Assessing gonadal hormone contributions to affective psychopathologies across humans and animal models.

Despite increasing acknowledgement of hormonal contributions to mood and anxiety disorders, the underlying mechanisms by which gonadal hormones influence psychopathology-related behaviours remain unknown. This review focuses on recent research that examines the influence of gonadal steroid hormones, including androgens, oestrogens, and progesterone, on mood and anxiety-related behaviours in human health and disease. To this aim, the literature was surveyed for studies that assess conditions with suspected underlying hormonal imbalances in otherwise healthy participants (e.g., premenstrual dysphoric disorder, postmenopausal depression) as well as conditions linked to congenital endocrine abnormalities (e.g., Turner Syndrome, Klinefelter Syndrome, polycystic ovary syndrome, congenital adrenal hyperplasia, familial male precocious puberty, androgen insensitivity syndrome). Furthermore, to better inform clinical work and to create a translational bridge, a second goal was to set human psychopathologies and animal models of these conditions side-by-side. In the second part of the review, based on consistencies revealed in the existing literature across conditions, a new model for the impact of gonadal hormones on anxious and depressed behavioural states is proposed. Finally, we conclude by proposing directions for future research, including the development of specific tasks suitable for cross-species comparisons to increase our knowledge of the role of gonadal hormones in mood and anxiety.

Acetylcholine mediates the estrogen-induced increase in NMDA receptor binding in CA1 of the hippocampus and the associated improvement in working memory.

Elevated levels of circulating estrogen in female rats result in increased spine and synapse density and parallel increases in NMDA receptor binding in area CA1 of the hippocampus. Estrogen also influences cholinergic neurochemistry in the basal forebrain and hippocampus. The objectives of the present study were to determine the role of acetylcholine in the estrogen-induced increase in NMDA receptor binding in CA1 of the hippocampus and to investigate the relationship between increased NMDA receptor binding in CA1 and performance on a task of working memory. In the current experiments, elevating endogenous levels of acetylcholine in ovariectomized rats by 3 d of continuous administration of physostigmine, an acetylcholinesterase inhibitor, increased NMDA receptor binding in CA1 as measured by quantitative autoradiography. This increase was comparable with the increase in NMDA receptor binding induced by injections of estradiol benzoate 72 and 48 hr before death. Additionally, the administration of 5,11-dihydro-8-chloro-11-[[4-[3-[(2,2-dimethyl-1-oxopentyl)ethylamino]propyl]-1-piperidinyl]acetyl]-6H-pyrido[2,3-b][1,4]benzodiazepin-6-one (BIBN 99), an M2 receptor antagonist, blocked the ability of both estrogen and physostigmine to increase NMDA receptor binding. The regimen of estradiol replacement that was demonstrated to increase NMDA receptor binding in CA1 of ovariectomized rats also improved arm-choice accuracy in a working memory task in an eight-arm radial maze. The estrogen-induced improvement in working memory performance was blocked by BIBN 99, which also blocked the increase in NMDA receptor binding. These results indicate that acetylcholine acts at M2 muscarinic receptors to mediate the estrogen-induced increase in NMDA receptor binding in CA1 of the hippocampus as well as the associated improvement in working memory.

Behavioral and anatomical correlates of chronic episodic hypoxia during sleep in the rat.

The role played by chronic episodic hypoxia (EHYP) in the neurocognitive morbidity of obstructive sleep apnea (OSA) is unknown. Sleep recordings, Morris water maze experiments, and immunohistochemistry for NMDA NR1 glutamate receptor, c-fos protein, and apoptosis [nuclear immunoreactivity for single-stranded DNA and terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling assay] were conducted in EHYP-exposed Sprague Dawley male rats. Exposures consisted of up to14 d in an environmental chamber in which O(2) concentrations were cycled between 10 and 21% every 90 sec or 30 min during 12 hr of daylight. For the remaining 12 hr, EHYP rats breathed room air, while controls spent 14 d in room air. Although EHYP induced significant disruption of sleep architecture during the initial day of exposure, sleep patterns normalized thereafter. Marked increases in apoptosis occurred in the CA1 hippocampal region (sevenfold) and cortex (Cx; eightfold) after 1-2 d of EHYP but not in CA3 and were followed by decreases toward normoxic levels by 14 d. Double labeling for NMDA NR1 and c-fos revealed marked architectural disorganization in CA1 and Cx with increases in c-fos over time. Rats exposed to EHYP displayed significantly longer escape latencies and swim path lengths to escape a hidden platform during 12 training trials given over 2 d. Differences in the performances of EHYP and control rats, although reduced, persisted after 14 d of recovery. We conclude that EHYP is associated with marked cellular changes over time within neural regions associated with cognitive functions. Furthermore, EHYP impaired performance during acquisition of a cognitive spatial task without affecting sensorimotor function. Such changes may underlie components of the learning and memory impairments found in OSA.

