Neuropeptides and the evolution of social behavior

Comparative studies over the past year have revealed two new insights into the role of neuropeptides in the evolution of social behaviors. First, across vertebrate taxa, certain neuropeptide effects appear to be gender-specific. Second, species variations in receptor gene structure can alter neuropeptide receptor distribution and thereby contribute to species differences in social behavior.     

Immunocytochemical localization of vasotocin and mesotocin in the hypothalamus of lacertilian reptiles

Summary The hypothalamic magnocellular neurosecretory system of lizards was studied with the unlabeled antibody peroxidase-antiperoxidase complex (PAP) technique at the light microscopic level. It was shown that vasotocin and mesotocin are synthesized in separate neurons. The vasotocinergic as well as the mesotocinergic perikarya are of different sizes. Both cell types occur in close juxtaposition, but without a distinct pattern of distribution. The external zone of the lacertilian median eminence contains numerous immunoreactive vasotocinergic fibers and only few immunoreactive mesotocinergic fibers. The general organization of the hypothalamic magnocellular neurosecretory system of lizards, as revealed by immunocytochemistry, is essentially similar to that revealed with unspecific staining methods.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

Immunocytochemical studies of vasotocin and mesotocin in the hypothalamo-hypophysial system of the chicken

Summary The hypothalamo-hypophysial system of the adult chicken has been studied with a monoclonal antibody that cross-reacts with arginine vasotocin and mesotocin. We have used this antibody on thick (100 m) sections in conjunction with a peroxidase-conjugated rabbit antimouse antibody that permits the visualization not only of entire perikarya, but also of long portions of their axons and dendrites. Our results confirm older concepts based on classical methods, but the more sensitive immunocytochemical method reveals that the system is more extensive than previously recognized. Immunostained neurons in the chicken are widely scattered in the hypothalamus. In the rostral preoptic region, there are three immunostained neuronal cell groups: 1) a prominent closely packed group that extends along the ventromedial surface, 2) a diffusely distributed lateral group, and 3) an external group that surrounds the lateral aspect of the septomesencephalic tract. Caudally in the preoptic area and in the anterior hypothalamus, the same groups are present; but there are also conspicuous periventricular perikarya. Many of them have processes that project to the lumen of the third ventricle, as well as parallel axons that arch lateroventrally in the hypothalamus. In the midhypothalamic area, the periventricular perikarya and processes are particularly numerous at the level of the palliai commissure. The dorsal periventricular group located at the level of the dorsomedial anterior nucleus of the thalamus are the most caudal perikarya. They extend laterally in a wing-like formation. The immunostained axons from all of these perikarya form a compact hypothalamo-hypophysial tract as they run from the midhypothalamus to the median eminence and converge beneath the third ventricle. Axons branching from this tract innervate the zone externa of the anterior median eminence; another group of axons running in the fibrous layer of the zona interna proceeds to the neural lobe.     

The topography of mesotocin and vasotocin systems in the brain of the domestic mallard and japanese quail: Immunocytochemical identification

Summary The neurosecretory systems producing mesotocin (MT) and vasotocin (VT) (the avian homologues of oxytocin and vasopressin, respectively) were characterized in the brains of the domestic mallard and Japanese quail by means of indirect immunofluorescence techniques using specific antisera. In the anterior preoptic region, including the organum vasculosum of the lamina terminalis, and at different levels of the supraoptic and paraventricular nuclei, separate mesotocin- and vasotocin-producing neurons were identified. Mesotocinergic and vasotocinergic neurons were also located in the tuberomammillary area, among the ectomammillary tract fibers. The supraoptico-neurohypophysial tract, formed by vasotocin- and mesotocincontaining axons, enters the internal zone of the median eminence and ends in the posterior lobe of the pituitary. The external zone of the rostral median eminence appears to contain vasotocin and mesotocin fibers, which terminate in close contact with the capillaries of the hypophysial portal system.With contributions by Dr. B. Kerdelhué, Laboratoire des Hormones Polypeptidiques du CNRS, 91190 Gif-sur-Yvette, France     

