Testicular hormone exposure during adolescence organizes flank-marking behavior and vasopressin receptor binding in the lateral septum

Adolescence is a period during which many social behaviors emerge. One such behavior, flank marking, is a testosterone-modulated scent marking behavior that communicates dominance status between adult male Syrian hamsters. Testosterone modulates flank-marking behavior by altering neural transmission of vasopressin within a forebrain circuit. This study tested whether testicular hormones secreted during adolescence play purely a transient activational role in the display of flank-marking behavior, or whether adolescent steroid hormone secretions also cause long-term organizational changes in vasopressin binding within brain regions underlying flank-marking behavior. We tested this hypothesis by manipulating whether testicular secretions were present during adolescent development and then tested for flank-marking behavior and vasopressin receptor binding within the flank-marking neural circuit in young adulthood. Specifically, males were gonadectomized immediately before or after adolescence, replaced with testosterone 6 weeks following gonadectomy in young adulthood, and behavior tested 1 week later. Adult testosterone treatment activated flank-marking behavior only in males that were exposed to testicular hormones during adolescence. In addition, males exposed to testicular hormones during adolescence exhibited significantly less vasopressin receptor binding within the lateral septum than males deprived of adolescent hormones, suggesting that hormone-dependent remodeling of synapses normally occurs in the lateral septum during adolescence. These data highlight the importance of gonadal steroid hormone exposure during adolescence for the organization of neural circuits and social behavior.     

Sex Steroid–Opioid Interactions Associated with the Temporal Component of Avian Calling Patterns

Isolation from conspecifics in young, precocial birds predictably induces distress vocalizations (DV) and androgens change this type of vocalization into male typical “crowing” (CR). In addition, opioid peptides are known to exert potent effects on avian vocal behavior. Here we investigate the organizational and activational correlates of sex-steroid actions on opioid-receptor organization and their relevance to the temporal evolution of DV and CR. From the effects of pre- and postnatal steroid applications and postnatal [³H]etorphin binding studies, we find that early steroidal effects become manifested at the behavioral level by changing the characteristic duration of vocalizations. In the male quail this extension of calling duration is accompanied by a clear decrease in opiate binding, whereas in the female there is a moderate increase in binding sites. The transition from DV to CR (within hours) induced by testosterone is correlated with “upregulation” of opiate receptor sites within unilateral brainstem areas of young male quail. Based on these findings, we suggest that organizational steroid effects change the characteristic duration of isolation-induced vocalizations and these effects appear to be manifested at the level of opioid-receptor distribution.     

Sexual experience and testosterone during adolescence alter adult neuronal morphology and behavior

Steroid hormones released immediately before and after birth provoke sexual differentiation of neural circuits. Further, steroid hormones secreted during adolescence also exert long lasting effects on the nervous system. Hormones secreted during development may act through two distinct pathways: (1) hormones can directly affect neuron and synapse elimination and (2) endocrine changes in the nervous system may occur secondary to changes in social behaviors. Therefore, a critical period for organization of the nervous system by steroid hormones during adolescence may also be a sensitive period for the effects of social experience. The overall goal of this experiment was to determine whether the opportunity to mate with a sexually receptive female during this adolescent critical period would have enduring effects on behavior and neuronal morphology into adulthood. A second question was to determine the extent to which testosterone mediated the effects of these social interactions on adult outcomes. Compared to sexually inexperienced hamsters and those that experienced sex for the first time in adulthood, hamsters that experienced adolescent sexual experience displayed increased anxiety- and depressive-like behavioral responses. Adolescent sexual experiences decreased the complexity and length of dendrites on prefrontal cortical neurons and increased the expression of the pro-inflammatory cytokine interleukin 1β (IL-1β) in the PFC. In a second experiment, administration of testosterone during the adolescent period largely recapitulated the effects of adolescent sexual experience. These data support the overall hypothesis that a sensitive period extends into adolescence and that salient social stimuli during this time can significantly and persistently alter adult phenotype.     

