The effect of rapid and depot testosterone and estradiol on spatial performance in water maze

Men and women differ in some cognitive functions including spatial abilities. These differences seem to be affected by sex steroids, but the results are controversial. The aim of this work is to describe the effects of rapid or depot testosterone and estradiol on spatial memory in rats. Thirty-two adult male Wistar rats were divided into 6 groups. Five groups were gonadectomized, and one group was left as control. Castrated groups received sterile oil, testosterone isobutyras, testosterone propionate, estradiol dipropionate or estradiol benzoate. We evaluated spatial performance (escape latency, overall improvement, and time in the quadrant after platform removal) of the rats in a spatial water maze. Animals receiving exogenous sex steroids showed higher plasma concentrations of the particular hormones. Experimental groups improved during the acquisition spatial trials in the water maze. No significant differences between the groups during probe trial were found. In overall improvement, the testosterone depot and estradiol depot groups showed less improvement in comparison to the control groups (P<0.05). No differences in respect to administered hormones were found in corresponding receptor gene expression in hippocampus. In conclusion, exogenous testosterone affects spatial memory of adult castrated males.     

Progesterone protects normative anxiety-like responding among ovariectomized female mice that conditionally express the HIV-1 regulatory protein,Tat, in the CNS

Increased anxiety is co-morbid with human immunodeficiency virus (HIV) infection. Actions of the neurotoxic HIV-1 regulatory protein, Tat, may contribute to affective dysfunction. We hypothesized that Tat expression would increase anxiety-like behavior of female GT-tg bigenic mice that express HIV-1 Tat protein in the brain in a doxycycline-dependent manner. Furthermore, given reports that HIV-induced anxiety may occur at lower rates among women, and that the neurotoxic effects of Tat are ameliorated by sex steroids in vitro, we hypothesized that 17β-estradiol and/or progesterone would ameliorate Tat-induced anxiety-like effects. Among naturally-cycling proestrous and diestrous mice, Tat-induction via 7 days of doxycycline treatment significantly increased anxiety-like responding in an open field, elevated plus maze and a marble-burying task, compared to treatment with saline. Proestrous mice demonstrated less anxiety-like behavior than diestrous mice in the open field and elevated plus maze, but these effects did not significantly interact with Tat-induction. Among ovariectomized mice, doxycycline-induced Tat protein significantly increased anxiety-like behavior in an elevated plus maze and a marble burying task compared to saline-treated mice, but not an open field (where anxiety-like responding was already maximal). Co-administration of progesterone (4 mg/kg), but not 17β-estradiol (0.09 mg/kg), with doxycycline significantly ameliorated anxiety-like responding in the elevated plus maze and marble burying tasks. When administered together, 17β-estradiol partially antagonized the protective effects of progesterone on Tat-induced anxiety-like behavior. These findings support evidence of steroid-protection over HIV-1 proteins, and extend them by demonstrating the protective capacity of progesterone on Tat-induced anxiety-like behavior of ovariectomized female mice.     

Sex differences in odor-stimulated flank marking in the golden hamster (Mesocricetus auratus).

To determine whether sex differences exist in the frequency of odor-stimulated flank marking, intact male and female hamsters were exposed to the recently vacated home cages of male stimulus hamsters for a 10-min test on 4 consecutive days. Females were found to mark at significantly higher levels than males. To investigate the role of gonadal hormones in the sex differences in flank marking, gonadectomized male and female hamsters were implanted with Silastic capsules containing estradiol or testosterone. Females exhibited twofold higher levels of odor-stimulated flank marking than males, and the amount of flank marking was significantly higher when the hamsters were administered testosterone than when they were administered estradiol. These data demonstrate that sex differences exist in the frequency of flank marking stimulated by the odors of male hamsters, and that these sex differences do not appear to result from the typical sex-specific patterns of circulating levels of estradiol and testosterone.     