Effects of ovarian hormones and environment on radial maze and water maze performance of female rats.

The effects of gonadal hormones and environment on performance in an eight-arm radial maze and in the Morris water maze were determined in female rats. Long-Evans female rats were ovariectomized or sham ovariectomized at 35 days of age, and housed in complex environments or in isolation for the duration of the study. One month following surgeries, spatial working memory performance in the radial maze was assessed. Exposure to complex environmental conditions independently enhanced performance, as indicated by increased arm choice accuracy during 20 days of maze training. Additionally, gonadally intact females significantly outperformed ovariectomized females before cyclicity was disrupted by food deprivation. Following radial maze training, spatial reference memory performance was assessed in the same females utilizing the Morris water maze. Gonadally intact females housed in isolation performed significantly more poorly during 16 days of place training trials and displayed significantly shorter times in the platform quadrants and fewer target crossings during probe trials than gonadally intact and ovariectomized females housed in complex environments and ovariectomized rats housed in isolation. Consequently, acquisition and retention of the water maze was impaired by the presence of ovaries, and this impairment was counteracted by exposure to complex environments. Performance did not differ between groups on cued trials, indicating that sensorimotor and motivational functions did not differ between groups. Results of these experiments indicate that endogenous gonadal hormones can differentially affect performance on tasks of spatial working and spatial reference memory, and that environmental conditions can interact with gonadal hormones to affect behavior.

Estrogen improves working but not reference memory and prevents amnestic effects of scopolamine of a radial-arm maze.

This study investigated the effect of estrogen treatment on working memory and reference memory of female rats. In addition, the impact of estrogen on the sensitivity of these two types of memory to the cholinergic antagonist scopolamine was investigated. At 35 days of ages, rats were ovariectomized and implanted chronically with Silastic capsules containing either 25% crystalline estradiol or 100% cholesterol. Thirty days after surgery, animals were trained on an eight-arm radial maze with four arms baited to assess both working and reference memory performance. Following training, females were given scopolamine hydrobromide (0.2 mg/kg i.p.) prior to retesting on the task. Results indicated that estrogen treatment improved working memory performance during maze acquisition but did not affect reference memory performance. Scopolamine treatment impaired performance on the working memory component, but not the reference memory component, while estrogen prevented the impairment of working memory by scopolamine. Results support previous evidence that estrogen selectively enhances performance on tasks that depend on working memory.

Estrogen improves performance of reinforced T-maze alternation and prevents the amnestic effects of scopolamine administered systemically or intrahippocampally.

In a previous study, administration of high doses of estradiol benzoate (100 microgram/kg for 3 days im) to ovariectomized Long-Evans rats counteracted impairments of reinforced T-maze alternation induced by systemic administration of scopolamine, a muscarinic receptor blocker. In the current study, daily administration of lower doses of estradiol benzoate (5 microgram/kg for 3 weeks sc) increased the number of correct reinforced alternations during T-maze acquisition in ovariectomized rats compared to oil-treated controls and prevented impairments of reinforced alternation induced by injection of scopolamine hydrobromide (0.2 mg/kg ip). Furthermore, scopolamine (20 microgram) delivered bilaterally to the dorsal hippocampus reduced reinforced T-maze alternation in ovariectomized rats previously trained to complete this task while daily treatment with estradiol benzoate (5 microgram/kg sc) for 1 week prior to scopolamine infusion counteracted this impairment. In summary, physiological levels of estrogen improved performance during acquisition of reinforced T-maze alternation and prevented impairments induced by scopolamine administered systemically or intrahippocampally.

