Sex differences in open-field behavior in the rat: The inductive and activational role of gonadal hormones

Male rats are generally found to be significantly less active and to defecate more than female rats in the open-field test. In developmental studies, these differences were shown to emerge in the peripuberal period at 50–60 days of age. After gonadectomy in adult life, the sex difference in open-field activity was reduced in magnitude but did persist indicating that it was not solely dependent on the presence of gonadal secretions in either sex. In contrast, the sex difference in open-field defecation did not reach statistical significance after gonadectomy with ovariectomized females exhibiting increased defecation frequency. Estrogen-progesterone replacement stimulated activity and decreased defecation scores to pre-gonadectomy levels in females but lacked effect in males. Administration of testosterone propionate (TP) to female rats in neonatal life produced developmental changes in open-field behavior similar to those seen in the male. In addition, the response of TP treated females to gonadectomy and hormone replacement was similar to that found for the male. These results suggest that the presence of testosterone in neonatal life plays a role in the masculinisation of open-field behavior and that its effect is partly mediated by the presence of cyclic ovarian secretions in the adult female rat and their absence in the male and TP treated female.     

Endothelial-independent prevention of high blood pressure in L-NAME-treated rats by angiotensin II type I receptor antisense gene therapy

It has previously been established that a single systemic administration of retroviral vector containing angiotensin II type I receptor antisense (AT(1)R-AS) in the neonatal spontaneously hypertensive rat (SHR) prevents development of hypertension, and in addition cardiac hypertrophy and endothelial dysfunction. However, these studies could not determine whether the effects of AT(1)R-AS on high blood pressure (BP) and endothelial function were independent. Angiotensin receptor blockers have been shown to reduce BP in the L-NAME (N (omega)-nitro-L-arginine methyl ester hydrochloride)-induced rat model of hypertension. Our objective in the present study was to use the L-NAME model of hypertension to determine whether AT(1)R-AS treatment would lower high BP and attenuate cardiac hypertrophy under conditions of permanent endothelial damage. A single bolus of LNSV-AT(1)R-AS viral particles in neonatal Wistar-Kyoto (WKY) rats was without affect on basal BP. Efficacy of the transgene incorporation was assessed by observing a significant reduction in angiotensin-induced dipsogenic response in the AT(1)R-AS-treated animals. Introduction of L-NAME in the drinking water for 10 weeks resulted in the establishment of hypertension only in the WKY rats treated with vector alone. These hypertensive (BP, 179 +/- 4 mmHg) animals showed a 17 % increase in heart weight/body weight ratio and a 60 % reduction in ACh-induced vasorelaxation in phenylephrine-preconstricted arteries. The L-NAME-induced high BP and cardiac hypertrophy were attenuated in rats expressing AT(1)R-AS. However, endothelial dysfunction could not be prevented with the antisense therapy. These observations demonstrate that attenuation of endothelial dysfunction is not a prerequisite for the antihypertensive effects of AT(1)R-AS treatment.     

Chronic stress effects on dendritic morphology in medial prefrontal cortex: sex differences and estrogen dependence

A growing body of work has documented sex differences in many behavioral, neurochemical, and morphological responses to stress. Chronic stress alters morphology of dendrites in medial prefrontal cortex in male rats. However, potential sex differences in stress-induced morphological changes in medial prefrontal cortex have not been examined. Thus, in Experiment 1 we assessed dendritic morphology in medial prefrontal cortex in male and female rats after chronic stress. Male and female rats underwent either 3 hours of restraint daily for 1 week or were left unhandled except for weighing. On the final day of restraint, all rats were euthanized and brains were stained using a Golgi–Cox procedure. Pyramidal neurons in layer II–III of medial prefrontal cortex were drawn in three dimensions, and morphology of apical and basilar arbors was quantified. In males, stress decreased apical dendritic branch number and length, whereas in females, stress increased apical dendritic length. In Experiment 2, we assessed whether estradiol mediates this stress-induced dendritic hypertrophy in females by assessing the effects of restraint stress on female rats that had received either ovariectomy with or without 17-β-estradiol replacement or sham ovariectomy. Brains were processed and neurons reconstructed as described in Experiment 1. Both sham-operated and ovariectomized rats with estradiol implants showed stress-induced increases in apical dendritic material, whereas ovariectomy without estradiol replacement prevented the stress-induced increase. Thus, the stress-induced increase in apical dendritic material in females is estradiol-dependent.     