Organizational effects of estrogen on male-type vulnerability to early weaning

We previously reported that early-weaned (postnatal day 14) male ICR mice, compared to normally weaned animals, exhibited a persistent increase in anxiety-related behavior in the elevated plus maze test. In this study, we examined whether steroid hormone manipulations on postnatal day 0 and at the ages of 2 or 3 weeks affected male-type vulnerability to early weaning. Neither castration nor ovariectomy at the age of 3 weeks affected male-type vulnerability. However, in males, castration at the age of 2 weeks attenuated the increased anxiety levels induced by early weaning, and the implantation of testosterone or estradiol, but not of dihydrotestosterone, restored the effects of early weaning. In contrast, in females, neonatal treatment with testosterone propionate together with testosterone at the age of 2 weeks, which reversed sexual behavior to the male type, did not affect anxiety levels in response to early weaning. When pregnant females were repeatedly treated with testosterone propionate on embryonic days 14, 17, and 19, in addition to testosterone treatment at the age of 2 weeks, the anxiety levels in female were increased by early weaning. Furthermore, the prenatal treatment of estradiol benzoate, but not dihydrotestosterone, induced enhanced anxiety levels by early weaning in females. These results suggest that neural systems are masculinized by estrogen from the embryonic phase to the early postnatal period and are responsible for the high levels of anxiety elicited by early weaning.     

Differential effects of the perinatal steroid environment on three sexually dimorphic parameters of the rat brain

Gonadectomy of male rats was performed at 0, 6-7 (6h), 12-13 (12h), or 24 h postnatally in order to examine the influence of testosterone exposure on sexual differentiation of the brain. The indices examined were: the volume of the sexually dimorphic nucleus of the preoptic area (SDN-POA) and luteinizing hormone (LH) and follicle-stimulating hormone (FSH) titers following estradiol benzoate (EB) and progesterone (P) administration. Control animals were sham-operated at 0 h and gonadectomized at 29 days of age (sham). A decrease in the percentage of males with elevated plasma LH levels following P was found with increasing delay before gonadectomy. Significant (P less than 0.001) differences existed in the amplitude of plasma LH titers 5 h following P administration between sham, 0 h, and 6 h groups. Follicle-stimulating hormone was also elevated in all neonatally gonadectomized male groups following P administration, but there was no difference between the groups. Volume of the SDN-POA was significantly (P less than 0.001) smaller in all gonadectomized males when compared to that of sham-operated males, but no differences existed between males gonadectomized at the different hours postpartum. In female rats gonadectomized at 0 h (F0h), LH levels were elevated 5 h following P, but only to a magnitude of 36% of that of sham-operated controls (P less than 0.001). Volume of the SDN-POA of the F0h group was significantly reduced (P less than 0.05) when compared to that of sham females. Thus, in males, the presence of the tests prenatally may be responsible for the initiation of masculinization of LH release mechanisms and the SDN-POA, but both require further androgen exposure for their completion. In addition, the LH and FSH regulating systems show a differential sensitivity to the steroid hormone environment during development that shapes the animal's response to steroid as an adult.     

New knockout model confirms a role for androgen receptors in regulating anxiety-like behaviors and HPA response in mice

Men are less likely than women to suffer from anxiety disorders. Because gonadal hormones play a crucial role in many behavioral sex differences, they may underlie sex differences in human anxiety. In rodents, testosterone (T) exerts anxiolytic effects via the androgen receptor (AR): we found that male mice with a naturally-occurring mutation rendering the AR dysfunctional, referred to as spontaneous testicular feminization mutation (sTfm), showed more anxiety-like behaviors than wildtype (WT) males. Here, we used Cre–lox recombination technology to create another dysfunctional allele for AR. These induced Tfm (iTfm) animals also displayed more anxiety-like behaviors than WTs. We further found that AR-modulation of these behaviors interacts with circadian phase. When tested in the resting phase, iTfms appeared more anxious than WTs in the open field, novel object and elevated plus maze tests, but not the light/dark box. However, when tested during the active phase (lights off), iTfms showed more anxiety-related behavior than WTs in all four tests. Finally, we confirmed a role of T acting via AR in regulating HPA axis activity, as WT males with T showed a lower baseline and overall corticosterone response, and a faster return to baseline following mild stress than did WT males without T or iTfms. These findings demonstrate that this recombined AR allele is a valuable model for studying androgenic modulation of anxiety, that the anxiolytic effects of AR in mice are more prominent in the active phase, and that HPA axis modulation by T is AR dependent.     

