Neural androgen receptors affect the number of surviving new neurones in the adult dentate gyrus of male mice.

Adult hippocampal neurogenesis occurs in many mammalian species. In rats, the survival of new neurones within the hippocampus is modulated by the action of androgen via the androgen receptor (AR); however, it is not known whether this holds true in mice. Furthermore, the evidence is mixed regarding whether androgens act in neural tissue or via peripheral non-neural targets to promote new neurone survival in the hippocampus. We evaluated whether the action of androgen via AR underlies the survival of new neurones in mice, and investigated whether increasing AR selectively in neural tissue would increase new neurone survival in the hippocampus. We used the cre-loxP system to overexpress AR only in neural tissues (Nestin-AR). These males were compared with wild-type males, as well as control males with 1 of the 2 mutations required for overexpression. Mice were gonadectomised and injected with the DNA synthesis marker, bromodeoxyuridine (BrdU) and for 37 days (following BrdU injection), mice were treated with oil or dihydrotestosterone (DHT). Using immunohistochemistry, proliferation (Ki67) and survival (BrdU) of new neurones were both evaluated in the dorsal and ventral dentate gyrus. Dihydrotestosterone treatment increased the survival of new neurones in the entire hippocampus in wild-type mice and control mice that only have 1 of 2 necessary mutations for transgenic expression. However, DHT treatment did not increase the survival of new neurones in mice that overexpressed AR in neural tissue. Cell proliferation (Ki67) and cell death (pyknotic cells) were not affected by DHT treatment in wild-type or transgenic males. These results suggest that androgens act via neural AR to affect hippocampal neurogenesis by promoting cell survival; however, the relationship between androgen dose and new neurone survival is nonlinear.

Neuronal Gonadotrophin-Releasing Hormone (GnRH) and Astrocytic Gonadotrophin Inhibitory Hormone (GnIH) Immunoreactivity in the Adult Rat Hippocampus.

Gonadotrophin-releasing hormone (GnRH) and gonadotrophin inhibitory hormone (GnIH) are neuropeptides secreted by the hypothalamus that regulate reproduction. GnRH receptors are not only present in the anterior pituitary, but also are abundantly expressed in the hippocampus of rats, suggesting that GnRH regulates hippocampal function. GnIH inhibits pituitary gonadotrophin secretion and is also expressed in the hippocampus of a songbird; its role outside of the reproductive axis is not well established. In the present study, we employed immunohistochemistry to examine three forms of GnRH [mammalian GnRH-I (mGnRH-I), chicken GnRH-II (cGnRH-II) and lamprey GnRH-III (lGnRH-III)] and GnIH in the adult rat hippocampus. No mGnRH-I and cGnRH-II+ cell bodies were present in the hippocampus. Sparse mGnRH-I and cGnRH-II+ fibres were present within the CA1 and CA3 fields of the hippocampus, along the hippocampal fissure, and within the hilus of the dentate gyrus. No lGnRH-III was present in the rodent hippocampus. GnIH-immunoreactivity was present in the hippocampus in cell bodies that resembled astrocytes. Males had more GnIH+ cells in the hilus of the dentate gyrus than females. To confirm the GnIH+ cell body phenotype, we performed double-label immunofluorescence against GnIH, glial fibrillary acidic protein (GFAP) and NeuN. Immunofluorescence revealed that all GnIH+ cell bodies in the hippocampus also contained GFAP, a marker of astrocytes. Taken together, these data suggest that GnRH does not reach GnRH receptors in the rat hippocampus primarily via synaptic release. By contrast, GnIH might be synthesised locally in the rat hippocampus by astrocytes. These data shed light on the sites of action and possible functions of GnRH and GnIH outside of the hypothalamic-pituitary-gonadal axis.

Effects of chronic oestradiol, progesterone and medroxyprogesterone acetate on hippocampal neurogenesis and adrenal mass in adult female rats.

Both natural oestrogens and progesterone influence synaptic plasticity and neurogenesis within the female hippocampus. However, less is known of the impact of synthetic hormones on hippocampal structure and function. There is some evidence that the administration of the synthetic progestin, medroxyprogesterone acetate (MPA) is not as beneficial as natural progesterone and can attenuate oestrogen-induced neuroprotection. Although the effects of oestradiol have been well studied, little is known about the effects of natural and synthetic progestins alone and in combination with oestradiol on adult neurogenesis in females. In the present study, we investigated the effects of chronic oestradiol, progesterone, MPA and the co-administration of each progestin with oestradiol on neurogenesis within the dentate gyrus of adult ovariectomised female rats. Twenty-four hours after a bromodeoxyuridine (BrdU; 200 mg/kg) injection, female rats were repeatedly administered either progesterone (1 or 4 mg), MPA (1 or 4 mg), oestradiol benzoate (EB), progesterone or MPA in combination with EB (10 µg), or vehicle for 21 days. Rats were perfused on day 22 and brain tissue was analysed for the number of BrdU-labelled and Ki67 (an endogenous marker of cell proliferation)-expressing cells. EB alone and MPA + EB significantly decreased neurogenesis and the number of surviving BrdU-labelled cells in the dorsal region of the dentate gyrus, independent of any effects on cell proliferation. Furthermore, MPA (1 and 4 mg) and MPA + EB treated animals had significantly lower adrenal/body mass ratios and reduced serum corticosterone (CORT) levels. By contrast, progesterone + EB treated animals had significantly higher adrenal/body mass ratios and 1 mg of progesterone, progesterone + EB, and EB significantly increased CORT levels. The results of the present study demonstrate that different progestins alone and in combination with oestradiol can differentially affect neurogenesis (via cell survival) and regulation of the hypothalamic-pituitary-adrenal axis. These findings have implications for women using hormone replacement therapies with MPA for both neuroprotection and stress-related disorders.