Testosterone-dependent transgene expression in the liver of the CAG-lacZ transgenic rat

Many endogenous gene expressions in the liver are well known to be predominant in males, compared with those of females. In contrast, the fate of hepatic transgene expression between sexes is not fully understood. Here we studied whether sex hormones changed hepatic transgene expression in the ubiquitous CAG promoter-driven lacZ transgenic (Tg) rat. Both sexes of CAG-lacZ Tg rats received gonadectomy. Liver biopsy was taken weekly to determine the change of transgene expression. Histological result of adult males showed mosaic lacZ expression but it was negative in adult females, while livers in neonatal stage showed comparable expression of lacZ. Other organs exhibited equal expression in both sexes. At 2 weeks after castration, lacZ expression in male liver was significantly decreased and became negative after 4 weeks while no significant difference was observed in the lacZ expression pattern in other organs. After ovariectomy, lacZ expression in female liver remained undetectable. Moreover, testosterone treatment to gonadectomized rats of both sexes could enhance lacZ expression in the liver. In summary, we report that CAG-lacZ Tg rats demonstrate sexual dimorphism of transgene expression specifically only in the liver. Testosterone administration mediated upregulation of liver lacZ expression. Our findings suggested that androgen, especially testosterone, plays an important role in the hepatic transgene expression.     

Sex differences in hippocampal slice excitability: role of testosterone

In vivo fluctuations in gonadal hormones alter hippocampal excitability and modulate both physiological and pathological hippocampal processes. To assess hormonal effects on excitability within a functional hippocampal circuit, extracellular CA1 field responses were compared in slices from intact male, intact female, orchidectomized male, and ovariectomized female rats. Secondly, the effects of in vitro applications of 17-beta estradiol, progesterone, or testosterone on baseline excitability of slices from gonadectomized rats were assessed versus pre-hormone baseline measures. Finally, using the in vitro kindling model of slice epileptogenesis, steroid hormone effects on interictal-like activity were also examined. Significant sex differences in excitatory postsynaptic potential amplitude were observed, with slices from males having larger excitatory postsynaptic potential amplitudes than those from females. Gonadectomy significantly decreased excitatory postsynaptic potential amplitude in slices from male rats. Slices from gonadectomized male and female rats also showed a decreased dendritic excitatory postsynaptic potential slope relative to slices from intact male and females rats. In vitro application of testosterone significantly increased excitatory postsynaptic potential amplitudes in slices from both orchidectomized males and ovariectomized females and the population spike amplitude of slices from ovariectomized females. Following in vitro kindling, slices from intact males showed greater spontaneous burst rates than slices from intact females, further suggesting an excitatory effect of testosterone.These results suggest: (1) a sex difference in the level of baseline excitability between slices from intact males and females as measured by excitatory postsynaptic potential amplitudes, (2) testosterone has excitatory effects on baseline physiology and kindled hippocampal responses, and (3) slices from males show a greater level of excitability than those from females in the in vitro kindling model.     

Endothelium-dependent relaxation in pulmonary arteries of L-NAME-treated Wistar and stroke-prone spontaneously hypertensive rats.

