These results demonstrate regionally and temporally specific regulation of ER protein in the brain during pregnancy. Alterations in the levels of ER at critical times in regions such as the MPN and VMN may underlie the unique expression of maternal and sexual behavior that occur during pregnancy and at the time of parturition.

Neurons in the paraventricular nucleus of the hypothalamus that project to the sexually dimorphic lower lumbar spinal cord concentrate 3H-estradiol in the male rat.

The location and distribution of estradiol-concentrating neurons in the hypothalamus afferent to segments of lumbar spinal cord that contain the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB) were determined by combining retrograde fluorescent tract tracing with steroid hormone autoradiography. Injections of Fluorogold were made into segments of L5-L6 of the spinal cord of adult male rats and 12 days later animals were castrated. One week following castration, males received injections of [3H]estradiol and were perfused. Their brains were then processed for steroid hormone autoradiography. Following exposure times of 11 to 12 months, autoradiograms were developed and the hypothalamus was analyzed for neurons that concentrate estradiol and project to the spinal cord. Numerous neurons in the hypothalamus projected to the spinal cord, specifically neurons in the paraventricular nucleus (PVN), the lateral hypothalamus and the dorsal area of the hypothalamus. Although many subnuclei of PVN, as well as lateral hypothalamus, contained Fluorogold labelled neurons and estradiol concentrating neurons, the majority of double labeled cells were found in the lateral parvocellular (LP) subnucleus of PVN. Approximately 30% of the neurons in the lp subnucleus that projected to spinal cord also concentrated estradiol. Up to one half of the estradiol-concentrating neurons in lp sent axons to the lower lumbar spinal cord. These results suggest that some of the effects of gonadal steroid hormones on SNB development, plasticity and function may in fact, be indirect, via steroid-sensitive afferents.

Regulation of sex differences in progesterone receptor expression in the medial preoptic nucleus of postnatal rats.

The medial preoptic nucleus (MPN) of the rat, an excellent model for understanding the mechanisms involved in sexual differentiation, is highly sensitive to gonadal hormones during both pre- and post-natal life. Progesterone receptor (PR) expression is sexually dimorphic in the prenatal MPN. Males have significantly higher levels of PR-immunoreactivity (PRir) than females from approximately embryonic day 19 through at least the day of birth, suggesting that PR may play a role in sexual differentiation. Because the MPN is still sensitive to steroid hormones postnatally, the present study investigated PR expression in the MPN of males and females after birth using immunocytochemistry. Results indicate that a sex difference in PR expression persists until at least postnatal day (P) 28. However, females begin to express PR around P10. Because oestradiol regulates PR expression in the adult brain, this study also examined the influence of gonadal hormones on PR expression in the neonatal male and female MPN. Castration on the day of birth significantly reduced levels of PRir in the MPN by 24 h following surgery. Ovariectomy on P4, before the onset of ovarian steroidogenesis, prevented the induction of PR expression in the female MPN, observed in controls by P13. In both sexes, the presence of PRir in the MPN is dependent on gonadal hormone exposure. These findings suggest that differences in steroid secretion by the neonatal male and female gonads are responsible for producing sex differences in the level of PR expression in the postnatal MPN.

Effects of neonatal RU486 on adult sexual, parental, and fearful behaviors in rats.

Exposure to gonadal hormones during perinatal life influences later behavior. The finding that sex differences exist in progestin receptor expression in the perinatal rat brain suggests differential sensitivity of male and female brains to progesterone (C. K. Wagner, A. N. Nakayama, & G. J. De Vries, 1998). Because these sex differences are in neural sites that influence sexually differentiated sexual, parental, and fearful behaviors in adults, this study examined the effects of administering the progestin receptor antagonist RU486 for the first 10 days after birth on these behaviors in adulthood. Neonatal RU486 significantly reduced sexual behavior in males but did not impair reproduction in females. Neonatal RU486 did not affect parental responses of virgin rats exposed to pups (sensitization) but reduced fear in the elevated plus-maze in both sexes. Treatment of pups with RU486 affected neither mother-litter interactions nor plasma testosterone levels in males during or after treatment. These results suggest that neonatal exposure to progesterone, in addition to androgens and estrogens, influences behavioral development in rats.