Organizational and activational effects of hormones on insect behavior

The concepts of hormone organization and activation provide a framework for thinking about the influence of hormones on development, brain, and behavior in vertebrates. There is good evidence for activational effects of hormones on the nervous system and behavior in insects, but organizational effects are almost never discussed in the insect literature. This paper explores the utility of the concepts of hormonal organization and activation of behavior in insects. We describe the two concepts as developed from studies of vertebrates, review some insect examples that appear to fit this classification scheme, and consider how explicit use of the concept of organization might benefit studies of the insect brain and behavior.     

Androgen Exposure and Reproductive Behavior of an Induced Ovulator, the Pine Vole (Microtus pinetorum)

The actions of steroid hormones on brain and behavior are classically divided into organizational effects that are permanent and occur early in development and activational effects that are temporary and occur throughout life. Here, we test the hypothesis that in an induced ovulator, testosterone defeminizes only those neural tissues that rely on synergistic interactions of estrogen and progesterone for normal function in adulthood. Female voles,Microtus pinetorum,injected with testosterone (T) or oil neonatally were paired with males for an 8-week period. During the pairing, androgenized and oil-treated females spent a similar amount of time investigating the caudal and rostral regions of the males. Males spent significantly less time investigating the caudal and rostral regions of androgenized females. Androgenized females mounted males, did not exhibit lordosis, and were less likely to be mounted by males. Moreover, none of the 10 androgenized females gave birth, whereas 8 of 9 control females gave birth. Androgenized females were also not capable of being stimulated into reproductive condition by males. Injection of 0.5 μg of estradiol benzoate for 4 consecutive days resulted in reduced uterine hypertrophy in androgenized females. These results support the original organizational–activational hypothesis by showing that neonatal androgenization defeminizes and masculinizes female pine voles.     

Behavioral and hormonal correlates of alternative reproductive strategies in a polygynous lizard: Tests of the relative plasticity and challenge hypotheses

Species with alternative reproductive tacts are good models to investigate the poorly understood question of whether individual variation within sexes results from the same physiological mechanisms that control variation between sexes. We have shown previously that adult male tree lizards, Urosaurus ornatus, of different throat color morphs express different levels of aggression in the laboratory. Further field results support the suggestion that the two morphs practice alternative reproductive tactics because the two morphs express different levels of aggressive behavior under field conditions and exhibit dramatic and opposite responses to aggressive challenges. However, despite these behavioral differences, the two morphs do not differ in levels of testosterone or corticosterone either in undisturbed situations or following aggressive challenge. These results are consistent with the relative plasticity hypothesis which proposes that organizational, rather than activational, actions of steroid hormones will be more important in morph differentiation when morphs are fixed in adult life, as they are in tree lizards. These results also support the hypothesis that steroid hormonal levels are insensitive to social modulation in males of species such as U. ornatus without paternal care.     

Gonadal hormones and sex differences in nonreproductive behaviors in rodents: organizational and activational influences