Hormonal regulation of vasotocin receptor mRNA in a seasonally breeding songbird

Behaviors associated with breeding are seasonally modulated in a variety of species. These changes in behavior are mediated by sex steroids, levels of which likewise vary with season. The effects of androgens on behaviors associated with breeding may in turn be partly mediated by the nonapeptides vasopressin (VP) and oxytocin (OT) in mammals, and vasotocin (VT) in birds. The effects of testosterone (T) on production of these neuropeptides have been well-studied; however, the regulation of VT receptors by T is not well understood. In this study, we investigated steroid-dependent regulation of VT receptor (VTR) mRNA in a seasonally breeding songbird, the white-throated sparrow (Zonotrichia albicollis). We focused on VTR subtypes that have been most strongly implicated in social behavior: V1a and oxytocin-like receptor (OTR). Using in situ hybridization, we show that T-treatment of non-breeding males altered V1a and OTR mRNA expression in several regions associated with seasonal reproductive behaviors. For example, T-treatment increased V1a mRNA expression in the medial preoptic area, bed nucleus of the stria terminalis, and ventromedial hypothalamus. T-treatment also affected both V1a and OTR mRNA expression in nuclei of the song system; some of these effects depended on the presence or absence of a chromosomal rearrangement that affects singing behavior, plasma T, and VT immunolabeling in this species. Overall, our results strengthen evidence that VT helps mediate the behavioral effects of T in songbirds, and suggest that the chromosomal rearrangement in this species may affect the sensitivity of the VT system to seasonal changes in T.     

Vasotocin treatment inhibits courtship in male zebra finches; concomitant androgen treatment inhibits this effect

Zebra finches evolved in arid areas of Australia. Their reproduction is stimulated by water availability, which is unpredictable. Cheng (Poult. Sci. Rev. 5 (1993) 37) hypothesized that the primary mechanism controlling reproduction in species relying on unpredictable cues should be inhibitory. The onset of stimulatory environmental conditions terminates the inhibition, allowing rapid initiation of reproduction. As the primary hormone regulating water balance in birds, arginine vasotocin (AVT) appears a likely candidate to modulate reproduction in finches. Drought conditions cause sustained AVT release, which in other species inhibits androgen production. To determine whether increased AVT inhibits reproductive behavior, intact males were tested with females and divided into three groups matched for courtship behavior. Osmotic minipumps containing (a) saline, (b) 264 ng AVT, or (c) 1320 ng AVT in saline were implanted subcutaneously and males tested 48 h later. AVT-treated males socialized with females, but the high dose significantly reduced singing and courtship displays. To determine whether AVT acted by depressing androgen secretion, additional males were given subcutaneous androgen implants and divided into two groups matched for courtship behavior. Males were then implanted with minipumps containing (a) saline or (b) the high AVT dose. Males treated with AVT plus androgen showed no deficits in courtship behavior. These data suggest that AVT secretion during periods of drought may inhibit reproduction by inhibiting androgen production. Inhibition of reproductive behavior by AVT may be a more general phenomenon. Large quantities of AVT or, in mammals the closely-related peptide vasopressin (VP), are released when animals are stressed, and high levels of AVT/VP may inhibit reproductive behavior. The extremely short half-life of these peptides means that once proximal factors become more favorable, the gonads should rapidly be released from the peptides' inhibitory actions.     

Immunocytochemical study of the development of vasotocin/mesotocin in the hypothalamo-hypophysial system of the chick embryo

