Steroid autoradiography of the sexually dimorphic nucleus of the preoptic area

Autoradiography was performed to determine if the neurons of the sexually dimorphic nucleus of the preoptic area (SDN-POA) in the adult rat accumulate estradiol (E2), testosterone (T), and/or dihydrotestosterone (DHT). Three days prior to steroid administration, adult male and female Sprague-Dawley rats were gonadectomized and adrenalectomized. Animals were then given either [3H]T, [3H]E2, or [3H]DHT through an indwelling jugular cannula. One hour later, animals were decapitated and brain sections processed for thaw mount autoradiography. The autoradiograms which contained the SDN-POA and an adjacent area of the medial preoptic area (MPOA) were quantitatively analyzed using the 3 times background, 5 times background, and Poisson criteria for labeled cells. In general, cells in the SDN-POA and the MPOA accumulate T, E2, or DHT. For both sexes, there is a greater percentage of labeled cells in the SDN-POA than in the MPOA, and a greater percentage of labeled cells following E2 exposure than following T or DHT exposure. In addition, there is a sex difference (male greater than female) in the percentage of labeled cells following T exposure. In summary, these data indicate that adult SDN-POA neurons do accumulate gonadal steroids.     

Neurogenesis of the sexually dimorphic nucleus of the preoptic area in the rat

The sexually dimorphic nucleus of the preoptic area (SDN-POA) of Sprague-Dawley rats is larger in volume in the male and hormone-dependent early in postnatal life. In the present study, we compared for each sex the time course of neuroblast proliferation which forms SDN-POA or adjacent medial preoptic area (MPOA) neurons. Additionally, we investigated whether there is a temporal gradient of production of neurons in relation to their final position within the SDN-POA. On day 14, 15, 16, 17, or 18 postfertilization (pf) pregnant rats were given a single injection of ³H-thymidine (*thy). At 30 postnatal days of age the pups were sacrificed and brain sections were prepared and processed for autoradiography. Three sections of the SDN-POA and an adjacent area just lateral to it in the MPOA wer also analyzed. In the MPOA and the SDN-POA the percentage (%) of labeled neurons decreases as the day of injection of *thy approaches the end of gestation, but the time period in which neuroblast divisions occurred is markedly different for the SDN-POA as compared to that for the MPOA. DNA synthesis occurs as late as day 18pf for neurons which form the SDN-POA but ceases on day 16pf for those destined for the MPOA. There is a sex difference in neuronal production on both day 14 and 17pf for neurons destined for the SDN-POA. After injection on day 14pf the % labeled neurons is larger in the female than in the male but after injection on day 17pf this is reversed. There are also significant sex differences as well as a temporal gradient associated with the % labeled neurons in the SDN-POA in relation to their final anterior-posterior position. In addition, this study confirms our previous results which justify labeling the SDN-POA a nucleus, since neuronal density in this region in the male and female is significantly greater than that in the surrounding MPOA. These data illustrate that the specific neurons which comprise the SDN-POA in both the male and female are being produced as late as day 18pf, whereas neurons located in the MPOA but not in the SDN-POA have all been born by day 16pf. Neuroblast division which produces the neurons of the SDN-POA may begin earlier and terminate sooner in the female than in the male. These differences in neuronal production may partially account for the sexual dimorphism seen in the volume and neuronal number of the SDN-POA of the adult rat.     