Sex-influenced population kinetics of Leishmania donovani in hamsters

Susceptibility of animals to infections depends upon various factors including sex of the host which plays a pivotal role. The intake of L. donovani was investigated in male and female hamsters as also in gonadectomized and hormone (sex) treated animals. Male hamsters developed more parasites (55/100 cell nuclei) than their female counterparts (22/100 cell nuclei). The hamsters receiving testosterone (250 micrograms/animal for 7 days) exogenously (im) had enhanced parasitic count (1.1-fold in male and 1.5-fold in females with respect to their respective controls). Administration of estradiol (3 micrograms/animal for 3 days) suppressed the infection in males by 2.5-fold and in female by 1.94-fold. Castration lowered the parasite 'in take' while ovarectomy promoted infection. In these (gonadectomized) animals the administration of testosterone in males restored parasite load while the estradiol therapy in females suppressed the infection. The results suggest a definite modulatory role of sex hormone, in the susceptibility of hamsters to L. donovani infection.     

Testosterone and imipramine have antidepressant effects in socially isolated male but not female rats

RationaleAffective disorders are twice as likely to occur in women as they are in men suggesting a critical role for gonadal hormones in their etiology. In particular, testosterone has been shown to have protective effects in men.ObjectiveTo investigate antidepressant effects and interactions between testosterone and imipramine in socially isolated male and female rats.MethodsA chronic social isolation model was used to induce an anxiety and depressive-like state in adult gonadectomized (Gnx) male and ovariectomized (Ovx) female rats receiving chronic testosterone and imipramine treatments. Their anxiety and depression-like behaviors were examined using the light–dark box, elevated plus maze, open field, sucrose preference and novelty induced hypophagia tests.ResultsIn socially isolated rats, the anxiolytic and antidepressant effects of testosterone and imipramine were limited to male rats. Additionally, testosterone enhanced the neurogenic effect of imipramine on hippocampal cell proliferation in male rats. Although female rats exhibited signs of anxiety and depressive-like behaviors following social isolation, testosterone and/or imipramine administration had no anxiolytic or antidepressant effects in Ovx females.ConclusionsTestosterone and imipramine had anxiolytic and antidepressant effects in socially isolated male, but not female rats. Testosterone enhanced the effect of imipramine on cell proliferation in the hippocampus of male rats.     

The Protective Effect of Semax in a Model of Stress-Induced Impairment of Memory and Behavior in White Rats

The effect of acute restrained stress on cognitive functions and anxiety-like behavior in white rats has been studied; furthermore, the influence of the fragment ACTH(4–10) analog Semax on the stress effects was evaluated. It was shown that stress exposure leads to impaired retention of previously acquired food-motivated maze task as well as reduced anxiety-like behavior in the elevated plus maze in rats. Preliminary intraperitonial administration of Semax (0.1 mg/kg) attenuates cognitive impairment caused by acute restrained stress, but it does not affect stress-induced changes in anxiety.     

A hormonal basis for sex differences in the self-grooming of rats

The self-grooming behavior of prepubescent male and female rats is described. Sex differences were observed in components of grooming addressed to the genitals, but not in other aspects of grooming. A hormonal basis for the sex difference was examined in two experiments. When females were injected with testosterone propionate (TP) on the day of birth, their subsequent grooming was found to be no different from that of control-treated females. However, males and females gonadectomized at weaning and treated daily with TP each performed significantly more genital self-grooming than oil-treated controls. There were no sex differences in gonadectomized, oil-treated rats, and sex differences in response to TP were limited to greater responsiveness of females to a 50-micrograms, but not 200-micrograms, TP dose. These results lead to the conclusion that sex differences in self-grooming can be accounted for primarily by differences in testosterone availability during the peripubertal period.     