Sex, hormones and neurogenesis in the hippocampus: hormonal modulation of neurogenesis and potential functional implications.

The hippocampus is an area of the brain that undergoes dramatic plasticity in response to experience and hormone exposure. The hippocampus retains the ability to produce new neurones in most mammalian species and is a structure that is targeted in a number of neurodegenerative and neuropsychiatric diseases, many of which are influenced by both sex and sex hormone exposure. Intriguingly, gonadal and adrenal hormones affect the structure and function of the hippocampus differently in males and females. Adult neurogenesis in the hippocampus is regulated by both gonadal and adrenal hormones in a sex- and experience-dependent way. Sex differences in the effects of steroid hormones to modulate hippocampal plasticity should not be completely unexpected because the physiology of males and females is different, with the most notable difference being that females gestate and nurse the offspring. Furthermore, reproductive experience (i.e. pregnancy and mothering) results in permanent changes to the maternal brain, including the hippocampus. This review outlines the ability of gonadal and stress hormones to modulate multiple aspects of neurogenesis (cell proliferation and cell survival) in both male and female rodents. The function of adult neurogenesis in the hippocampus is linked to spatial memory and depression, and the present review provides early evidence of the functional links between the hormonal modulation of neurogenesis that may contribute to the regulation of cognition and stress.

Androgens increase survival of adult-born neurons in the dentate gyrus by an androgen receptor-dependent mechanism in male rats.

Gonadal steroids are potent regulators of adult neurogenesis. We previously reported that androgens, such as testosterone (T) and dihydrotestosterone (DHT), but not estradiol, increased the survival of new neurons in the dentate gyrus of the male rat. These results suggest androgens regulate hippocampal neurogenesis via the androgen receptor (AR). To test this supposition, we examined the role of ARs in hippocampal neurogenesis using 2 different approaches. In experiment 1, we examined neurogenesis in male rats insensitive to androgens due to a naturally occurring mutation in the gene encoding the AR (termed testicular feminization mutation) compared with wild-type males. In experiment 2, we injected the AR antagonist, flutamide, into castrated male rats and compared neurogenesis levels in the dentate gyrus of DHT and oil-treated controls. In experiment 1, chronic T increased hippocampal neurogenesis in wild-type males but not in androgen-insensitive testicular feminization mutation males. In experiment 2, DHT increased hippocampal neurogenesis via cell survival, an effect that was blocked by concurrent treatment with flutamide. DHT, however, did not affect cell proliferation. Interestingly, cells expressing doublecortin, a marker of immature neurons, did not colabel with ARs in the dentate gyrus, but ARs were robustly expressed in other regions of the hippocampus. Together these studies provide complementary evidence that androgens regulate adult neurogenesis in the hippocampus via the AR but at a site other than the dentate gyrus. Understanding where in the brain androgens act to increase the survival of new neurons in the adult brain may have implications for neurodegenerative disorders.

Distribution of androgen receptor immunoreactivity in the brainstem of male rats.

Gonadal steroids such as testosterone and estrogen are necessary for the normal activation of male rat sexual behavior. The medial preoptic area (MPOA), an important neural substrate regulating mating, accumulates steroids and also expresses functional androgen receptors (AR). The MPOA is intimately connected with other regions implicated in copulation, such as the bed nucleus of the stria terminalis and medial amygdala. Inputs to the MPOA arise from several areas within the brainstem, synapsing preferentially onto steroid sensitive MPOA cells which are activated during sexual activity. Given that little is known about the distribution of AR protein in the brainstem of male rats, we mapped the distribution of AR expressing cells in the pons and medulla using immunocytochemistry. In agreement with previous reports, AR immunoreactivity (AR-ir) was detected in ventral spinal motoneurons and interneurons. In addition, AR-ir was detected in areas corresponding to the solitary tract, lateral paragigantocellular and alpha and ventral divisions of the gigantocellular reticular nuclei, area postrema, raphe pallidus, ambiguus nucleus, and intermediate reticular nucleus. Several regions within the pons contained AR-ir, such as the tegmental and central gray, parabrachial nucleus, locus coeruleus, Barrington's nucleus, periaqueductal gray, and dorsal raphe. In contrast with in situ hybridization studies, auditory and somatosensory areas were AR-ir negative, and, except for very light staining in the prepositus nucleus, areas carrying vestibular information did not display AR-ir. Additionally, cranial nerve motoneurons of the hypoglossal, facial, dorsal vagus, and spinal trigeminal did not display AR-ir in contrast to previous reports. The data presented here indicate that androgens may influence numerous cell groups within the brainstem. Some of these probably constitute a steroid sensitive circuit linking the MPOA to motoneurons in the spinal cord via androgen responsive cells in the caudal ventral medulla.