Sexual behavior of Flinders Line female rats bred for differential cholinergic sensitivities.

Flinders Lines are two strains of rats selectively bred for their divergent physiological responses to cholinergic drug challenges. Flinders Sensitive Line (FSL) rats are highly sensitive to cholinergic stimulation of various autonomic and behavioral responses compared to Flinders Resistant Line (FRL) rats. Because cholinergic innervation contributes to the regulation of female sexual behaviors in rats, a study was conducted to compare the sexual responses of FSL females to those of FRL females, as well as to those of Long-Evans (LE) females, a conventional rat strain. Ovariectomized FSL rats exhibited significantly higher incidences of lordosis and proceptive behaviors than ovariectomized FRL and LE rats over a range of estrogen doses (2, 3, 4, 5, or 20 microgram(s)/kg estradiol benzoate at 48 h before testing) administered in combination with progesterone (1 or 2 mg/kg at 4-6 h before testing). In addition, the muscarinic antagonist scopolamine inhibited lordosis behavior strongly in FRL females over a range of doses (0.25, 0.5, 1, 2, or 4 mg/kg), but failed to inhibit lordosis in FSL females. Results indicate that FSL females are highly sensitive to the behavioral effects of gonadal steroids and highly insensitive to the effects of a muscarinic antagonist. The enhanced sexual behavioral responses of FSL females could be a consequence of their well-established cholinergic hypersensitivity or a consequence of other undocumented characteristics of FSL females such as hypersensitivity to ovarian hormones. FSL females could provide a valuable model for the study of estrogen action at behavioral, cellular, and molecular levels.

Estrogen enhances performance of female rats during acquisition of a radial arm maze.

Estrogen can influence the expression of behaviors not associated directly with reproduction, including learning and memory. However, the effects of estrogen on learning and memory in mammals are complex, dependent on a variety of factors. The radial arm maze is a traditional experimental task that takes advantage of the natural foraging strategy of rats and provides an appropriate measure for studying the effects of estrogen on working memory in this species. In the experiments reported here, ovariectomized rats were implanted subcutaneously with 5-mm Silastic capsules containing 25% estradiol diluted with cholesterol. Control females received 5-mm Silastic capsules containing 100% cholesterol. Results of three separate experiments demonstrated that estradiol administered by Silastic implants for 30 days prior to eight-arm radial maze training, during the 24 days of maze training, or both significantly improved working memory performance compared to females treated with cholesterol alone, as indicated by improved arm choice accuracy over trials. The positive effect of estradiol exposure prior to training suggests that estrogen may induce neuronal changes that persist beyond the period of exposure with functional consequences for behavior.

Mate preference and avoidance in female rats following treatment with scopolamine.

The effects of the muscarinic antagonist, scopolamine, on the behavior of female rats in a mate preference test were determined. Ovariectomized rats were primed with sufficient doses of estradiol benzoate (0.25 micrograms for 3 days) and progesterone (500 micrograms) to activate full sexual receptivity. Beginning 15 min after intraperitoneal injection of saline (0.4 ml/kg) or scopolamine hydrochloride (1 mg/kg), females were tested in an arena divided into three compartments. Females treated with scopolamine clearly avoided a compartment occupied by a gonadally intact stimulus male, instead preferring a compartment occupied by a castrated male or an empty neutral compartment. Direct contact with an intact male was a critical feature of this avoidance because females treated with scopolamine did not avoid intact males that were confined within a chamber that prevented physical contact between the male and female. Results indicate that female rats treated with scopolamine avoid direct physical contact with sexually active males, possibly due to an alteration in sexual motivation and/or the aversive nature of this interaction.

Inhibition of lordosis in female hamsters and rats by 8-OH-DPAT treatment.