Orexin mediates initiation of sexual behavior in sexually naive male rats, but is not critical for sexual performance

The hypothalamic neuropeptide orexin mediates arousal, sleep, and naturally rewarding behaviors, including food intake. Male sexual behavior is altered by orexin receptor-1 agonists or antagonists, suggesting a role for orexin-A in this naturally rewarding behavior. However, the specific role of endogenous orexin-A or B in different elements of male sexual behavior is currently unclear. Therefore, the current studies utilized markers for neural activation and orexin cell-specific lesions to test the hypothesis that orexin is critical for sexual motivation and performance in male rats. First, cFos expression in orexin neurons was demonstrated following presentation of a receptive or non-receptive female without further activation by different elements of mating. Next, the functional role of orexin was tested utilizing orexin-B conjugated saporin, resulting in orexin cell body lesions in the hypothalamus. Lesions were conducted in sexually naive males and subsequent sexual behavior was recorded during four mating trials. Lesion males showed shortened latencies to mount and intromit during the first, but not subsequent mating trials, suggesting lesions facilitated initiation of sexual behavior in sexually naive, but not experienced males. Likewise, lesions did not affect sexual motivation in experienced males, determined by runway tests. Finally, elevated plus maze tests demonstrated reduced anxiety-like behaviors in lesioned males, supporting a role for orexin in anxiety associated with initial exposure to the female in naive animals. Overall, these findings show that orexin is not critical for male sexual performance or motivation, but may play a role in arousal and anxiety related to sexual behavior in naive animals.     

Ingestion of Lactobacillus strain reduces anxiety and improves cognitive function in the hyperammonemia rat

Evidence suggests that the hyperammonemia(HA)-induced neuroinflammation and alterations in the serotonin(5-HT)system may contribute to cognitive decline and anxiety disorder during hepatic encephalopathy(HE).Probiotics that maintain immune system homeostasis and regulate the 5-HT system may be potential treatment for HA-mediated neurological disorders in HE.In this study,we tested the efficacy of probiotic Lactobacillus helveticus strain NS8 in preventing cognitive decline and anxiety-like behavior in HA rats.Chronic HA was induced by intraperitoneal injection of ammonium acetate for four weeks in male Sprague-Dawley rats.HA rats were then given Lactobacillus helveticus strain NS8(109 CFU mL?1)in drinking water as a daily supplementation.The Morris water maze task assessed cognitive function,and the elevated plus maze test evaluated anxiety-like behavior.Neuroinflammation was assessed by measuring the inflammatory markers:inducible nitric oxide synthase,prostaglandin E2,and interleukin-1βin the brain.5-HT system activity was evaluated by measuring 5-HT and its metabolite,5-HIAA,and the 5-HT precursor,tryptophan.Probiotic treatment of HA rats significantly reduced the level of inflammatory markers,decreased 5-HT metabolism,restored cognitive function and improved anxiety-like behavior.These results indicate that probiotic L.helveticus strain NS8 is beneficial for the treatment of cognitive decline and anxiety-like behavior in HA rats.     

The role of testicular hormones and luteinizing hormone in spatial memory in adult male rats

Attempts to determine the influence of testicular hormones on learning and memory in males have yielded contradictory results. The present studies examined whether testicular hormones are important for maximal levels of spatial memory in young adult male rats. To minimize any effect of stress, we used the Object Location Task which is a spatial working memory task that does not involve food or water deprivation or aversive stimuli for motivation. In Experiment 1 sham gonadectomized male rats demonstrated robust spatial memory, but gonadectomized males showed diminished spatial memory. In Experiment 2 subcutaneous testosterone (T) capsules restored spatial memory performance in gonadectomized male rats, while rats with blank capsules demonstrated compromised spatial memory. In Experiment 3, gonadectomized male rats implanted with blank capsules again showed compromised spatial memory, while those with T, dihydrotestosterone (DHT), or estradiol (E) capsules demonstrated robust spatial memory, indicating that T's effects may be mediated by its conversion to E or to DHT. Gonadectomized male rats injected with Antide, a gonadotropin-releasing hormone receptor antagonist which lowers luteinizing hormone levels, also demonstrated spatial memory, comparable to that shown by T-, E-, or DHT-treated males. These data indicate that testicular androgens are important for maximal levels of spatial working memory in male rats, that testosterone may be converted to E and/or DHT to exert its effects, and that some of the effects of these steroid hormones may occur via negative feedback effects on LH.     

Effects of testosterone and estrogen on hepatic levels of cytochromes P450 2C7 and P450 2C11 in the rat.