Sex differences in many nonreproductive behaviors have been described in rodents. Among the behaviors that are sexually dimorphic in the rat are activity, aggression, pain, and taste sensitivity, food intake and body weight regulation, the learning and retention of certain kinds of mazes, avoidance responses, taste aversion, and performance on certain schedules of reinforcement. Gonadal hormones seem to be responsible, in part, for sex differences in these behaviors, but their contribution varies greatly with the behavior in question. Frequently, these sexually dimorphic behaviors are influenced both by organizational and activational actions of sex hormones. In other instances (e.g., maze learning and the acquisition of shuttle-box avoidance responses) organizational influences predominate. And while there is no sexually dimorphic behavior surveyed that can be shown to be influenced only by activational effects, wheel-running activity is clearly more strongly subject to activational than to organizational effects of the gonadal hormones. In general, only rudimentary information exists regarding the temporal limits of the period in development when organizational influences on nonsexual behaviors occur. The suggestion can be made that organizational influences often occur outside of the critical period for differentiation of the neuroendocrine system regulating cyclic release of gonadotrophins. Even for behaviors where organizational effects usually occur during a roughly delimited period of development, data for other behavioral systems suggest that the time limits during which organizational effects can occur are not rigidly fixed. Very little information exists regarding biochemical or neural mechanisms by which organizational or activational effects on sexually dimorphic nonreproductive behaviors are expressed. It is important to recognize for many of the sexually dimorphic behaviors in the rat that differences between the sexes are neither large nor absolute. This is especially true of several kinds of learning situations where groups of males and females typically differ in average levels of performance. Ostensibly minor variations in test procedure can abolish or accentuate the average difference in performance between the sexes. We are a long way from an adequate understanding of what factors are important, but such information could be quite helpful in estimating whether sex differences in certain laboratory learning tasks have any adaptive significance.Sex differences in nonreproductive behaviors may be influenced by many factors other than hormonal status. This greatly complicates a comparative analysis, but such an analysis will ultimately be necessary. What limited data exist on rodents suggest that: (1) Sexually dimorphic responses in the rat are often not similarly differentiated in the hamster, the gerbil, or the mouse; and (2) major differences exist among rodent species in hormonal effects on such responses.Over the last decade it has become clear that the behavioral effects of deliberate neurological insult are not necessarily the same in male and female rats (or in one case, in rhesus monkeys). Sex differences in the behavioral effects of ventromedial hypothalamic, lateral hypothalamic, septal, and striatal lesions in the rat and of orbital prefrontal cortex lesions in the monkey have been described. While information regarding hormonal modulation of these differences in response to brain damage is very limited, available data suggest both organizational and activational effects of sex hormones may be involved. It is too early to tell where this line of research may ultimately lead, but rather striking sex differences in the incidence of certain neurological disorders in humans suggest that further research may have both practical and theoretical significance.     

Application of organization-activation theory to alternative male reproductive strategies: a review

Many species have extreme within-sex morphological and behavioral polymorphisms, most commonly different male phenotypes that practice different reproductive strategies. Although much is known about the role of hormones in sexual differentiation, little is known about what role hormones might play in within-sex differentiation. The relative plasticity hypothesis is derived from the classical organization-activation model of hormone action. It distinguishes between two types of polymorphic systems: a fixed system in which individual males assume one phenotype for their adult lives and a plastic system in which individual males can change phenotypes at least once. By analogy to sexual differentiation, the relative plasticity hypothesis generally predicts that organizational influences of hormones will be more important in fixed systems and activational influences of hormones will be more important in plastic systems. A review of our knowledge of the role of hormones in differentiation of within-sex polymorphisms indicates that the relative plasticity hypothesis accounts for otherwise diverse and contradictory results. This further supports the hypothesis that the organizational-activational model of hormone action derived from sexual differentiation generalizes to within-sex polymorphisms. However, studies of the effects of hormone manipulations on within-sex differentiation are rare but are desperately needed to further our understanding of this problem. Further studies of discontinuous behavioral variation characteristic of polymorphic species may further our understanding of the physiological basis of within-sex behavior variation in all species.     

Temperament moderates the influence of periadolescent social experience on behavior and adrenocortical activity in adult male rats

Adolescence is a period of significant behavioral and physiological maturation, particularly related to stress responses. Animal studies that have tested the influence of adolescent social experiences on stress-related behavioral and physiological development have led to complex results. We used a rodent model of neophobia to test the hypothesis that the influence of adolescent social experience on adult behavior and adrenocortical function is modulated by pre-adolescent temperament. Exploratory activity was assessed in 53 male Sprague–Dawley rats to classify temperament and then they were housed in one of the three conditions during postnatal days (PND) 28–46: (1) with familiar kin, (2) with novel social partners, or (3) individually with no social partners. Effects on adult adrenocortical function were evaluated from fecal samples collected while rats were individually-housed and exposed to a 1-hour novel social challenge during PND 110–114. Adolescent-housing with novel or no social partners led to reduced adult glucocorticoid production compared to adolescent-housing with familiar littermates. Additionally, highly-exploratory pre-weanling rats that were housed with novel social partners during adolescence exhibited increased exploratory behavior and a more rapid return to basal glucocorticoid production in adulthood compared to those housed with familiar or no social partners during adolescence and compared to low-exploratory rats exposed to novel social partners. In sum, relatively short-term adolescent social experiences can cause transient changes in temperament and potentially longer-term changes in recovery of glucocorticoid production in response to adult social challenges. Furthermore, early temperament may modulate the influence of adolescent experiences on adult behavioral and adrenocortical function.     