Neuroanatomical distribution of aromatase MRNA in the rat brain: indications of regional regulation.

Aromatase, the enzyme responsible for the conversion of testosterone to estradiol, is found in the rat brain and is present in regions of the preoptic area, hypothalamus, and limbic system. Gonadal steroid hormones regulate aromatase activity levels in many brain regions, but not all. Using in situ hybridization, we examined the distribution of aromatase mRNA in the adult male forebrain, as well as the levels of aromatase mRNA in the brains of males and females, and the regulation by gonadal steroid hormones. In the adult male, many heavily labelled cells were found in the encapsulated bed nucleus of the stria terminalis (BNST), the medial preoptic nucleus (MPN), the ventromedial nucleus (VMN), the medial amygdala (mAMY) and the cortical amygdala (CoAMY). The regional distribution of aromatase mRNA was similar in males and females, but males tended to have a greater number of aromatase mRNA-expressing cells in each region compared to females. Aromatase mRNA levels in the BNST, MPN, VMN and mAMY tended to be lower in castrated males than in intact males, whereas aromatase mRNA levels were unaltered by castration in the CoAMY. Further analysis of individual cells expressing aromatase mRNA suggests that aromatase mRNA may be regulated by steroid hormones differentially in specific populations of cells in regions where enzyme activity levels are steroid-hormone-dependent.

Projections of the paraventricular nucleus of the hypothalamus to the sexually dimorphic lumbosacral region of the spinal cord.

Lumbar regions L5-L6 of the spinal cord of the male rat contain the sexually dimorphic motor nuclei, the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN), which innervate perineal muscles, the bulbocavernosus and the ischiocavernosus, respectively. This neuromuscular system controls penile reflexes which are essential to male reproductive success. Oxytocin has been shown to induce penile reflexes and the site of action for these effects is the PVN. Since PVN is known to project to cervical and thoracic levels of spinal cord, the present study examined projections of the PVN to the L5-L6 region of the spinal cord. WGA-HRP was injected into the region of L5-L6, aimed at the SNB and DLN and their dendritic extents, in intact male, castrated male and female rats. WGA-HRP-labelled cells bodies were found in the parvocellular subnuclei of PVN, as well as regions of the lateral hypothalamus and the dorsal area of the hypothalamus. These results demonstrate that the PVN projects to lumbar levels of the spinal cord that are sexually dimorphic and androgen-dependent. This suggests that PVN may modulate the activity of these motoneurons.

Anatomical organization of the sexually dimorphic perineal neuromuscular system in the house mouse.

The anatomy of the sexually dimorphic motoneuron nuclei of the bulbocavernosus (BC) and ischiocavernosus (IC) muscles, as well as the non-sexually dimorphic external and sphincter (EAS), was examined in hybrid B6D2F1 mice using the retrograde tracer, cholera toxin-bound horseradish peroxidase. Motoneurons innervating the BC were located in the dorsomedial nucleus (DM), as well as in the ventral nucleus (V) and in the mid-region of the ventral horn (MVH). Following injections restricted to the IC, labelled neurons were found in the dorsolateral nucleus (DL), as well as in V, DM and MVH. Cells innervating the EAS in both males and females were located in the DM, as well as in the V and MVH. An elaborate network of dendrites extended between all labelled nuclei. The present results demonstrate that the anatomical specificity of the sexually dimorphic neuromuscular system of the mouse differs from that observed in other species.

Neurochemical identification of A13 dopaminergic neuronal projections from the medial zona incerta to the horizontal limb of the diagonal band of Broca and the central nucleus of the amygdala.