In the present experiments, the ability of the 5-HT1A agonist, 8-OH-DPAT, to inhibit lordosis was determined in ovariectomized hamsters and rats under various hormonal conditions. Systemic administration of 8-OH-DPAT (0.25 mg/kg) significantly inhibited lordosis duration in ovariectomized hamsters treated on 3 consecutive wk with estradiol benzoate (3 or 10 micrograms) and progesterone (500 micrograms). Similarly, systemic administration of 8-OH-DPAT (0.25 mg/kg) significantly inhibited lordosis frequency in ovariectomized rats treated on 3 consecutive wk with estradiol benzoate (0.25, 0.5, or 25 micrograms for 3 days) and progesterone (500 micrograms), although females treated with the higher doses of estrogen were slightly less inhibited on the second and third wk of testing. Results indicate that the 5-HT1A agonist, 8-OH-DPAT, effectively inhibited lordosis in female hamsters, as well as female rats, under varied hormonal conditions.

Androgenic-anabolic steroids modify beta-endorphin immunoreactivity in the rat brain.

Immunocytochemical localization of beta-endorphin in the brains of intact and castrated male rats was conducted after the administration of high levels of androgenic-anabolic steroids (AAS; 14 daily injections of sesame oil or a cocktail of 2 mg/kg testosterone cypionate, 2 mg/kg nandrolone decanoate, and 1 mg/kg boldenone undecylenate) at doses commonly self-administered by athletes who are considered 'heavy abusers'. In normal intact oil-treated males, cytoplasmic immunoreactivity was prevalent throughout the arcuate nucleus while intense fiber tract immunoreactivity was most prevalent in the bed nucleus of the stria terminalis and the paraventricular hypothalamic nucleus. Administration of AAS significantly decreased the number of neurons exhibiting cytoplasmic immunoreactivity only in the rostral region of the arcuate nucleus. AAS treatment had no effect on beta-endorphin immunoreactivity in the middle or caudal aspects of the arcuate nucleus.

Effects of hormonal treatment and history on scopolamine inhibition of lordosis.

The muscarinic receptor blocker, scopolamine, inhibits the display of lordosis behavior in female rats but its effectiveness depends on hormonal conditions. In these experiments, systemic administration of scopolamine (0.031-4 mg/kg) inhibited lordosis in ovariectomized rats brought into receptivity by treatment with a low dose of estradiol benzoate (EB, 0.25 micrograms for 3 days) with progesterone (P, 500 micrograms for 1 day), or a high dose of EB (25 micrograms for 3 days) with and without P. However, the effectiveness of scopolamine was reduced at the high dose of EB and with the addition of P. Furthermore, scopolamine failed to inhibit lordosis in females treated on a second week with the high dose of EB with or without P, unless an interval of at least 3 weeks separated the two sets of steroid treatments. The reduced effectiveness of scopolamine cannot be explained by peripheral mechanisms because its inhibitory effect on lordosis also was reduced following repeated hormonal exposure even when scopolamine was infused directly into the lateral ventricles.

Estrogen and estrogen-progesterone treatments counteract the effect of scopolamine on reinforced T-maze alternation in female rats.

The purpose of the experiments was to determine if steroid hormone treatments would attenuate the effect of the muscarinic receptor blocker scopolamine on a memory task. Ovariectomized rats were trained first to alternate for food reward between the arms of a T maze. Following training, females treated with scopolamine hydrobromide (0.2 mg/kg ip) did not alternate correctly between the arms of the T maze and responded at chance levels. However, when estradiol benzoate (25 micrograms) was administered 72, 48, and 24 hr before testing alone or in combination with progesterone (500 micrograms) administered 4-6 hr before testing, females alternated successfully between the arms of the T maze following scopolamine administration. Results indicate that gonadal steroids can completely counteract the impairment of T maze performance induced by scopolamine in female rats.

Estrogen dependence of cholinergic systems that regulate lordosis in cycling female rats.

Previous evidence indicated that physostigmine, an acetylcholinesterase inhibitor, facilitated lordosis behavior when administered intraventricularly to cycling female rats on proestrus prior to the onset of natural sexual receptivity, but not when administered to rats on mid-diestrus or diestrus II. In the present experiments, intraventricular infusion of physostigmine (10 micrograms bilaterally) facilitated lordosis on mid-diestrus and diestrus II if females were primed with two injections of estradiol (0.2, 0.1, or 0.05 micrograms) administered 20 h and 32 h prior to infusion of physostigmine. Despite unequal levels of endogenous progesterone, physostigmine facilitated lordosis equally on mid-diestrus and diestrus II following estradiol priming. Finally, intraventricular infusion of the muscarinic receptor blocker scopolamine (20 micrograms bilaterally) reduced the incidence of lordosis in females that displayed lordosis on mid-diestrus following estrogen priming. Results confirm that cholinergic mechanisms influence sexual behavior displayed by cycling female rats. Data further indicate that sufficient estrogen stimulation is necessary for cholinergic neurons to facilitate lordosis. However, progesterone does not play a major role in the regulation of lordosis by cholinergic systems.