Summary The hypothalamo-hypophysial system of the chick embryo has been studied with a monoclonal antibody which cross-reacts with arginine vasotocin and mesotocin, using thick (100 m) sections in conjunction with a peroxidase-conjugated rabbit anti-mouse antibody. Although weakly stained perikarya occur occasionally in the tuberal region on embryonic days 6 and 7, the most consistent immunostaining of perikarya is found in the periventricular region of the caudal midhypothalamus at the level of the optic chiasm after embryonic day 8 1/2. Synthesis of peptides, therefore, takes place while the cells are close to their site of origin. Between embryonic days 9 and 10, beaded axons run along the anterior median eminence closely apposed to the adenohypophysis, thereby forming the anlage of the zona externa. The axons of the hypothalamo-neurohypophysial tract surround the neural lobe between embryonic days 11 and 12. The caudal to rostral wave of neuronal maturation that occurs during development appears to be due to a progressive differentiation of the periventricular zone, as well as the migration of perikarya. The early periventricular perikarya at embryonic day 8 1/2 send processes rostrally in a wing-shaped formation that extends both dorso- and ventrolaterally. From embryonic days 10 to 12, perikarya can be observed in the wing-like extensions, apparently migrating to rostral levels. The dorsolateral pathway gives rise at its midportion to the lateral cell group, whereas those perikarya migrating more laterally form the anlage of the external supraoptic nucleus. The ventrolateral wing-shaped extension of perikarya appears to be directed toward the ventral group and those lateral perikarya continuous with it. The location of mature neuronal cell groups is well established by embryonic day 17.     

Sex-specific activity and function of hypothalamic nonapeptide neurons during nest-building in zebra finches

Vertebrate species from fish to humans engage in a complex set of preparatory behaviors referred to as nesting; yet despite its phylogenetic ubiquity, the physiological and neural mechanisms that underlie nesting are not well known. We here test the hypothesis that nesting behavior is influenced by the vasopressin–oxytocin (VP–OT) peptides, based upon the roles they play in parental behavior in mammals. We quantified nesting behavior in male and female zebra finches following both peripheral and central administrations of OT and V_1a receptor (OTR and V1aR, respectively) antagonists. Peripheral injections of the OTR antagonist profoundly reduce nesting behavior in females, but not males, whereas comparable injections of V1aR antagonist produce relatively modest effects in both sexes. However, central antagonist infusions produce no effects on nesting, and OTR antagonist injections into the breast produce significantly weaker effects than those into the inguinal area, suggesting that antagonist effects are mediated peripherally, likely via the oviduct. Finally, immunocytochemistry was used to quantify nesting-induced Fos activation of nonapeptide neurons in the paraventricular and supraoptic nuclei of the hypothalamus and the medial bed nucleus of the stria terminalis. Nest-building induced Fos expression within paraventricular VP neurons of females but not males. Because the avian forms of OT (Ile⁸-OT; mesotocin) and VP (Ile³-VP; vasotocin) exhibit high affinity for the avian OTR, and because both peptide forms modulate uterine contractility, we hypothesize that nesting-related stimuli induce peptide release from paraventricular vasotocin neurons, which then promote female nesting via peripheral feedback from OTR binding in the oviduct uterus.     

Hormonal regulation of agonistic and affiliative behavior in female mongolian gerbils (Meriones unguiculatus)

Ovarian steroids and oxytocin (OT) have been implicated in the regulation of social behaviors. The purpose of the present study was to examine hormonal substrates of aggression and affiliation in the female Mongolian gerbil (Meriones unguiculatus), a highly social, monogamous rodent. Sexually naive adult females were paired with sexually experienced males for 48 h and their interactions videotaped. Females were gonadally intact and tested during vaginal estrus (INT) or ovariectomized and observed after the following treatments, administered by means of sc injections: EBEB (7 days of estradiol-benzoate); EBP (2 days of EB followed by progesterone), SALEB (saline, days 1-5 then 2 days of EB), OTEB (OT for days 1-5 then 2 days of EB); OTOIL (OT for days 1-5 then 2 days of OIL); or SALOIL (saline days 1-5 then 2 days of OIL). During the first hour of pairing INT females displayed higher levels of affiliation and lower levels of sniffing and agonistic behavior than SALOIL females. All hormonal treatments reduced agonistic behaviors when compared to SALOIL, although none of the hormonal treatments restored affiliation to INT levels. During the 48-h test overt aggression varied by treatment with INT, EBEB, EBP, and OTEB females displaying lower levels than SALOIL, while all groups displayed similar levels of affiliation. The results indicate that OT and E play a significant role in regulating male-directed aggressive behavior in females and that the presence of ovarian hormones as well as OT can increase affiliation during initial contact. Over a sustained period of cohabitation social cues appear to be more important in regulating affiliation than gonadal hormones.     

