Vasopressin infusion into the lateral septum of adult male rats rescues progesterone-induced impairment in social recognition.

It is well established that social recognition memory is mediated, in part, by arginine vasopressin (AVP). AVP cells within the bed nucleus of the stria terminalis (BST) and medial amygdala (MeA) send AVP-ergic projections to the lateral septum (LS). We have demonstrated that progesterone treatment decreases AVP immunoreactivity within the BST, the MeA and the LS, and that progesterone treatment impairs social recognition. These data suggested that progesterone may impair social recognition memory by decreasing AVP. In the present experiment, we hypothesized that infusions of AVP into the LS would rescue the progesterone-induced impairment in social recognition within adult male rats. One week after adult male rats underwent cannula surgery, they were given systemic injections of either a physiological dose of progesterone or oil control for 3 days. Four hours after the last injection, we tested social recognition memory using the social discrimination paradigm, a two-trial test that is based on the natural propensity for rats to be highly motivated to investigate novel conspecifics. Immediately after the first exposure to a juvenile, each animal received bilateral infusions of either AVP or artificial cerebrospinal fluid into the LS. Our results show that, as expected, control animals exhibited normal social discrimination. In corroboration with our previous results, animals given progesterone have impaired social discrimination. Interestingly, animals treated with progesterone and AVP exhibited normal social discrimination, suggesting that AVP treatment rescued the impairment in social recognition caused by progesterone. These data also further support a role for progesterone in modulating vasopressin-dependent behavior within the male brain.

Regulation of progestin receptor expression in the developing rat brain by a dopamine d1 receptor antagonist.

Steroid receptors within the developing brain influence a variety of cellular processes that endure into adulthood, altering both behaviour and physiology. Therefore, it is important to understand how steroid receptor expression is regulated during early brain development. Most studies indicate that oestradiol, by acting upon oestrogen receptors, increases the expression of progestin receptors in the developing brain. We have recently observed an additional mechanism by which dopamine can increase the expression of progestin receptors in developing female rat brain. That is, we found that a dopamine D1 receptor agonist can further increase progestin receptor expression by activating oestrogen receptors in a ligand-independent manner within restricted areas of female brain; however, it is unclear whether dopamine D1 receptors are involved in the normally occurring expression of progestin receptors in developing male and female brain. To investigate this, we examined whether a dopamine D1 receptor antagonist can disrupt the normal developmental expression of progestin receptors in both male and female rat brain. We report that treatment with a dopamine D1 receptor antagonist reduces progestin receptor expression within some, but not all, regions of the developing rat brain in a sex-specific manner. Some of the current findings also suggest that dopamine might be acting to prevent sex differences in progestin receptor expression in some areas while contributing to a sex difference in other areas.

Progesterone treatment of adult male rats suppresses arginine vasopressin expression in the bed nucleus of the stria terminalis and the centromedial amygdala.

The steroid sensitive vasopressin cells of the bed nucleus of the stria terminalis (BST) and centromedial amygdala (CMA) are involved in numerous behavioural and physiological functions. These cells are known to be greatly influenced by gonadal steroids. Castration reduces and testosterone replacement restores arginine vasopressin (AVP)-immunoreactive (-ir) labelling and AVP mRNA expression in the BST and CMA. Gonadal steroids appear to act directly in AVP-expressing cells within the BST and CMA, because the majority of AVP-ir cells in these areas contain oestrogen and androgen receptor immunoreactivity. Recently, we have localised progestin receptor immunoreactivity in virtually all of the AVP-ir cells in the BST and CMA. To understand the role played by progestin receptors in AVP cells within the BST and CMA, we treated male rats with 1 mg of progesterone or oil for 5 days, and then examined AVP immunoreactivity within the brain. We found that progesterone decreased AVP-ir labelling within the BST and CMA, as well as in two of the projection sites of these cells, the lateral septum and lateral habenula. Progesterone treatment did not alter testosterone secretion from the testes, nor did it alter adult male sexual behaviour. These data illustrate an additional mechanism by which the AVP cells in the BST and CMA can be regulated. These data also suggest that progesterone may act in the male brain to influence behaviours that are AVP-dependent.

Sex differences in mouse cortical thickness are independent of the complement of sex chromosomes.