Studies utilizing fluorescent histochemical techniques first revealed that A13 dopaminergic (DA) perikarya located in medial zona incerta (MZI) project to various regions within the hypothalamus; accordingly, these DA neurons were designated the 'incertohypothalamic' DA neuronal system. More recently, it has been shown that the anterograde neuronal tract tracer Phaseolus vulgaris leucoagglutinin, after injection into MZI, is identified in nerve terminals outside of the hypothalamus; for example, in horizontal limb of the diagonal band of Broca (HDB) and central nucleus of the amygdala (cAMY). The purpose of the present study was to determine, using neurochemical techniques, if A13 DA neurons project to the HDB and cAMY. Concentrations of dopamine and one of its metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) were determined in HDB and cAMY following: (1) electrical stimulation of MZI, (2) electrolytic lesion or knife ablation of MZI, and (3) administration of gamma-hydroxybutyric acid (GHBA) into MZI. For comparison, similar measurements were made in nucleus accumbens (N. Acc.), a terminal region of A10 DA neurons in the ventral tegmental area (VTA). Electrical stimulation of MZI increased DOPAC concentrations in HDB and cAMY, whereas electrolytic or ablative lesions of MZI decreased dopamine concentrations in both of these regions. By contrast, neither stimulation nor lesion of MZI had any effect on DOPAC or dopamine concentrations in N. Acc. Intracerebral injection of GHBA into MZI increased dopamine concentrations in MZI and HDB, but not in cAMY or N. Acc. Intracerebral administration of GHBA into VTA increased dopamine concentrations in HDB and N. Acc., but not in MZI or cAMY.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcitonin gene-related peptide-like immunoreactivity in spinal motoneurons of the male mouse is affected by castration and genotype.

Calcitonin gene-related peptide (CGRP) is found in motoneurons of the mammalian spinal cord, including motoneurons of the androgen-dependent spinal nucleus of the bulbocavernosus (SNB) of the mouse. Motoneurons of the SNB innervate the bulbocavernosus (BC), a striated muscle involved in penile reflexes. CGRP is though to be a trophic factor produced by motoneurons to regulate the expression of the acetylcholine receptor at the neuromuscular junction. In rats, the number of SNB motoneurons containing CGRP is increased by gonadal steroids. This regulation appears to rely on an activity-dependent factor produced by the BC muscle. The purpose of the present study was to examine, using immunohistochemistry, the steroid dependence of CGRP in the SNB of male house mice. Genotypic differences in the steroid regulation of CGRP immunoreactivity were examined in three strains of mice that differ in their behavioral sensitivity to castration. The results demonstrate that castration reduces the number of CGRP-positive SNB motoneurons in mice. The magnitude of the change in CGRP in response to castration and the length of time required following castration to alter CGRP were dependent on genotype. Interestingly, the effect of castration in mice, to reduce the number of CGRP-immunoreactive SNB motoneurons, is opposite in direction from the increase in CGRP SNB motoneurons observed in rats observed following castration. These experiments suggest that androgens may alter neuromuscular junction function of mouse SNB by regulating the production of CGRP in a species-specific, genotypically dependent fashion.

Efferent projections from the region of the medial zona incerta containing A13 dopaminergic neurons: a PHA-L anterograde tract-tracing study in the rat.

Potential efferent projections of A13 dopaminergic (DA) neurons were identified in the present study by examining the distribution of labelled fibers following iontophoretic injection of the anterogradely transported lectin Phaseolus vulgaris leucoagglutinin (PHA-L) into the medial zona incerta (MZI), the region of the diencephalon containing A13 DA neuronal perikarya. One week after injection, PHA-L labelled fibers were found throughout the brain with the heaviest labelling occurring ipsilateral to the injection site in the anterior hypothalamic area, lateral hypothalamus, lateral preoptic area, horizontal diagonal band of Broca, and parvocellular region of the paraventricular nucleus. Moderate labelling was observed in the ipsilateral median preoptic nucleus, lateral septum, lateral aspect of the bed nucleus of the stria terminalis, and central nucleus of the amygdala. Moderate labelling was also found in the contralateral MZI and parvocellular region of the paraventricular nucleus. Light labelling was detected in the ipsilateral medial preoptic area, supraoptic nucleus, ventromedial nucleus, arcuate nucleus, vertical limb of the diagonal band of Broca, and in the contralateral lateral hypothalamus. Few immunopositive fibers were present in the dorsomedial nucleus of the hypothalamus or the magnocellular region of the paraventricular nucleus. These results reveal that neurons located in the MZI (possibly A13 DA neurons) have ipsilateral efferent axonal projections to a variety of brain regions including the lateral hypothalamus, lateral preoptic area, and the limbic structures at the diencephalic-telencephalic juncture.