Cognitive effects of variations in pubertal timing: Is puberty a period of brain organization for human sex-typed cognition?

There is considerable interest in the organizational effects of pubertal sex hormones on human sex-related characteristics. Recent evidence from rodents suggests that there is a decreasing window of sensitivity to sex hormones throughout adolescence. If adolescence also represents a period of brain organization in human beings, then the timing of exposure to sex-typical hormones at puberty should have long-term effects on sex-typed characteristics: individuals with early timing should be more sex-typed than individuals with late timing. We tested this hypothesis in 320 young adults by relating their pubertal timing (retrospective comparison to peers) to cognitive abilities that show sex differences. Results provide partial support for the hypothesis. For men, pubertal timing was inversely related to scores on a test of three-dimensional mental rotations. Effects do not appear to be due to duration of hormone exposure (time since puberty), but other potential influences need further study.     

Organizational and activational effects of sex steroids on brain and behavior: A reanalysis

The actions of sex steroids on brain and behavior traditionally have been divided into organizational and activational effects. Organizational effects are permanent and occur early in development; activational effects are transient and occur throughout life. Over the past decade, experimental results have accumulated which do not fit such a simple two-process theory. Specifically, the characteristics said to distinguish organizational and activational effects on behavior are sometimes mixed, as when permanent effects occur in adulthood. Attempts to determine whether specific cellular processes are uniquely associated with either organizational or activational effects are unsuccessful. These considerations blur the organizational-activational distinction sufficiently to suggest that a rigid dichotomy is no longer tenable.     

Sexual differentiation in quail: Conversion of androgen to estrogen mediates testosterone-induced demasculinization of copulation but not other male characteristics

Previous research has shown that administration of either testosterone or estradiol to male quail embryos will demasculinize behavior and morphology. Six experiments in which embryos were treated were conducted to test the hypothesis that this testosterone-induced demasculinization is due to conversion of testosterone to estrogen (aromatization). In Experiment 1, dihydrotestosterone propionate, a nonaromatizable androgen, failed to demasculinize copulatory behavior, but did demasculinize crowing, strutting, and proctodeal glands. In Experiment 2, injection of the aromatizable androgens testosterone propionate (TP), testosterone, or androstenedione demasculinized copulatory behavior, the nonaromatizable androgen androsterone failed to have such an effect, and all androgens demasculinized proctodeal glands. In Experiment 3, Silastic implants of testosterone demasculinized all male characteristics, whereas implants of androsterone demasculinized only proctodeal glands. In Experiment 4, the antiestrogen tamoxifen prevented TP from demasculinizing copulatory behavior, but had no such effect with respect to crowing and strutting. In Experiments 5 and 6, the aromatization inhibitor 1,4,6-androstatrien-3,17-dione (ATD) prevented TP but not estradiol benzoate from demasculinizing copulatory behavior. Thus (1) in quail, testosterone-induced demasculinization of copulatory behavior is due to androgen aromatization, whereas testosterone-induced demasculinization of crowing, strutting, and proctodeal glands is not; (2) the distinct components of normal male reproductive behavior exhibit different patterns of steroid specificity during the organizational period, as was previously shown for the activational period; (3) the steroid specificity of crowing, strutting, and proctodeal glands changes between the organizational and activational periods. During organization, there is little specificity, whereas during activation, these characteristics respond only to androgens, never to estrogens. This difference suggests that developmental changes have occurred in the underlying biochemical substrates.     