Although the morphology of the cerebral cortex is known to be sexually dimorphic in several species, to date this difference has not been investigated in mice. The present study is the first to report that the mouse cerebral cortex is thicker in males than in females. We further asked if this sex difference is the result of gonadal hormones, or alternatively is induced by a direct effect of genes encoded on the sex chromosomes. The traditional view of mammalian neural sexual differentiation is that androgens or their metabolites act during early development to masculinize the brain, whereas a feminine brain develops in the relative absence of sex steroids. We used mice in which the testis determination gene Sry was inherited independently from the rest of the Y chromosome to produce XX animals that possessed either ovaries or testes, and XY animals that possessed either testes or ovaries. Thus, the design allowed assessment of the role of sex chromosome genes, independent of gonadal hormones, in the ontogeny of sex differences in the mouse cerebral cortex. When a sex difference was present, mice possessing testes were invariably masculine in the morphology of the cerebral cortex, independent of the complement of their sex chromosomes (XX vs. XY), and mice with ovaries always displayed the feminine phenotype. These data suggest that sex differences in cortical thickness are under the control of gonadal steroids and not sex chromosomal complement. However, it is unclear whether it is the presence of testicular secretions or the absence of ovarian hormones that is responsible for the thicker male cerebral cortex.

Expression of cAMP response element binding protein-binding protein in the song control system and hypothalamus of adult European starlings (Sturnus vulgaris).

In songbirds, the initiation of song behaviour and the neural substrate of this system are highly influenced by gonadal steroids. Receptors for gonadal steroid hormones, such as androgens and oestrogens, have been localized within select nuclei of the song system. An important step in steroid receptor action is the recruitment of nuclear receptor coactivators. The coactivator, cAMP response element binding protein (CREB)-binding protein (CBP), has been implicated in both androgen and oestrogen receptor transactivation. Although the role of CBP in transcriptional mechanisms has been widely studied, little is known about CBP expression in the brain. The association between the distribution of CBP and oestrogen receptors in the hippocampus has been related to long-term memory. However, the distribution of brain CBP has not been related to the expression of gonadal steroid receptors in a system as relevant to reproductive behaviour as the avian song system. Western immunoblotting of European starling (Sturnus vulgaris) brain tissue reveals a band at 265 kDa. Immunohistochemical localization of CBP in starling brain indicates wide, but heterogeneous expression. CBP-immunoreactive (CBP-ir) cells define the boundaries of song control nuclei. In HVc (sometimes called the High Vocal Center) and the robust nucleus of the archistriatum (RA), there is a higher density of CBP-ir cells within the boundaries of these nuclei than in adjacent neostriatum or archistriatum, for HVc and RA, respectively. We also report that the distribution of CBP-ir cells varies among different nuclei within the song control system. CBP-ir cells within area X (also a part of the song system) and HVc are densely packed into clusters, whereas cells can be easily discriminated in RA. CBP is also highly expressed in hypothalamic areas, indicating that areas rich in steroid receptors also contain CBP. These data suggest that CBP is important for modulating transcriptional activities in the song system and other sites in the songbird brain that express gonadal steroid receptors.

Progestin receptor immunoreactivity within steroid-responsive vasopressin-immunoreactive cells in the male and female rat brain.

Progestin receptor immunoreactivity is found in the same regions of the bed nucleus of stria terminalis (BST) and centromedial amygdala (CMA) as steroid-responsive vasopressin immunoreactive (AVP-ir) cells. To test whether AVP-ir cells express progestin receptors, brains of male rats were stained immunocytochemically for arginine vasopressin as well as progestin receptors. In BST and CMA, over 95% of AVP-ir cells contained progestin receptor immunoreactivity. In contrast, none of the AVP-ir cells in the suprachiasmatic, supraoptic and paraventricular nuclei expressed progestin receptor immunoreactivity. To study whether progestin receptor expression in AVP-ir cells in the BST and CMA is responsive to gonadal steroids, male and female rats were castrated and implanted with either empty capsules or capsules filled with testosterone or oestradiol, respectively. Ten days later, brains were processed for AVP and progestin receptor immunoreactivity. Although there was no effect of hormonal status on the percentage of colocalized cells, the level of progestin receptor immunoreactivity was higher in rats that received gonadal steroids than those that did not. The presence of progestin receptor immunoreactivity in steroid responsive AVP-ir cells, and the responsiveness of this expression to gonadal hormones, is consistent with the possibility that the effects of gonadal steroids on AVP-ir expression in the BST and CMA may be mediated at least in part by progestin receptors.