Effects of a muscarinic antagonist on various components of female sexual behavior in the rat.

The effects of the muscarinic antagonist scopolamine on lordosis, solicitation, pacing, approach, attractivity, and activity were evaluated in ovariectomized rats brought into sexual receptivity with estrogen and progesterone. Systemic (1 mg/rat) or intraventricular (10 micrograms bilaterally) administration of scopolamine significantly reduced the incidence of lordosis and solicitation behaviors and disrupted typical pacing of sexual contacts with a stimulus male. In addition, females avoided contact with a stimulus male, but not a stimulus female, following intraventricular infusion of scopolamine. The levels of general activity and frequencies of sexual contacts were similar in females treated intraventricularly with scopolamine and vehicle solutions. Consequently, scopolamine disrupted various components of sexual behavior, including lordosis, solicitation, pacing, and approach, without altering female attractivity or general activity.

The effect of estrogen treatment on scopolamine inhibition of lordosis.

Previous evidence indicates that the cholinergic muscarinic antagonist, scopolamine, inhibits lordosis in female rats. In the experiments reported here, the effects of various doses and repeated administrations of estrogen on the scopolamine inhibition of lordosis were examined. In the first experiment, intraperitoneal injections of scopolamine (1 mg/rat) completely inhibited lordosis in ovariectomized rats primed with low doses of estradiol benzoate (0.25 or 0.5 micrograms for 3 days) and progesterone (500 micrograms). However, scopolamine was significantly less effective in inhibiting lordosis in females primed with a higher dose of estradiol benzoate (25 micrograms for 3 days) and progesterone (500 micrograms). When hormone priming was repeated on subsequent weeks, scopolamine continued to inhibit lordosis in females that received 0.25 micrograms estradiol benzoate but was less effective in females primed with 0.5 micrograms. Scopolamine failed to inhibit lordosis in females treated with 25 micrograms estradiol benzoate on these later tests. In the second experiment, various doses of scopolamine (1, 2, or 4 mg/rat) were administered intraperitoneally to females primed with the highest dose of estradiol benzoate (25 micrograms) and progesterone (500 micrograms). Lordosis was inhibited equally by all scopolamine doses during the first week. As in the first experiment, scopolamine failed to inhibit lordosis at all doses on subsequent weeks of testing. These results indicate that the ability of scopolamine to inhibit lordosis is reduced by increasing the dose or the number of estrogen exposures. Because higher doses of scopolamine failed to restore its inhibitory effect on lordosis an upregulation of muscarinic receptors by estrogen cannot account for the reduced effectiveness of scopolamine.

Muscarinic receptor subtypes and sexual behavior in female rats.

Cholinergic muscarinic systems are involved in the regulation of female sexual behavior in rats and hamsters. This series of experiments was designed to determine whether sexual behavior in female rats is controlled preferentially by one of the traditional muscarinic receptor subtypes. Intraventricular infusion of the muscarinic antagonist scopolamine (10 micrograms bilaterally) which binds with high affinity to both M1 and M2 subtypes inhibited sexual behavior, as indicated by the incidence of lordosis, in ovariectomized rats treated with estrogen and progesterone. In contrast, the M1-selective antagonist pirenzepine failed to reduce the incidence of lordosis following intraventricular infusion (10 to 80 micrograms bilaterally). Biochemical analyses revealed that intraventricular infusion of scopolamine (10 micrograms bilaterally) inhibited both M1 and M2 binding in brain tissues while intraventricular infusion of pirenzepine (10 micrograms bilaterally) completely inhibited M1 binding without affecting M2 binding. Intraventricular infusions of the acetylcholinesterase inhibitor physostigmine (10 micrograms bilaterally), the cholinergic agonist carbachol (1 microgram bilaterally), and the muscarinic agonist oxotremorine-M (0.1 micrograms bilaterally) activated lordosis in ovariectomized females primed with low doses of estrogen. In contrast, the putative M1 agonist McN-A-343 failed to significantly increase lordosis following intraventricular infusions (1, 10, 20 micrograms bilaterally). According to biochemical results, the ability of these agents to activate lordosis in female rats was related to their affinities for M2 binding sites not M1 binding sites. In a final experiment, estrogen treatment of ovariectomized rats did not alter muscarinic subtype binding in several brain areas as measured by the M1-selective ligand [3H] pirenzepine and the M2-selective ligand [3H] oxotremorine-M. The results of these experiments confirm that muscarinic systems contribute to the regulation of lordosis in female rats and indicate that M2 binding sites rather than M1 binding sites may be a critical component of this regulation.