To evaluate whether the elevated blood pressure induced by chronic treatment with N(omega)-nitro-L-arginine methyl ester (L-NAME) contributes to an impairment of endothelium-dependent relaxation (EDR), the effects of chronic treatment of Wistar rats with L-NAME on systolic blood pressure, pulmonary arterial blood pressure and EDR of the pulmonary arteries were studied and compared with those of stroke-prone spontaneously hypertensive rats (SHRSP). While the systolic blood pressure (SBP) of Wistar rats was increased above that of controls by chronic treatment with L-NAME, it was still significantly lower than that of SHRSP. Chronic treatment with L-NAME did not affect pulmonary arterial blood pressure. On the other hand, the pulmonary arterial blood pressure of SHRSP was slightly but significantly higher than that of the control normotensive Wistar Kyoto rats (WKY). EDR in response to acetylcholine in the pulmonary artery of L-NAME-treated rats was significantly smaller than that in control Wistar rats. The EDR markedly increased in the presence of L-arginine and completely disappeared in the presence of N(omega)-nitro-L-arginine. Indomethacin hardly affected EDR. In preparations from SHRSP, the EDR was not different from that in those from WKY. Relaxation induced by sodium nitroprusside was identical in all preparations. Elevation of SBP and the impairment of EDR observed in L-NAME-treated rats recovered two weeks following cessation of treatment. These results suggest that the impaired EDR in the pulmonary artery of L-NAME-treated rats is not due to an L-NAME-induced increase in blood pressure but due to the inhibition of nitric oxide synthase by the drug remaining in the endothelium.     

Sexual dimorphism in the contribution of protein kinase C isoforms to nociception in the streptozotocin diabetic rat

The contribution of second messenger signaling, glucose level and sex hormones to sexual dimorphism in the streptozotocin model of diabetic painful peripheral neuropathy was evaluated. Streptozotocin induced elevation of blood glucose and mechanical hyperalgesia (measured by the Randall-Selitto paw-withdrawal test) were both greater in female rats. Ovariectomy abolished and estrogen implants reconstituted this sexual dimorphism; gonadectomy in males had no effect. An inhibitor of protein kinase Cε attenuated hyperalgesia in males and ovariectomized females, but not in normal females or in ovariectomized females with estrogen implants, whereas inhibitors of protein kinase Cδ attenuated hyperalgesia in females but not in males. Inhibitors of protein kinase A, protein kinase C (non-selective), protein kinase G and nitric oxide synthase attenuated hyperalgesia equally in both sexes. Higher blood glucose levels in diabetic females were also sex hormone dependent, and magnitude of hyperalgesia correlated with blood glucose level in diabetic male and female rats. These results demonstrate sexual dimorphism in diabetic hyperalgesia, mediated by sex hormone dependent differences in protein kinase Cε and protein kinase Cδ signaling and blood glucose levels and suggest that sex may be an important factor to be considered in the treatment of symptomatic diabetic neuropathy.     

Developmental effects of neonatal sex hormones on spatial and activity skills in the white rat.

The purpose of this experiment is to extend these experiments by using varying neonatal hormonal differentiation processes and to examine the effects of these different methods of neonatal feminization on changes in spatial and activity skills. The experiment involved three neonatally feminized groups of male rats: 'castration alone', 'estrogen alone', and 'estrogen injected castrates' together with one female sample, 'testosterone injected castrates', while treated and untreated male and female controls were also used. The data provide partial support for the major hypotheses that neonatal gonadectomy and opposite sex hormones administered to male and female castrated rats (together and independently for males) would reverse the normal sex-associated abilities of the white rate (higher male spatial learning and higher female activity). However, the feminization effect for the male 'estrogen alone' and 'castration alone' experimental groups was much greater than for the male estrogen injected castrates. The masculinized females, testosterone injected castrates, also had higher spatial learning and lower activity levels, while the feminized male's spatial and activity skills were also reversed. This confirmed in part the extent to which neonatal gonadal sex hormones are effective at critical periods of development in programming the brain in terms of sex-associated spatial and activity skills. Adult hormonal replacement therapy was also administered at 12 months and supported the hypothesis that sex hormones in adults would be mainly activational and have less marked effects than the significant directional changes obtained by these neonatal sex hormones and castration techniques.     

Photoperiod and gonadal hormones influence odor preferences of the male meadow vole, Microtus pennsylvanicus.

Male meadow voles housed in a long photoperiod (14 h light/day, LP) preferred female to male odors, whereas males maintained in a short photoperiod (10 h light/day, SP) did not display preferences for odors of either sex. These odor-preference patterns matched those of free-living males during spring and autumn, respectively. The preference of LP male voles for female over male odors was eliminated by gonadectomy and reinstated by treatment with testosterone. In SP males, although gonadectomy did not affect odor choices, a preference for female odors was induced by testosterone treatment. Treatment with estradiol did not alter odor preferences of LP or SP males. In conjunction with previous result, the present findings suggest that hormonal responsiveness of neural substrates that control odor preferences are sexually dimorphic and may reflect sex differences in reproductive strategies.     