Ontogeny of the sexually dimorphic nucleus of the preoptic area

An area of increased cell density in the medial preoptic area of the adult rat brain which is markedly larger in volume in the male has been named the Sexually Dimorphic Nucleus of the Preoptic Area (SDN-POA). It has been further shown that the presence of the testes or exogenous androgen during the first week of postnatal life significantly increases the volume of the SDN-POA in the brain of the adult. The present study was conducted to describe the time course of the prenatal and postnatal development of the SDN-POA in the intact male and female rat. Sprague-Dawley females were housed with males on proestrus. The presence of sperm in the vaginal smear on estrus was used to define day 1 of gestation. Male and female prenatal and postnatal pups were sacrificed and perfused with 10% neutral formalin on days 20, 21, 22, 23, 24, 25, 26, 27, 28, 30, and 32 postfertilization. Following histological sectioning at 60 m?m and staining with thionin, three investigators independently drew the boundaries of the SDN-POA on successive sections, using a microprojector at a magnification of 43.5. A fourth investigator averaged the three drawings; from this average, the nuclear volume was determined with a calibrated planimeter. All drawings and measurements were performed without knowledge of age or sex since the brains were coded according to a random number generator. The volume of the SDN-POA was found to be significantly larger in males than in females on days 23, and 25 through 32. Moreover, the volume of the SDN-POA increased significantly with age in the male, but there was no change in SDN-POA volume in the female. In order to test the specificity of the sexual dimorphism of the SDN-POA, five different linear measurements were taken of brain size. There was a significant increase in these parameters in both sexes with increasing age; however, there was no significant sex difference found. Thus, the sex difference in volume of the SDN-POA cannot be accounted for by sex differences or age-related changes in brain size per se. These data suggest that the development of the SDN-POA (as measured by volume) is itself sexually dimorphic. There are dramatic increases in the male, but not in the female, during a time-period which is known to be critical for sexual differentiation of the brain.     

Evidence for the existence of a sexually dimorphic nucleus in the preoptic area of the rat

The volume of an intensely staining component of the preoptic area of the male rat is markedly larger than that of the female. Moreover, its volume in both sexes is altered by perinatal hormone exposure consistent with the view that this brain region undergoes hormone dependent sexual differentiation. The present study was carried out to determine if this sexually dimorphic area of the brain has a greater cell density than that of the surround, and if a unique population or distribution of cells, either within one sex or between males and females, characterized this region. A single coronal paraffin section (10 m?m) through the approximate center of this sexually dimorphic area in four adult gonadectomized rats of each sex was evaluated systematically. Each cell was labelled as being inside or outside of the sexually dimorphic area. In addition to cell density per unit area the following parameters were evaluated through a closed circuit video system: cell size, staining intensity, shape, and the presence of processes and of a nucleolus. The presence of a nucleolus was further used to identify neurons within the total population of almost 5000 cells that was evaluated. In both sexes, the sexually dimorphic area was characterized by a significantly increased cell density per unit area compared to that of the surround. On this basis, the term, the Sexually Dimorphic Nucleus of the Preoptic Area (SDN-POA) is proposed, for this region. Moreover, the SDN-POA of the male was characterized by increased neuronal density per unit area. The SDN-POA in the male was also found to contain larger cells and neurons, as determined by direct measurement of their greatest diameter, as well as a greater percentage of cells and neurons rated large on a three-point scale (small, medium, and large). No consistent differences in frequency distribution by stain intensity, shape, or the presence of cell processes were found to characterize the SDN-POA or contribute to the sexual dimorphism. It is concluded that the marked sex difference in the volume of the SDN-POA is due principally to an increase in the male of the total area of higher cell and neuronal density. However, the present results do not eliminate the possibility that more subtle differences in neuronal characteristics may exist in the SDN-POA.     

Sex difference in dendritic development of the sexually dimorphic nucleus of the preoptic area in the rat

Sex differences in the growth and dendritic development of neurons in the sexually dimorphic nucleus of the preoptic area were examined with quantitative Golgi techniques during early postnatal life in rats. Neuronal size and dendritic extent were found to increase more in males than in females during the first 10 postnatal days, while the numbers of primary and terminal dendrites were similar in the two sexes. The onset of greater dendritic growth in males occurs just after the volume of the nucleus begins to exhibit sexual dimorphism, between 24 and 26 days after fertilization. Growth of dendrites in this region may be related to the presence of sex hormones during the critical period of sexual brain differentiation.     

Development of the sexually dimorphic nucleus of the preoptic area and the influence of estrogen-like compounds

One of the wel -defined sexual y dimorphic structures in the brain is the sexual y dimorphic nucleus, a cluster of cells located in the preoptic area of the hypothalamus. The rodent sexual y dimorphic ...     