Perinatal modification of a sexually dimorphic motor nucleus in the spinal cord of the B6D2F1 house mouse.

The sexually dimorphic dorsomedial nucleus (DM) of the spinal cord of mice is affected by gonadal steroids in adulthood and these effects are dependent upon genotype. Following castration of adult mice there is a decrease in DM cell size in DBA/2J and hybrid B6D2F1 strains and a decrease in the number of cells staining with thionin in C57B1/6J and B6D2F1 strains. The effects of androgens on development of the DM nucleus were examined in B6D2F1 mice, which exhibit both characteristics in adulthood. Testosterone propionate (TP) administered to females pre- or postnatally resulted in a significantly larger number of motoneurons in the region of the DM when compared to administration of the vehicle alone, while soma area remained unchanged. Adult males castrated on the day of birth had significantly fewer cells in the DM than did intact males. Differences in cell size between shams and castrates were dependent upon age.

Anatomically-specific actions of oestrogen receptor in the developing female rat brain: effects of oestradiol and selective oestrogen receptor modulators on progestin receptor expression.

Steroid hormones largely exert their actions by activating nuclear receptors, which, as transcription factors, powerfully influence fundamental processes of neural development. Often, steroid receptor action demonstrates remarkable specificity under different developmental, anatomical or hormonal conditions. Yet, the mechanisms underlying such specificity are poorly understood. The present study examined the anatomically-specific regulation of progestin receptor (PR) expression by oestrogen receptor (ER) activation in the ventromedial nucleus (VMN) of the hypothalamus and the medial preoptic nucleus (MPN) of the neonatal female rat brain, using the selective oestrogen receptor modulators (SERMs), tamoxifen and ICI 182780 (ICI), in the presence or absence of oestradiol benzoate (EB) treatment. The results demonstrate that PR immunoreactivity (PR-ir) in the neonatal female MPN was significantly increased by EB and this increase was abolished by either tamoxifen or ICI treatment. In contrast, within the VMN of the same animals, EB had no effect on PR-ir and the SERMs only modestly decreased PR-ir. Interestingly, ICI acted as a true antagonist regardless of EB treatment, whereas tamoxifen acted as an ER agonist in the absence of EB in the MPN, but not the VMN, representing one of the first in vivo demonstrations of tissue-specific and oestradiol-independent effects of tamoxifen on ER activation. The present results indicate that PR expression is highly dependent on oestradiol and its receptor in the MPN, although it is independent of both oestradiol and ER activation within the neonatal VMN. These findings demonstrate the anatomically-specific actions of oestradiol and its receptor to induce PR in two brain regions controlling different aspects of female reproductive behaviours in adulthood.

Maternal hypothyroidism decreases progesterone receptor expression in the cortical subplate of foetal rat brain.

Steroid hormones exert profound effects on the development of brain areas controlling complex cognitive function in adulthood. One class, progestins, may contribute by acting on the progestin receptor (PR), which is transiently expressed in a critical layer of developing cortex: the subplate. PR expression in the subplate coincides with the establishment of ongoing cortical connectivity and may play an important organisational role. Identification of the factor(s) that regulate the precise timing of PR expression within subplate may help elucidate the function of PR. Thyroid hormone may interact with hormone response elements within the PR gene. The present study examined the effects of maternal hypothyroidism on levels of PR immunoreactivity (PR-IR) within the foetal subplate. Pregnant rats were made hypothyroid by the administration of methimazole and potassium perchlorate in drinking water. Maternal hypothyroidism significantly decreased PR-IR within the foetal subplate. Using the incorporation of 5-bromo-2'-deoxyuridine (BrDU) during subplate cell neurogenesis (embryonic day 13.5) to determine subplate cell survival in hypothyroid animals, we found that decreases in PR-IR cannot be attributed to significant subplate cell loss but are more likely the result of altered PR expression. Gestational thyroxine replacement to hypothyroid dams prevented the decrease in PR-IR within the subplate. These results identify thyroid hormone as a potential factor in the regulation of PR expression in the developing brain. These results are consistent with the idea that endocrine cross-talk between progesterone and thyroid hormone may be one mechanism by which maternal hypothyroidism alters normal cortical development.