Behavior of male and female rats with septal lesions: Influence of prior gonadectomy

The influence of gonadectomy at different ages upon the behavioral changes induced by septal lesions in adulthood was examined in male and female rats. In both sexes septal lesions increased emotionality, facilitated acquisition of shuttle avoidance responding, increased escape from light onset, depressed rearing in the open field, increased the number of shocks received during passive avoidance acquisition, but did not affect the number of sessions required to acquire or to extinguish the passive avoidance response. In males prepuberal, but not neonatal or adult gonadectomy attenuated hyperemotionality, but gonadectomy had little influence on other behavioral changes produced by septal lesions. Ovariectomy, either prepuberally or in adulthood, also attenuated hyperemotionality in females with lesions. Adult, but not prepuberal ovariectomy rendered females with septal lesions somewhat hypoactive compared to other female groups. Among animals without lesions, females reared more, entered more squares and were less likely to defecate than males during open field tests. Neonatal, but not prepuberal or adult gonadectomy increased both rearing and activity in males. Females also acquired active avoidance behvior and extinguished passive avoidance behavior more rapidly than males and gonadectomy at any age did not abolish the sex differences in these behaviors. Sex differences in behavior and the effects of gonadectomy were generally similar in controls and in animals with septal lesions, suggesting that differences in septal function are not likely to underly sex differences in these behaviors.     

Discriminated lever press avoidance conditioning in male and female rats

Discriminated lever press avoidance and step through passive avoidance conditioning were studied in male and female rats. In lever press avoidance female rats showed superior avoidance performance, less intertrial responding and shorter durations of leverholding than male rats. Castration of males tended to improve their avoidance performance, whereas ovariectomy had no effect. In passive avoidance more males than females showed an avoidance response. From these results it is concluded that aversive stimulation seems to elicit different behavior tendencies in male and female rats rendering males less prepared for active avoidance, but at the same time more prepared for passive avoidance conditioning. The data do not support the hypothesis that sex differences in activity underly the dimorphism in active and passive avoidance conditioning.     

Acute effects of sex-specific sex hormones on heat shock proteins in fast muscle of male and female rats

Heat shock protein (HSP) expression and sex hormone levels have been shown to influence several aspects of skeletal muscle physiology (e.g., hypertrophy, resistance to oxidative stress), suggesting that sex hormone levels can effect HSP expression. This study evaluated the effects of differing levels of sex-specific sex hormones (i.e., testosterone in males and estrogen in females) on the expression of 4: HSP70, HSC70, HSP25, and αB-crystallin in the quadriceps muscles of male and female rats. Animals were assigned to 1 of 3 groups (n = 5 M and F/group). The first group (Ctl) consisted of typically cage-housed animals that served as controls. The second group (H) was gonadectomized and received either testosterone (males) or estradiol (females) via injection for 12 consecutive days. The third group (Gx) was gonadectomized and injected as above, but with vehicle only, rather than hormones. Significant sex by condition interactions (P < 0.05 by two-way MANOVA) were found for all 4 proteins studied, except for HSP70, which exhibited a significant effect of condition only. The expression of all HSPs was greater (1.9–2.5-fold) in males vs. females in the Ctl group, except for HSP70, which was no different. Generally, gonadectomy appeared to have greater effects in males than females, but administration of the exogenous sex hormones tended to produce more robust relative changes in females than males. There were no differences in myosin composition in any of the groups, suggesting that changes in fiber type were not a factor in the differential protein expression. These data may have implications for sex-related differences in muscular responses to exercise, disuse, and injury.     

Hypertension and impairment of endothelium-dependent relaxation of arteries from spontaneously hypertensive and L-NAME-treated Wistar rats.