Ontogeny of a sexually dimorphic opioid system in the preoptic area of the rat

A striking sexual dimorphism exists in the distribution of Met-enkephalin (m-ENK) immunoreactive fibers in the preoptic area of the rat brain. A dense plexus of m-ENK fibers, approximately 100 microns wide, is present in the periventricular part of the preoptic area (pePOA) in adult females, but not in males. In the present study, we have examined the time of first expression of this female-typical system during the course of normal brain development. The female-typical plexus of m-ENK fibers in the pePOA is not expressed prepubertally, but first becomes evident during the late peri-pubertal period (usually by 40 days of age). In adult females, the maintenance of immunohistochemically detectable levels of m-ENK in this fiber system is dependent upon the presence of gonadal steroids, especially estradiol. Therefore, we examined whether exposure of pre-pubertal females to estradiol would result in precocious expression of the m-ENK fiber plexus. The results of this experiment demonstrate that exposure to estradiol for 7 days induces the full expression of the m-ENK fiber system in the pePOA of prepubertal females, such that it was indistinguishable from that seen in mature animals. These results demonstrate that while the sexually dimorphic m-ENK system of the pePOA is not normally expressed prepubertally, the neural substrate is nevertheless in place and capable of being activated by exposure to exogenous estradiol.     

Implication of testosterone metabolism in the control of the sexually dimorphic nucleus of the quail preoptic area

In quail, testosterone (T) activates male copulation and affects the volume and cytoarchitectonic organization of the medial preoptic nucleus (POM). T metabolism (especially its aromatization) is critical for the production of these behavioral effects. We wondered whether T metabolism is also playing a role in the induction of the morphological changes in POM. We compared the effects of T and of its metabolites in this nucleus. To obtain an independent evaluation of the role played by aromatase, morphological effects of T associated or not with the aromatase inhibitor R76713 were also assessed. As previously observed, T increased the POM volume and the cross-sectional area of the neurons in the lateral part of the nucleus. The effects of T on the neurons in the lateral POM were mimicked in part by the combined treatment with estradiol and 5α-dihydrotestosterone. They were also blocked by the aromatase inhibitor. This suggests that T aromatization plays a critical role in the mediation of the cytoarchitectonic effects of T. A specific role for androgens alone remains to be established.     

Intrauterine proximity to male fetuses affects the morphology of the sexually dimorphic nucleus of the preoptic area in the adult rat brain

Previous studies on polytocous rodents have revealed that the fetal intrauterine position influences its later anatomy, physiology, reproductive performance and behavior. To investigate whether the position of a fetus in the uterus modifies the development of the brain, we examined whether the structure of the sexually dimorphic nucleus of the preoptic area (SDN-POA) of rat brains accorded to their intrauterine positions. Brain sections of adult rats gestated between two male fetuses (2M) and between two female fetuses (2F) in the uterus were analysed for their immunoreactivity to calbindin-D28k, which is a marker of the SDN-POA. The SDN-POA volume of the 2M adult males was greater than that of the 2F adult males, whereas the SDN-POA volume of the 2M and 2F adult females showed no significant difference. This result indicated that contiguous male fetuses have a masculinizing effect on the SDN-POA volume of the male. To further examine whether the increment of SDN-POA volume in adulthood was due to exposure to elevated steroid hormones during fetal life, concentrations of testosterone and 17beta-estradiol in the brain were measured with 2M and 2F fetuses during gestation, respectively. On gestation day 21, the concentrations of testosterone and 17beta-estradiol in the brain were significantly higher in the 2M male rats as compared with the 2F male rats. The results suggested that there was a relationship between the fetal intrauterine position, hormone transfer from adjacent fetuses and the SDN-POA volume in adult rat brains.     

Sexual differentiation of the human hypothalamus: ontogeny of the sexually dimorphic nucleus of the preoptic area

Sexual differentiation of the hypothalamus of the human brain is generally believed to take place around midpregnancy and thought to be related to the development of sexual orientation and gender identity. The present life span study on the human sexually dimorphic nucleus (SDN) of more than a hundred subjects revealed, however, that at the age of 2-4 years the SDN cell number reaches a peak value, and that only after this age sexual differentiation becomes manifest. Furthermore, the SDN cell number in homosexual men was not different from that of the male reference group, but significantly larger than the cell number in age-matched women.     