The effects of sex steroids and vasotocin on behavioral responses to visual and olfactory sexual stimuli in ovariectomized female roughskin newts

Previous studies have found that vasotocin (AVT) administration to male roughskin newts (Taricha granulosa) enhances courtship clasping as well as appetitive responses to specific sexual stimuli and that treating female newts with androgens plus AVT induces the expression of male-typical courtship clasping (the selective clasping of females). However, the unique and/or interactive effects of sex steroids and AVT on appetitive responses to specific sexual stimuli have not yet been determined. To first identify male-typical, sexually dimorphic appetitive responses to female sexual stimuli, we tested intact newts during the breeding season and found that males, but not females, are attracted to female visual and pheromonal sexual stimuli. We then used ovariectomized (ovx) females implanted with empty silastic capsules (Blk) or with capsules containing testosterone (T), dihydrotestosterone (DHT), or estradiol (E2) and then injected with either saline or AVT to determine the effects of steroids and AVT, alone or in combination with each other, on male-typical behavioral responses to those stimuli. E2 treatment depressed responses toward female visual stimuli independently of AVT. On the other hand, only T-implanted, AVT-injected females displayed male-typical behavioral responses toward female olfactory stimuli, preferring to spend more time in proximity to female-scented than unscented newt models and selectively clasping the female-scented models. Together, these results support the conclusion that sex steroids and AVT influence behavioral responses to sexual stimuli via sensory-specific mechanisms. Furthermore, they suggest that T and AVT interact within the brain to influence sensorimotor processing in the pathways that integrate olfactory sexual stimuli into male-typical courtship behaviors.     

Tactile contact is required for early estrogen treatment to alter the sexual partner preference of female zebra finches

Sex steroid actions during early development appear to play a role in the development of sex differences in sexual partner preference (SPP, preference for males vs. females) in several species of mammals and in the socially monogamous pair bonding zebra finch (Taeniopygia guttata). Female finches treated with estrogen as nestlings exhibit varying degrees of masculinized SPP as adults, but only if they have been housed in all-female groups during the juvenile and young adult period, suggesting that the estrogen effect may involve social experience and possibly sexual imprinting. Because tactile contact is important for consolidation of imprinted preferences in this species, it was predicted that early estrogen treatment would alter preferences of females only if they were allowed to have tactile contact with other females. Subjects were injected with estradiol benzoate or with oil (normal controls) daily for the first 2 weeks post-hatching. At age 45 days, they were housed in a mixed sex aviary (normal controls), in an all-female aviary allowing tactile contact (group EB-TC), or in an all-female aviary with no tactile contact (group EB-NTC). At 100+ days, birds were given two-choice SPP tests followed by aviary tests of SPP. EB-TC females did not show the sex-typical preference for male stimuli, and differed significantly from the controls on several measures. EB-NTC females preferred males and never differed significantly from controls. These results show that tactile contact after age 45 days is essential for an EB effect on SPP, supporting the hypothesis that hormones and sexual imprinting together contribute to SPP.     

Both oxytocin and vasopressin may influence alloparental behavior in male prairie voles

Neuropeptides, especially oxytocin (OT) and arginine vasopressin (AVP), have been implicated in several features of monogamy including alloparenting. The purpose of the present study was to examine the role of OT and AVP in alloparental behavior in reproductively na?ve male prairie voles. Males received intracerebroventricular (ICV) injections of artificial cerebrospinal fluid (aCSF), OT, an OT receptor antagonist (OTA), AVP, an AVP receptor antagonist (AVPA), or combinations of OTA and AVPA and were subsequently tested for parental behavior. Approximately 45 min after treatment, animals were tested for behavioral responses to stimulus pups. In a 10-min test, spontaneous alloparental behavior was high in control animals. OT and AVP did not significantly increase the number of males that showed parental behavior, although more subtle behavioral changes were observed. Combined treatment with AVPA and OTA (10 ng each) significantly reduced male parental behavior and increased attacks; following a lower dose (1 ng OTA/1 ng AVPA), males were less likely to display kyphosis and tended to be slower to approach pups than controls. Since treatment with only one antagonist did not interfere with the expression of alloparenting, these results suggest that access to either OT or AVP receptors may be sufficient for the expression of alloparenting.     