Effects of chronic treatment of normotensive Wistar rats with N(omega)-nitro-L-arginine methyl ester (L-NAME) on blood pressure and on endothelium-dependent relaxation of the aorta, carotid and iliac arteries were studied. The endothelium-dependent relaxation was compared in arteries from normotensive Wistar Kyoto rats (WKY) and genetically hypertensive rats (stroke-prone spontaneously hypertensive rats, SHRSP). Chronic treatment of normotensive Wistar rats with L-NAME caused an elevation of blood pressure. The elevated blood pressure at 15 weeks of age was significantly higher in these animals than that of untreated Wistar rats, but lower than that of SHRSP. Endothelium-dependent relaxation of the arteries induced by acetylcholine (ACh) was almost abolished by chronic treatment with L-NAME. The remaining small relaxation in arteries from L-NAME-treated rats was completely inhibited by application of L-NAME (10(-4) M). In such preparations, higher concentrations of ACh induced a contraction, which was abolished by removal of the endothelium or by an application of indomethacin (10(-5) M). Endothelium-independent relaxation induced by sodium nitroprusside was similar between preparations from untreated and L-NAME-treated Wistar rats. Endothelium-dependent relaxation was significantly impaired in preparations from SHRSP, when compared with that in those from WKY. However, the impairment was less prominent in preparations from SHRSP than in those from L-NAME-treated rats. These results suggest that the impairment of endothelium-dependent relaxation in the arteries from L-NAME-treated rats is not due to the elevated blood pressure resulting from the chronic treatment, and that impairment of NO synthesis by the endothelium does not play a major role in the initiation of hypertension in SHRSP.     

Ontogeny of sex differences in LH and FSH levels 48 h after castration in the rat

Serum gonadotropin levels were measured 12, 24, and 48 h after gonadectomy in male and female rats (ages, 22--60 days) to assess when during development the rate of rise of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) after castration approximates that seen in the gonadectomized adult. In females serum LH levels 48 h after ovariectomy were increased above sham levels only when the ovaries were removed prior to vaginal opening. Ovariectomy on the day of vaginal opening or at older ages resulted in no increase in LH levels by 48 h after surgery. Serum FSH levels at 24 and 48 h after ovariectomy declined with increasing age at the time of ovariectomy. In males serum LH levels at 48 h after castration increased with increasing age at the time of gonadectomy. Serum FSH levels at either 12, 24, or 48 h after orchidectomy did not change appreciably with age at the time of surgery. It is concluded that the acute pituitary secretion of gonadotropins after removal of testes in the immature male resembles that seen in the mature male early in the course of the development of sexual maturity. In contrast, the acute pituitary secretion of gonadotropins after removal of the ovaries in the immature female does not resemble that seen in the ovariectomized adult until she is mature and capable of ovulating. Thus, the observed delay in the rise of LH seen in ovariectomized adults may be a function of some aspect of the hormonal changes associated with the estrous cycle.     

雄激素对大鼠心内神经节神经生长因子表达的影响

目的 观察神经生长因子(NGF)在成年雄性大鼠、雌性大鼠、雌性大鼠睾酮处理组、睾丸切除大鼠、睾丸切除后睾酮替代大鼠心房后壁心内神经节的表达及变化。方法 免疫组织化学染色。结果 各组大鼠心内神经节均存在NGF阳性神经元,但睾丸切除组大鼠心内神经节NGF阳性神经元的数量明显减少,表达明显降低。结论 心内神经节细胞内含NGF;雄激素可能影响心内神经节细胞的NGF表达。     

Effects of sex hormones on induction of intestinal metaplasia by X-irradiation in rats.

The influence of sex hormones on induction of intestinal rnetaplasia was examined in 5 week old Crj: CD (SD) rats of both sexes. At the age of 4 weeks, the animals were gonadectornized and given testosterone or dimethyl estradiol (DES). One week after operation, they were irradiated with two 10 Gy doses of X-rays to the gastric region at a 3 day interval for a total of 20 Gy. At the termination of the experiment, 6 months after the X-irradiation, the incidence of intestinal metaplasia with alkaline phosphatase (ALP) positive foci in males was significantly higher than in females, in orchidectomized males or orchidectomized plus DES treated rats (P <0.01). On the other hand, the incidence of intestinal metaplasia with ALPpositive foci in normal females appeared lower than in ovariectomized females (P < 0.01), and was increased in rats by treatment with testosterone or decreased by DES. Numbers of foci of intestinal metaplasias with Paneth cells and total numbers appeared to increase in males treated with DES. The results suggested a promoting role for testosterone in the development of ALP positive lesions and indicated considerable heterogeneity between intestinal metaplasia subtypes.     