Cholecystokinin synapses in the sexually dimorphic central part of the medial preoptic nucleus

The distribution of immunoreactive cholecystokinin synapses in the medial preoptic nucleus was studied. At the light microscopic level, the relative density of cholecystokinin in the nucleus corresponds to its three subdivisions. The sexually dimorphic central part of the nucleus was filled with a very dense plexus of cholecystokinin fibers/terminals. At the ultrastructural level, cholecystokinin-positive boutons contained labeled, dense core vesicles and clear vesicles as well as a population of unlabeled vesicles. The cholecystokinin boutons formed axodendritic, axosomatic, and axoaxonic interactions. Very few of the boutons were found in apposition with unlabeled elements but without any evidence of contact, i.e., synaptoid-type. Interestingly, cholecystokinin-positive, dendrite-like structures occasionally extended thin processes that were observed to end in a bouton containing both light and dense core vesicles. These varicosities were observed to have synaptoid endings. The dense innervation and synaptoid interaction of cholecystokinin elements in the central part of the medial preoptic nucleus may help to clarify the peptide's function in the medial preoptic area.     

Ontogeny of a sexually dimorphic nucleus in the preoptic area of the Japanese quail (Coturnix japonica).

The nucleus preopticus medianus (POMn) is a sexually dimorphic nucleus in Japanese quail (Coturnix japonica) that is critically involved in the hormonal activation of male copulatory behavior. The larger volume apparent in males appears to depend upon circulating testosterone [Brain Res., 416 (1987) 59-68; J. Comp. Neurol., 303 (1991) 443-456]. The present study determined when during normal development this nucleus becomes dimorphic. POMn and a control nucleus, the nucleus commissurae pallii (nCPa), were traced from Nissl-stained coronal sections (40 microns) from animals sacrificed at 2, 3, 4, 5, 6 or 7 weeks of age. Areas were measured and used to calculate volume. POMn volumes were not significantly different in males and females through 5 weeks of age. The dimorphism in POMn volume then became apparent at 6 weeks of age as a function of an increase in male POMn volume between 5 and 6 weeks of age. No significant differences were apparent at any developmental age in nCPa volume. The appearance of a sexual dimorphism in POMn volume is coincident with the pubertal surge in testosterone that occurs between 5 and 6 weeks of age [Horm. Behav., 11 (1978) 175-182], and is also coincident with behavioral sexual maturity.     

Perinatal androgenization prevents age-related neuron loss in the sexually dimorphic nucleus of the preoptic area in female rats

To investigate the effect of perinatal testosterone exposure, which simulates the endogenous testosterone peak, on neuron loss during aging, nuclear morphology was evaluated in male and female rats as well as in female rats treated with testosterone perinatally followed by ovariectomy (TE/Ovx). Additionally, neuronal apoptosis, which occurred primarily at postnatal day 8 (PND8), was identified by in situ TUNEL staining. Neuronal density, nuclear volume, total neuronal number and pyknotic ratio were estimated after HE stain at PND8, middle age and old age. The results showed that age-related decrease in neuronal nuclear volume and total neuron number in the sexually dimorphic nucleus of the preoptic area (SDN-POA) of female rats was significantly diminished by TE/Ovx. The pyknotic ratio in the SDN-POA of female rats at PND8 was significantly higher than that of males, and neuronal death was reversed by testosterone exposure, while no significant difference of pyknotic ratios was observed among male, female and TE/Ovx female rats at both middle and old age. Moreover, the high apoptotic incidence of female rats at PND8 was significantly diminished by testosterone exposure. These results suggest that neuron loss in the SDN-POA during aging may be predominantly determined by perinatal testosterone through modulation of postnatal neuronal apoptosis.     

Photoperiod affects the morphology of a sexually dimorphic nucleus within the preoptic area of male Japanese quail

Changes in the gross and cellular morphology of the nucleus preopticus medianus (POMn) were measured in response to changes in photoperiod in adult male Japanese quail (Coturnix japonica). POMn volume and the soma size of a dorsolateral population of neurons within POMn decreased when birds were moved from long day housing conditions (16L,8D) to short day housing conditions (8L,16D), and then increased again when birds were moved back to long day conditions, presumably as a function of the changes in circulating testosterone that accompanied changing daylengths. Male Japanese quail exhibit sexual behavior only when housed under long day housing conditions that approximate the photoperiod of the spring/summer breeding season, and do not exhibit sexual behavior when housed under short day conditions characteristic of fall/winter. Because POMn is known to be critically involved in the expression of male copulatory behavior, these morphological changes in the adult brain likely represent key functional events associated with the seasonal regulation of sexual behavior in male Japanese quail.     