Expression of the nuclear receptor coactivator, cAMP response element-binding protein, is sexually dimorphic and modulates sexual differentiation of neonatal rat brain.

Recent studies indicate that the transcriptional activity of steroid receptors is governed by proteins called nuclear receptor coactivators. Using immunocytochemistry, we found that on the day of birth (postnatal d 0) males express higher levels of the nuclear receptor coactivator, cAMP response element binding protein-binding protein (CBP), within the ventromedial hypothalamus, medial preoptic area, and arcuate nucleus. Using Western immunoblots, we confirmed that males have higher levels of CBP on postnatal d 0, 1, and 5; however, there was no sex difference on postnatal d 11. To examine the functional role of CBP, we infused oligodeoxynucleotides that were antisense to CBP mRNA or a scrambled sequence as a control into the hypothalamus of female rats on postnatal d 0, 1, and 2. On postnatal d 1, all rats were injected with 100 microg testosterone propionate to both masculinize (increase male) and defeminize (decrease female) sexual behavior. Rats were ovariectomized in adulthood and tested for adult sexual behavior. Neonatal CBP antisense oligodeoxynucleotides treatment interfered with the defeminizing, but not the masculinizing, actions of testosterone. These results indicate that CBP expression in developing rat brain is sexually dimorphic and an important modulator for steroid hormone action.

Androstenedione effects on the vasopressin innervation of the rat brain.

The steroid hormone androstenedione profoundly influences the development and expression of sexual and aggressive behavior. The neural basis of these effects are, however, poorly understood. In this study we evaluated androstenedione's ability to maintain vasopressin peptide levels in the gonadal steroid-responsive vasopressin cells of the bed nucleus of the stria terminalis and the centromedial amygdala, and their projections. Adult male rats were castrated and given testosterone, androstenedione or no hormonal treatment for five weeks. Their brains were then processed for vasopressin immunoreactivity. Androstenedione and testosterone treatment were equally effective in preventing the reduction of vasopressin immunoreactivity associated with castration. Androstenedione may therefore be able to mimic the effects of testosterone on testosterone-responsive neural systems.

Down-regulation of estrogen receptor immunoreactivity by 17 beta-estradiol in the guinea pig forebrain.

Low amplitude pulses of estradiol-17 beta (E2-17 beta) are more effective than large single bolus injections or constant exposure to E2-17 beta in inducing progesterone-facilitated sex behavior in female rats and guinea pigs. The present study examined whether the increased responsiveness to E2-17 beta is due to an increase in the number of estrogen receptors in the estrogen receptor rich areas of the hypothalamus and amygdala. Initial studies examined the rapid effects (20 min) of a high dose of E2-17 beta (50 micrograms) on estrogen receptor immunostaining using either the H222 antibody or the ER 21 antiserum. ER 21 immunostaining was not affected by the E2-17 beta treatment suggesting that it binds to both occupied and unoccupied estrogen receptors. Therefore the ER 21 antiserum was used to characterize the regulation of estrogen receptor immunoreactivity (ER-IR) by E2-17 beta. ER-IR was examined for 48 h and serum E2-17 beta for 24 h following a 2 micrograms s.c. injection of E2-17 beta (a dose similar to that used in multiple pulse paradigms). Serum E2-17 beta peaked 15 to 30 min following the injection and returned to baseline values by 1 h. In all but one area maximal suppression of ER-IR occurred at 12 h. In summary, 1) decreases in estrogen receptor immunoreactivity following E2-17 beta are consistent with studies in which estrogen receptors were assayed by binding assays and estrogen receptor mRNA was determined by in situ hybridization; 2) the ER 21 antiserum is able to detect both occupied and unoccupied estrogen receptors and 3) H222 immunoreactivity is influenced by the presence of E2-17 beta, so that the level of H222-IR is a reflection of ligand/receptor binding dynamics. The data suggest that up-regulation of estrogen receptors does not account for the increase in behavioral sensitivity which is observed following multiple pulses of E2-17 beta.