Do Sex Differences in the Brain Explain Sex Differences in the Hormonal Induction of Reproductive Behavior? What 25 Years of Research on the Japanese Quail Tells Us

Early workers interested in the mechanisms mediating sex differences in morphology and behavior assumed that differences in behavior that are commonly observed between males and females result from the sex specificity of androgens and estrogens. Androgens were thought to facilitate male-typical traits, and estrogens were thought to facilitate female-typical traits. By the mid-20th century, however, it was apparent that administering androgens to females or estrogens to males was not always effective in sex-reversing behavior and that in some cases a “female” hormone such as an estrogen could produce male-typical behavior and an androgen could induce female-typical behavior. These conceptual difficulties were resolved to a large extent by the seminal paper of C. H. Phoenix, R. W. Goy, A. A. Gerall, and W. C. Young in (1959,Endocrinology65, 369–382) that illustrated that several aspects of sexual behavior are different between males and females because the sexes have been exposed during their perinatal life to a different endocrine milieu that has irreversibly modified their response to steroids in adulthood. Phoenixet al.(1959) therefore formalized a clear dichotomy between the organizational and activational effects of sex steroid hormones. Since this paper, a substantial amount of research has been carried out in an attempt to identify the aspects of brain morphology or neurochemistry that differentiate under the embryonic/neonatal effects of steroids and are responsible for the different behavioral response of males and females to the activation by steroids in adulthood. During the past 25 years, research in behavioral neuroendocrinology has identified many sex differences in brain morphology or neurochemistry; however many of these sex differences disappear when male and female subjects are placed in similar endocrine conditions (e.g., are gonadectomized and treated with the same amount of steroids) so that these differences appear to be of an activational nature and cannot therefore explain sex differences in behavior that are still present in gonadectomized steroid-treated adults. This research has also revealed many aspects of brain morphology and chemistry that are markedly affected by steroids in adulthood and are thought to mediate the activation of behavior at the central level. It has been explicitly, or in some cases, implicitly assumed that the sexual differentiation of brain and behavior driven by early exposure to steroids concerns primarily those neuroanatomical/neurochemical characteristics that are altered by steroids in adulthood and presumably mediate the activation of behavior. Extensive efforts to identify these sexually differentiated brain characteristics over the past 20 years has only met with limited success, however. As regards reproductive behavior, in all model species that have been studied it is still impossible to identify satisfactorily brain characteristics that differentiate under early steroid action and explain the sex differences in behavioral activating effects of steroids. This problem is illustrated by research conducted on Japanese quail (Coturnix japonica), an avian model system that displays prominent sex differences in the sexual behavioral response to testosterone, and in which the endocrine mechanisms that control sexual differentiation of behavior have been clearly identified so that subjects with a fully sex-reversed behavioral phenotype can be easily produced. In this species, studies of sex differences in the neural substrate mediating the action of steroids in the brain, including the activity of the enzymes that metabolize steroids such as aromatase and the distribution of steroid hormone receptors as well as related neurotransmitter systems, did not result in a satisfactory explanation of sex differences in the behavioral effectiveness of testosterone. Possible explanations for the relative failure to identify the organized brain characteristics responsible for behavioral sex differences in the responsiveness to steroids are presented. It is argued that novel research strategies may have to be employed to successfully attack the fundamental question of the hormonal mechanisms regulating sex differences in behavior.     

Adolescents and androgens, receptors and rewards

Adolescence is associated with increases in pleasure-seeking behaviors, which, in turn, are shaped by the pubertal activation of the hypothalamo-pituitary-gonadal axis. In animal models of naturally rewarding behaviors, such as sex, testicular androgens contribute to the development and expression of the behavior in males. To effect behavioral maturation, the brain undergoes significant remodeling during adolescence, and many of the changes are likewise sensitive to androgens, presumably acting through androgen receptors (AR). Given the delicate interaction of gonadal hormones and brain development, it is no surprise that disruption of hormone levels during this sensitive period significantly alters adolescent and adult behaviors. In male hamsters, exposure to testosterone during adolescence is required for normal expression of adult sexual behavior. Males deprived of androgens during puberty display sustained deficits in mating. Conversely, androgens alone are not sufficient to induce mating in prepubertal males, even though brain AR are present before puberty. In this context, wide-spread use of anabolic-androgenic steroids (AAS) during adolescence is a significant concern. AAS abuse has the potential to alter both the timing and the levels of androgens in adolescent males. In hamsters, adolescent AAS exposure increases aggression, and causes lasting changes in neurotransmitter systems. In addition, AAS are themselves reinforcing, as demonstrated by self-administration of testosterone and other AAS. However, recent evidence suggests that the reinforcing effects of androgens may not require classical AR. Therefore, further examination of interactions between androgens and rewarding behaviors in the adolescent brain is required for a better understanding of AAS abuse.     