The effects of exogenous hormone treatment on the expression of cytochromes P450 2C7 and P450 2C11 were studied in neonatally gonadectomized and sham-operated male and female rats. Hepatic levels of cytochrome P450 2C7 were found to be two- to threefold higher in intact adult female versus male rats. Neonatal gonadectomy resulted in a reversal of the relative cytochrome P450 2C7 levels in male and female animals at maturity. Expression of this isozyme was restored in ovariectomized females by estradiol treatment. Furthermore, neonatal and/or pubertal administration of estradiol to intact male rats induced cytochrome P450 2C7 to adult female levels. On the other hand, administration of testosterone at all times examined had no effect in intact female rats, but decreased cytochrome P450 2C7 to normal levels in neonatally castrated males treated during adulthood. Neonatal testosterone treatment also increased hepatic cytochrome P450 2C7 content in both ovariectomized females and intact males. These results indicate that estrogen is required for full expression of cytochrome P450 2C7 while the effect of testosterone is ambiguous. In comparison, neonatal gonadectomy of male rats abolished the adult expression of cytochrome P450 2C11. Normal levels were restored only by treatment with testosterone during adulthood. Neonatal testosterone treatment did not induce cytochrome P450 2C11 levels in gonadectomized rats of either sex. In contrast, neonatal estrogen treatment suppressed cytochrome P450 2C11 expression in intact adult male rats to the same extent as neonatal castration. These results indicate that androgen exposure during the adult, and not the neonatal, phase is essential for full expression of cytochrome P450 2C11.     

Mice with the testicular feminization mutation demonstrate a role for androgen receptors in the regulation of anxiety-related behaviors and the hypothalamic–pituitary–adrenal axis

Testosterone (T) appears to play a role in anxiety and sensorimotor gating in rodents, but whether T acts through the androgen receptor (AR) to influence these behaviors is less clear. We compared adult genetic male mice with the testicular feminization mutation (Tfm), which lack functional ARs, to wild type male littermates (wt males) on an assay of sensorimotor gating (prepulse inhibition of the acoustic startle response; PPI) and several tests thought to reflect anxiety: open field exposure, novel object exposure, elevated plus maze (EPM), and light/dark (LD) box. PPI was similar between groups, but indices of anxiety in the novel object and LD box tests were increased in Tfm males with no significant differences found in the open field or EPM. Since Tfm male mice have decreased circulating T, the same tests were conducted in mice that were gonadectomized (wt males) or sham-operated (Tfm males) as adults and supplemented with T or nothing (B). While T treatment reduced indices of anxiety in the novel object and LD box tests in wt males, it was ineffective in Tfm males. Increased indices of anxiety in Tfm males appear to be related to hyper-activation of the hypothalamic–pituitary–adrenal axis since levels of the stress hormone corticosterone were elevated in Tfm males compared to wt males at baseline and at several time points after exposure to a novel object. These findings demonstrate that ARs influence anxiety and stress responses in mice.     

Differences in anxiety-related behavior between apolipoprotein E-deficient C57BL/6 and wild type C57BL/6 mice

The influence of ApoE gene deletion on the anxiety state has not been previously investigated. The elevated plus maze was used in this study to determine differences in anxiety-related behavior between apoE-deficient and wild type C57BL/6 mice. The apoE-deficient mice demonstrated less anxiety on the elevated plus maze by spending more time in the open arms of the elevated plus maze compared to wild type mice (p<0.001). Additionally, female apoE-deficient mice visited the open arm of the maze more often than their apoE-deficient male counterpart (p<0.05). The anxiety state and/or sex are possible variables to be considered when designing physiological and/or behavioral studies involving mice that are apoE-deficient.     

Sex differences in adrenocortical structure and function. VI. Long-term effect of gonadectomy and testosterone or estradiol replacement on rat adrenal cortex