Effects of prenatal stress on differentiation of the sexually dimorphic nucleus of the preoptic area (SDN-POA) of the rat brain

The present study was designed to determine the effects of prenatal malnutrition or environmental stress on the development of the sexually dimorphic nucleus of the preoptic area (SDN-POA). Pregnant rats were divided into a control group and two treatment groups (immobilization-illumination-heat or environmental stress, and nutritional stress). The two forms of stress were administered during the third trimester of gestation (days 14-20). Male and female offspring were sacrificed at birth, 20, and 60 days postnatally. The cross-sectional area of the SDN-POA was identified under light microscopy and was measured. The data confirm previous studies by showing a significant sex difference in the SDN-POA between control male and female rats. Prenatally stressed males sacrificed 20 and 60 days after birth showed SDN-POA areas 50% smaller than the nuclear areas of control males. The size of the SDN-POA of female offspring, however, was not significantly altered by prenatal treatments.     

Mu-opiate receptor binding in the medial preoptic area is cyclical and sexually dimorphic

The density and distribution of mu- and kappa-opiate receptors in the medial preoptic area (MPOA) of male and female rats across the estrous cycle was examined using quantitative in vitro autoradiography of [3H]D-Ala2,MePhe4,Gly-ol5-enkephalin (DAGO), [3H]naloxone and [3H]bremazocine binding. While no difference in kappa-receptor labeling was observed across sex or estrous stage, selective mu-receptor labeling with [3H]DAGO revealed a significant variation of density and distribution in the MPOA across the estrous cycle and between sexes. A dense concentration of mu-receptors located in the central, sexually dimorphic portion of the MPOA was observed during metestrus and diestrus in females, but not during proestrus nor in males. This region appeared to be the same as that labeled similarly using [3H]naloxone. These results suggest that a regional substrate for functional activation by endogenous opioid peptides (e.g. beta-endorphin) is cyclically regulated in females, which may explain the gonadal steroid-dependent effects of MPOA beta-endorphin on lordosis and luteinizing hormone secretion in females.     

Lesions of the sexually dimorphic nucleus of the preoptic area: effects upon LH, FSH and prolactin in rats

Bilateral lesions were placed in the sexually dimorphic nucleus of the preoptic area (SDN-POA) in castrated adult male rats in an attempt to determine a physiologic role for this nucleus. These lesions significantly attenuated the increase in plasma FSH and LH due to simultaneous castration at 24 hr, and at 7 and 14 days following surgery, and significantly decreased the levels of plasma prolactin on comparison with pre-operative values and those of castrated controls. When rats were castrated and lesions placed at 14 days following castration, plasma levels of FSH, LH and prolactin were significantly decreased at 24 hr and at 7 days following surgery. Lesions which were placed lateral or caudal to the SDN-POA simulated the effects of lesions placed within the SDN-POA upon plasma LH, FSH and prolactin, with lateral lesions being most effective. However, lesions which were placed dorsal or rostral to the SDN-POA had no effect. The results of these studies suggest that the SDN-POA may be involved in the regulation of LH, FSH, and prolactin release.     

Sexual differentiation and hormonal control of the sexually dimorphic medial preoptic nucleus in the quail

We recently identified a sexually dimorphic nucleus in the preoptic region of the Japanese quail, the medial preoptic nucleus (POM), which is significantly larger in males than in females. In the present study, we investigated the hormonal control of this morphological neuroanatomical difference and the possible relationships between the sexual dimorphism in POM volume and in copulatory behavior. Treatments which are known to affect sexual behavior were thus applied to different groups of birds and the POM volume was then measured. In one experiment, male and female quails were either gonadectomized, gonadectomized and treated with testosterone or left intact. The larger size of the POM in males was confirmed and treatments significantly affected the nucleus size which was decreased by gonadectomy and restored by testosterone treatment in both sexes to a level similar to that seen in intact males. In two other experiments, eggs were injected with estradiol benzoate on day 9 of incubation and the POM volume was measured in adulthood either in intact birds or in gonadectomized birds receiving a replacement therapy with testosterone. Despite the fact that estradiol benzoate treatment completely suppressed copulatory behavior, it did not affect the volume of the POM or slightly increased it. These data thus show that the POM volume is controlled by testosterone levels in adulthood and could thus be an interesting model for the study of the effects of steroids on the brain.