Hormonal Mechanisms of Mate Choice

SYNOPSIS. Mate choice is a critically important determinantof reproductive success. Because of its significance in theevolutionary process, it has received a great deal of attentionfrom animal behaviorists interested in ultimate causes of behavior.Much less effort has been directed at uncovering the physiologicalmechanisms of mate choice, including those operating duringontogeny that lead to adult mate preferences. As a result ofnatural and sexual selection, many aspects of mate choice aresexually dimorphic. How do adult males and females of the samespecies come to show different mating partner preferences? Onepossibility is that sex steroid hormones play important roles,acting either during early development to permanently establishsex differences or during adulthood to facilitate their expression,roles similar to the organizational and activational effectsof sex steroids on sexually dimorphic copulatory and courtshipbehavior patterns. This review (1) summarizes what is knownabout hormones and mate choice, highlighting those results mostrelevant to understanding proximate causation from an evolutionaryperspective; (2) describes recent work from the author's labtesting an organizational hormone hypothesis of mate choice,focusing on a particularly widespread and robust aspect of matechoice—preference for opposite sex partners—in apair bonding species—the zebra finch; and (3) suggestssome future directions for research that might integrate ultimateand proximate causation.     

The organizational–activational hypothesis as the foundation for a unified theory of sexual differentiation of all mammalian tissues

The 1959 publication of the paper by Phoenix et al. was a major turning point in the study of sexual differentiation of the brain. That study showed that sex differences in behavior, and by extension in the brain, were permanently sexually differentiated by testosterone, a testicular secretion, during an early critical period of development. The study placed the brain together in a class with other major sexually dimorphic tissues (external genitalia and genital tracts), and proposed an integrated hormonal theory of sexual differentiation for all of these non-gonadal tissues. Since 1959, the organizational–activational theory has been amended but survives as a central concept that explains many sex differences in phenotype, in diverse tissues and at all levels of analysis from the molecular to the behavioral. In the last two decades, however, sex differences have been found that are not explained by such gonadal hormonal effects, but rather because of the primary action of genes encoded on the sex chromosomes. To integrate the classic organizational and activational effects with the more recently discovered sex chromosome effects, we propose a unified theory of sexual differentiation that applies to all mammalian tissues.     

Gonadectomy negatively impacts social behavior of adolescent male primates

Social behavior changes dramatically during primate adolescence. However, the extent to which testosterone and other gonadal hormones are necessary for adolescent social behavioral development is unknown. In this study, we determined that gonadectomy significantly impairs social dominance in naturalistic settings and changes reactions to social stimuli in experimental settings. Rhesus macaques were castrated (n =  6) or sham operated (n = 6) at age 2.4 years, group-housed for 2 years, and ethograms were collected weekly. During adolescence the gonadally intact monkeys displayed a decrease in subordinate behaviors and an increase in dominant behaviors, which ultimately related to a rise in social status and rank in the dominance hierarchy. We measured monkey's reactions to emotional faces (fear, threat, neutral) of conspecifics of three ages (adult, peer, infant). Intact monkeys were faster to retrieve a treat in front of a threatening or infant face, while castrated monkeys did not show a differential response to different emotional faces or ages. No group difference in reaction to an innate fear-eliciting object (snake) was found. Approach and proximity responses to familiar vs unfamiliar conspecifics were tested, and intact monkeys spent more time proximal to a novel conspecific as compared to castrates who tended to spend more time with a familiar conspecific. No group differences in time spent with novel or familiar objects were found. Thus, gonadectomy resulted in the emergence of significantly different responses to social stimuli, but not non-social stimuli. Our work suggests that intact gonads, which are needed to produce adolescent increases in circulating testosterone, impact social behavior during adolescences in primates.