Gonadectomy or sham operations were performed on rats of Wistar strain weighing 150--170 g. Animals were sacrificed in groups, males 397 days and females 395 days after surgery. 14 days before autopsy some of the gonadectomized rats were injected with a single dose of depo-testosterone or depo-estradiol respectively. Absolute as well as relative adrenal weights were larger in female than in male rats. Long-term orchiectomy did not change these weights while testosterone, if compared with control, lowers the weights of the adrenals. Long-term ovariectomy lowers the weights of the glands and estradiol reversed them to the control level. Adrenal cortex of rats studied show irregular zonation. Within the zona fasciculata two types of cells were observed--with strongly eosinophilic and pale cytoplasm. In some cases groups of hypertrophied cells surrounded by spindle-shaped cells were observed. Neither long-term gonadectomy nor gonadal hormone replacement has a marked effect on the structure of the adrenal cortex. In male and female rats there was no difference in the volume of single zona fasciculata and reticularis cells. Long-term orchiectomy has no effect on this volume while testosterone resulted in a lowering of volume of zona fasiculata cells. Neither long-term ovariectomy nor estradiol has an effect on morphometric parameters studied in the fasciculata and reticularis zones. Distinct sex differences and dependence on sex hormones were found in reactions for alpha-glycerophosphate dehydrogenase and nonspecific esterases activity.     

Sex-related differences in the hepatobiliary transport of phenolsulfonphthalein in the rat

Sex-related differences in the hepatobiliary transport of phenolsulfonphthalein (PSP) were investigated in male and female Wistar rats. Maximal biliary excretion of unconjugated PSP was significantly higher in females while the excretion of the conjugated dye and liver UDP-glucuronosyltransferase activity toward PSP were higher in male animals. Orchidectomy decreased enzyme activity and excretion of the conjugate, whereas ovariectomy produced the opposite effect. Both in gonadectomized males and females maximal biliary excretion of the unconjugated dye was significantly reduced. Testosterone treatment increased the excretion of both conjugated and unconjugated PSP and transferase activity in orchidectomized males. Combined treatment of gonadectomized females with estradiol plus progesterone led to excretions of both conjugated and unconjugated PSP and UDP-glucoronosyltransferase activities similar to those found in control rats. These data indicate the existence of sex-related differences in the conjugation and biliary excretion of PSP in the rat and its modulation by sex hormones.     

Mice with the testicular feminization mutation demonstrate a role for androgen receptors in the regulation of anxiety-related behaviors and the hypothalamic-pituitary-adrenal axis

Testosterone (T) appears to play a role in anxiety and sensorimotor gating in rodents, but whether T acts through the androgen receptor (AR) to influence these behaviors is less clear. We compared adult genetic male mice with the testicular feminization mutation (Tfm), which lack functional ARs, to wild type male littermates (wt males) on an assay of sensorimotor gating (prepulse inhibition of the acoustic startle response; PPI) and several tests thought to reflect anxiety: open field exposure, novel object exposure, elevated plus maze (EPM), and light/dark (LD) box. PPI was similar between groups, but indices of anxiety in the novel object and LD box tests were increased in Tfm males with no significant differences found in the open field or EPM. Since Tfm male mice have decreased circulating T, the same tests were conducted in mice that were gonadectomized (wt males) or sham-operated (Tfm males) as adults and supplemented with T or nothing (B). While T treatment reduced indices of anxiety in the novel object and LD box tests in wt males, it was ineffective in Tfm males. Increased indices of anxiety in Tfm males appear to be related to hyper-activation of the hypothalamic–pituitary–adrenal axis since levels of the stress hormone corticosterone were elevated in Tfm males compared to wt males at baseline and at several time points after exposure to a novel object. These findings demonstrate that ARs influence anxiety and stress responses in mice.     

Effects of septal lesions on the lordotic behavior of weanling male and female rats

Twenty-four-day-old weanling male and female rats were either lesioned in the septal area, gonadectomized, lesioned and gonadectomized, or not treated. Tests for lordotic behavior were carried out at 27 and 28 days of age after priming with estradiol alone and with estradiol plus progesterone. A second set of tests for lordotic behavior was carried out at 47 and 48 days of age. In the interim period, some of the animals were given chronic estrogen treatment. In tests given at 27 days of age, septal lesions facilitated lordotic responding after estrogen priming; no differences were observed between male and female animals. At 47 and 48 days of age, however, unless male rats had been exposed to chronic estrogens following septal lesioning, no facilitation of estrogen-induced lordotic behavior occurred. In addition, it was found that female rats gonadectomized at 24 days of age and given no exposure to estrogens between the tests at 27 and 28 days and those at 47 and 48 days of age showed reduced sensitivity to estrogens, as compared to normal or estrogen-treated females, whether lesioned or not.