Physostigmine facilitation of lordosis in naturally cycling female rats.

Both systemic and intracerebral administrations of the cholinergic muscarinic antagonist, scopolamine, have been shown to inhibit naturally occurring sexual behavior in intact, cycling female rats. The present study examined the facilitative effects of the acetylcholinesterase inhibitor, physostigmine (eserine), on sexual behavior in intact, cycling female rats. Cycling was determined by daily monitoring of sexual behavior and vaginal cytology. When administered during either early proestrus or proestrus, physostigmine activated lordosis 15 min and 1 hr after intraventricular infusion (10 micrograms bilaterally). However, infusion of physostigmine failed to facilitate lordosis 15 min after administration during either diestrus I, mid-diestrus, or diestrus II. The administration of this cholinergic agent did not interrupt cyclicity patterns. Because estrogen levels are highest during proestrus and cholinergic facilitation appears to be limited to this time, it is suggested that estrogen priming of central cholinergic systems is necessary for the cholinergic regulation of sexual behavior in intact, cycling female rats.

Effects of uric acid and caffeine on A1 adenosine receptor binding in developing rat brain.

Previous studies have demonstrated that elevated levels of serum uric acid or caffeine are associated with increased locomotor activity in rats and humans. Since uric acid and caffeine are structurally similar, it was hypothesized that these compounds alter locomotor activity through a common neural mechanism, specifically by acting as receptor antagonists at adenosine A1 binding sites. In vitro competition of caffeine and uric acid against the A1 agonist, [3H] cyclohexyladenosine ([3H]CHA), was conducted using homogenates of adult rat forebrain. Caffeine effectively competed for the A1 binding site as previously reported (IC50 = 107 microM), but uric acid failed to compete with [3H]CHA binding at concentrations within a relevant physiological range. Nevertheless, in vivo experiments indicated that chronic elevation of uric acid following allantoxanamide treatment of male rats on days 4-27 of life significantly decreased A1 receptor binding in the striatum, a region traditionally implicated in mammalian locomotion. In contrast, chronic caffeine treatment on days 4-27 of life caused an increase in A1 receptor binding in the cortex similar to increases reported previously in whole brain. These changes in A1 receptor binding following chronic elevation of uric acid or caffeine did not persist in rats that had been withdrawn from allantoxanamide or caffeine treatment for 14 days. Results from in vitro and in vivo experiments indicate that despite a similar molecular structure uric acid does not act by the same mechanism as caffeine to increase locomotor activity in rats.

Cholinergic regulation of sexual behavior in female hamsters.

The effect of cholinergic manipulations on sexual behavior in female hamsters was determined in a series of experiments. The cholinergic receptor antagonist, scopolamine, reduced total lordosis duration following systemic (1 mg/kg) or intraventricular (10 and 20 micrograms bilaterally) administration to ovariectomized hamsters primed with estrogen and progesterone. The inhibitory effect of scopolamine on lordosis occurred within 15 min after either treatment route and persisted at 2 hr after systemic administration. Intraventricular administration of the acetylcholinesterase inhibitor, physostigmine (10 micrograms bilaterally), activated lordosis of short duration in ovariectomized hamsters primed only with estrogen. These results indicate that the cholinergic system plays a facilitative role in the regulation of sexual behavior in female hamsters similar to that demonstrated previously in female rats. The activational effect of cholinergic neurotransmission on female sexual behavior may be a neural mechanism common to a number of mammalian species.