Arginine vasotocin induces sexual behavior of newts by acting on cells in the brain

To investigate whether arginine vasotocin (AVT) acts on target cells in the brain of Taricha granulosa (a urodele amphibian), the behavioral effects of intracerebroventricular (ICV) and intraperitoneal (IP) injections of AVT were compared. Male newts exhibited the greatest sexual activity (amplectic clasping) following an ICV injection of 0.1 μg AVT. Another study showed that nanogram quantities of AVT, administered ICV, stimulated the behavior. An ICV injection of an antagonist to arginine vasopressin, d(CH₂)₅Tyr(Me)AVP, or an anti-AVT immune serum significantly inhibited the sexual behavior. Intracranial implants of 17β-estradiol (E₂) or 5α-dihydrotestosterone (DHT) in castrated males maintained the behavioral response to an injection of AVT. Another study found that an IP injection of DHT or E₂ did not increase the incidence of newt sexual behavior during the 8 hours following the injection.     

Effects of housing condition and early corticosterone treatment on learned features of song in adult male zebra finches

Early developmental stress can have long-term physiological and behavioral effects on an animal. Developmental stress and early corticosterone (Cort) exposure affect song quality in many songbirds. Early housing condition can act as a stressor and affect the growth of nestlings and adult song, and improvements in housing condition can reverse adverse effects of early stress exposure in rodents. However, little is known about this effect in songbirds. Therefore, we took a novel approach to investigate if housing condition can modify the effects of early Cort exposure on adult song in male zebra finches. We manipulated early housing conditions to include breeding in large communal flight cages (FC; standard housing condition; with mixed-sex and mix-aged birds) versus individual breeding cages (IBC, one male–female pair with small, IBC-S, or large clutches, IBC-L) in post-hatch Cort treated male birds. We found that Cort treated birds from IBC-S have higher overall song learning scores (between tutor and pupil) than from FC but there is no difference between these groups in the No-Cort treated birds. When examining the effects of Cort within each housing condition, overall song learning scores decreased in Cort treated birds from flight cages but increased in birds from IBC-S compared to controls. Likewise, the total number of syllables and syllable types increased significantly in Cort treated birds from IBC-S, but decreased in FC-reared birds though this effect was not statistically significant. These findings suggest that the effects of early Cort treatment on learned features of song depend on housing condition.     

雌雄子午沙鼠的认知行为及其神经内分泌水平的差异

本研究通过对雌雄子午沙鼠进行新物体识别和社会认知实验,运用免疫组化方法检测其相关脑区合成催产素（OT）、加压素（AVP）和多巴胺（DA）能的神经元数量,采用酶联免疫试验（ELISA）方法检测了其血清中OT、AVP的水平,探究了雌雄子午沙鼠的两性认知差异及其神经内分泌水平的差异。结果表明,雌雄子午沙鼠对新物体的探究时间均要显著高于旧物体,雌雄子午沙鼠的辨别指数无显著差异（P＞0.05）;雄性子午沙鼠随着探究次数的增加对重复刺激鼠a的探究时间不断减少,对陌生刺激鼠b的探究时间显著高于刺激鼠a（P＜0.05）;雌性子午沙鼠没有此趋势。雄性子午沙鼠OT能神经元数量在下丘脑室旁核(PVN)和视上核(SON)均要显著少于雌性（P＜0.05）;雄性个体DA能神经元数量在黑质显著高于雌性（P＜0.01）;然而雄性个体DA能神经元数量在腹侧被盖区显著少于雌性（P＜0.01）;雌雄子午沙鼠血清OT、AVP水平均无显著差异。综上所述,雌雄子午沙鼠对新物体的识别能力无显著差异,然而雄性子午沙鼠的社会认知能力强于雌性。在神经内分泌水平上,雌雄子午沙鼠PVN和SON中OT能神经元数量、黑质和腹侧背盖区的DA能神经元数量均呈现出了两性差异。     