Ontogeny of sex differences in open-field ambulation in the rat

The effects of age and gonads were studied in rats subjected to open-field tests, during which ambulation behavior was recorded. Subjects were three groups of male and female rats: sham-operation on day 1 and day 21; gonadectomy on day 1 and sham-operation on day 21; and sham-operation on day 1 and gonadectomy on day 21. Half of each group were tested in a circular open field (3 min/day, 3 consecutive days) on days 28-30; the others were tested on days 47-49. Representatives of both batches were tested again in a square open field on days 76-78. There was a sex difference in ambulation at 77 days, but not at earlier ages. In animals gonadectomized on day 1 or day 21 the sex difference in adulthood failed to occur, because castration caused the males to ambulate as much as sham-operated and ovariectomized females. On the basis of our results and reports in the literature it is suggested that testicular secretions around puberty have an organizing effect on ambulation behavior. The intact adult male rat ambulates less than the adult female and this difference persists after castration in adulthood. Castration well before puberty prevents the development of the adult sex difference.     

Effects of sex hormones on associative learning in spontaneously hypertensive rats

Pavlovian conditioning of a visual stimulus paired with food was examined in spontaneously hypertensive rats (SHR), which are a commonly used model for Attention-Deficit/Hyperactivity Disorder (ADHD), and in Wistar rats (normoactive control). In gonadally intact rats of both strains, males spent more time in the food cup following onset of the light than did females, indicating a stronger association of the conditioned stimulus (CS) with reward. Gonadectomy carried out in adulthood affected conditioning differently in the two strains. In Wistar rats, gonadectomy had no effect on conditioned responding in females, but reduced conditioned responding in males, effectively eliminating the sex difference in behavior. This result suggests that circulating androgens in male Wistar rats normally aid conditioning in this task. In contrast, gonadectomy enhanced conditioning in both sexes in the SHR rats, indicating that androgens and/or estrogens impair conditioned associations in this strain. These data indicate that gonadal steroids can influence conditioning in rats and that the valence of steroid action on this behavior is strain-dependent. To the extent that SHR serves as a model of ADHD in humans, the influence of steroids on associative learning may play a role in the expression of ADHD-like behaviors.     

Effects of sex,gonadectomy, and oestrogen substitution on ischaemic preconditioning and ischaemia-reperfusion injury in mice

AIM: Ischaemic preconditioning (IPC) has been demonstrated to protect heart function and viability, but has been predominantly studied in male animals. METHODS: We studied a possible influence of sex and oestrogen for protection in IPC. Infarct size and heart function after 40 min global ischaemia and 60 min reperfusion with or without preceding classic IPC was investigated in Langendorff-perfused hearts. Hearts were harvested from 10-week-old male and female C57BL6 mice with or without gonadectomy 6 weeks earlier, or gonadectomy and substitution with 17 beta-oestradiol for 4 weeks (n = 104). RESULTS: Classic IPC reduced depression of left ventricular developed pressure (P < 0.01), attenuated the increase of end-diastolic pressure (P < 0.01), and reduced infarct size (P < 0.01) in hearts of untreated male mice, but failed to protect untreated females which had improved functional recovery and smaller infarctions than untreated males. After gonadectomy of female mice, developed pressure was reduced (P < 0.01) and infarct size increased (P < 0.01) compared with normal females, with no protection of preconditioning. The changes were not reversed by 17 beta-oestradiol substitution. In hearts of gonadectomized males, the post-ischaemic increase of end-diastolic pressure was attenuated (P < 0.01), and enhanced after substitution with 17 beta-oestradiol (P < 0.01). The preconditioning effect disappeared after gonadectomy and gonadectomy with substitution in male mice. CONCLUSION: There is a sex difference in evoking preconditioning in male and female mice which is only partially dependent on sex hormones.