Hormonal specificity and activation of sexual behavior in male zebra finches

Castrated zebra finches receiving one of six hormone treatments were given three weekly tests with different females and their sexual behavior was contrasted with that of two control groups consisting of intact or castrated males given implants of cholesterol. The six hormone treatments were: two aromatizable androgens, testosterone (T) and androstenedione (AE); two nonaromatizable androgens, androsterone (AN) and dihydrotestosterone (DHT); an estrogen, estradiol (E); or a combination of E + DHT. Half the males receiving DHT received the 5α-isomer, half received the 5β-isomer. Castration significantly reduced the proportion of males which courted females, total courtship displays, high-intensity courtship displays, beak wiping activity, and significantly increased the latencies to show these behaviors compared to intact males. Castrated males never attempted to mount a female. All of these measures of courtship and copulatory behavior were restored to normal levels only by treatments providing both estrogenic and α-androgenic metabolites (i.e., T, AE, E + αDHT). AE was clearly the most effective of these, raising behavior significantly above normal on several measures. AN treatment was more effective than αDHT on all measures and not significantly different from intact birds on some. Treatment with E, αDHT, βDHT, or E + βDHT was totally ineffective. Surprisingly, females only solicited males whose hormone treatments provided estrogenic metabolites. Not only did they solicit males given aromatizable androgens, which showed high rates of courtship activity, they also solicited males given E or E + βDHT, some of which never even courted. Castration and hormone treatment also affected body and syringeal weight, but in opposite directions. Castration increased body weight while decreasing syringeal weight. Hormone treatments providing α-androgenic metabolites decreased body weight and increased syrinx weight. Treatments supplying estrogen as well were slightly more effective.     

Sexual behavior of female rats in a multiple-partner preference test

In this study, sexually experienced female rats were tested in a multiple-partner preference test (MPPT) in which they were allowed to pace their sexual contacts with four sexually active males. Four cylinders, with a small hole through which only the female could move freely from one cylinder to another, were assembled forming in the center an empty compartment. An intact female was placed in the central compartment and a sexually active male in each cylinder. Female sexual behavior was analyzed throughout the estrus cycle in four consecutive days. Each daily test lasted 15 min. The percentage of exits after intromission or ejaculation was significantly higher than the percentage of exits after each mount. The female spent significantly longer time with one of the males. We designated this male as the preferred male (PM). Although in each of the 4 days studied, females spent significantly longer time with the PM, however, the male selected was not the same throughout the estrus cycle. The number of entries into the compartment of the PM was significantly higher and increased around proestrus. Compared to previous studies, pacing behavior was notably lower in the conditions of the MPPT. No significant differences were observed during the estrous cycle concerning the other parameters recorded. The present results suggest that the MPPT could be a good model to study partner preference in the female rat.     

An experimental test of condition-dependent male and female mate choice in zebra finches

In mating systems with social monogamy and obligatory bi-parental care, such as found in many songbird species, male and female fitness depends on the combined parental investment. Hence, both sexes should gain from choosing mates in high rather than low condition. However, theory also predicts that an individual's phenotypic quality can constrain choice, if low condition individuals cannot afford prolonged search efforts and/or face higher risk of rejection. In systems with mutual mate choice, the interaction between male and female condition should thus be a better predictor of choice than either factor in isolation. To address this prediction experimentally, we manipulated male and female condition and subsequently tested male and female mating preferences in zebra finches Taeniopygia guttata, a songbird species with mutual mate choice and obligatory bi-parental care. We experimentally altered phenotypic quality by manipulating the brood size in which the birds were reared. Patterns of association for high- or low-condition individuals of the opposite sex differed for male and female focal birds when tested in an 8-way choice arena. Females showed repeatable condition-assortative preferences for males matching their own rearing background. Male preferences were also repeatable, but not predicted by their own or females' rearing background. In combination with a brief review of the literature on condition-dependent mate choice in the zebra finch we discuss whether the observed sex differences and between-studies differences arise because males and females differ in context sensitivity (e.g. male-male competition suppressing male mating preferences), sampling strategies or susceptibility to rearing conditions (e.g. sex-specific effect on physiology). While a picture emerges that juvenile and current state indeed affect preferences, the development and context-dependency of mutual state-dependent mate choice warrants further study.