Oxytocin and vasopressin receptors in human and uterine myomas during menstrual cycle and early pregnancy

The purpose of this study was to determine the specificity and concentration of oxytocin (OT) and arginine vasopressin (AVP) binding sites in non-pregnant (NP) human and rhesus monkey endometrium, myometrium and fibromyomas, and to determine the cellular localization of OT receptor (OTR). Besides [3H]AVP, [125I]LVA, a specific VP1 receptor subtype antagonist, was used to determine vasopressin receptor (VPR) concentrations. Samples were obtained from 42 pre-menopausal and three pregnant women (5, 13 and 35 weeks gestation), and several NP and pregnant monkeys. Specificity of binding was assessed in competition experiments with unlabelled agonists and antagonists of known pharmacological potency. Cellular localization of OTR was determined by immunohistochemistry. In NP human uterine tissues, [3H]AVP was bound with higher affinity and greater binding capacity than [3H]OT, whereas in pregnant women and in NP and pregnant rhesus monkeys, uterine OT binding capacity was greater. OT and AVP binding sites discriminated very poorly between OT and AVP; [125I]LVA binding sites were more selective than [3H]AVP. Their ligand specificity and binding kinetics indicated the presence of two distinct populations of binding sites for OT and AVP in primate uterus. Endometrium of NP women and monkeys had low OTR and VPR concentrations. Myometrial and endometrial OTR and VPR were down-regulated in midcycle and in early human pregnancy, they were up-regulated in the secretory phase and second half of pregnancy. Immunoreactive OTR in NP uterus was localized in patches of myometrial muscle cells and small numbers of endometrial epithelial cells.     

Expression and regulation of endothelin-1 and its receptors in human penile smooth muscle cells

We report for the first time that penile smooth muscle cells (SMC) not only respond to, but also synthesize, endothelin-1 (ET-1), one of the main regulators of SMC activity. Immunohistochemical studies indicated that, beside endothelial cells (EC), SMC of the human adult and fetal penis also express ET-1 and its converting enzyme, ECE-1. Accordingly, cultures of adult penile stromal cells express these genes. We also prepared and characterized penile SMC from human fetuses. These cells express SMC specific markers such as alpha smooth muscle actin and phosphodiesterase type 5A3 along with hallmarks of androgen-dependent cells (androgen receptor and 5alpha reductase type 2). Human fetal penile SMC (hfPSMC) are immunopositive for ET-1 and release ET-1. ET-1 expression in hfPSMC was strongly increased by several factors such as transforming growth factor-beta1 (TGF-beta1), interleukin-1alpha (IL-1alpha), ET-1 itself and prolonged (24 h) hypoxia. This latter condition not only affected ET-1 expression but also responsiveness. While at normal oxygen tension, hfPSMC responded to ET-1 with a decreased proliferation mediated by the endothelin-A receptors and TGF-beta1; however, during hypoxia, ET-1 stimulated cell growth. Accordingly, prolonged hypoxia up-regulated endothelin-B receptor mRNA expression. In conclusion, our results indicate that in penile tissues SMC produce ET-1 and that such production is modulated by factors involved in penile physiology and tissue remodelling. In addition, the hfPSMC we have characterized might be a useful model for studying biochemical aspects of the human erectile process in vitro.     

Distribution of vasopressin and oxytocin binding sites in the brain and upper spinal cord of the common marmoset

The aim of this study was to label selectively and to map central vasopressin (AVP) and oxytocin (OT) binding sites in the common marmoset. [¹²⁵I]VPA, a compound selective in rodents and human for the AVP V_1a receptor, yielded the same labeling pattern as [³H]AVP, thus suggesting that most AVP receptors present in the marmoset brain are of the V_1a subtype. Numerous areas exhibited AVP binding sites, among which the olfactory bulb, the accumbens nucleus, the bed nucleus of the stria terminalis, the hypothalamic suprachiasmatic, arcuate and ventromedial nuclei, the medial amygdaloid nucleus, the nucleus of the solitary tract and the cerebral cortex. Binding sites for [¹²⁵I]OTA, a selective OT receptor antagonist in rat and human, were markedly less abundant than [¹²⁵I]VPA ones, and, to a few exceptions, expressed in different areas. Neither AVP, nor OT binding sites were detected in the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei identified by neurophysin immunoreactivity. Marked species-related differences were observed in the distribution of both AVP and OT binding sites. Altogether, our data provide a morphological basis to investigate the function of central AVP and OT in the marmoset.     

Neonatal oxytocin manipulations have long-lasting, sexually dimorphic effects on vasopressin receptors

Developmental exposure to oxytocin (OT) or oxytocin antagonists (OTAs) has been shown to cause long-lasting and often sexually dimorphic effects on social behaviors in prairie voles (Microtus ochrogaster). Because regulation of social behavior in monogamous mammals involves central receptors for OT, arginine vasopressin (AVP), and dopamine, we examined the hypothesis that the long-lasting, developmental effects of exposure to neonatal OT or OTA might reflect changes in the expression of receptors for these peptides. On postnatal day 1, prairie voles were injected intraperitoneally with either OT (1 mg/kg), an OTA (0.1 mg/kg), saline vehicle, or were handled only. At approximately 60 days of age, vasopressin V1a receptors, OT receptors (OTR) and dopamine D2 receptor binding were quantified using receptor autoradiography in brain tissue taken from males and females. Significant treatment effects on V1a binding were found in the bed nucleus of the stria terminalis (BNST), cingulate cortex (CgCtx), mediodorsal thalamus (MdThal), medial preoptic area of the hypothalamus (MPOA), and lateral septum (LS). The CgCtx, MPOA, ventral pallidum, and LS also showed significant sex by treatment interactions on V1a binding. No significant treatment or sex differences were observed for D2 receptor binding. No significant treatment difference was observed for OTR receptor binding, and only a marginal sex difference. Changes in the neuropeptide receptor expression, especially the V1a receptor, may help to explain sexually dimorphic changes in behavior that follow comparable neonatal manipulations.     

Effects of hypoxia on endothelin-1 sensitivity in the corpus cavernosum

The penis remains in a hypo-oxygenated, flaccid state for a large majority of the time. In this study, we investigated the effect of changing oxygen tension on the expression and functional activity of endothelin-1 (ET-1) receptors in the penis. Experiments were performed in rabbit and human corpora cavernosa (CC) as well as in human fetal penile tissue and cell cultures [human fetal penile endothelial cells (hfPECs) and human fetal smooth muscle cells (hfPSMCs)]. Endothelin A (ETA) receptors are expressed by both endothelial and muscular cells in all tissues investigated. Only penile endothelial cells express endothelin B (ETB) receptors, which are further turned on during experimental hypoxia. In addition, hypoxia also allows ETB expression in the muscular compartment without affecting ETA expression. This hypoxia-induced over-expression of ETB decreased the contractile activity of ET-1 and increased ETB-mediated relaxation. The latter was essentially related to increased ETB-mediated nitric oxide formation in hfPEC and even in hfPSMC. Hypoxia also induced a time-dependent down-regulation of RhoA and Rho kinase (ROK) expression which, in turn, participated in the decreased contractile activity of ET-1 in the hypoxic penile tissue. Accordingly, during hypoxia, an ROK inhibitor, Y27632, was less effective in relaxing ET-1-precontracted strips. In conclusion, prolonged (24 h) hypoxia stimulated several counter-regulatory mechanisms in penile tissue, including up-regulation of ETB and down-regulation of RhoA/ROK pathways, which may help to preserve CC hypo-oxygenation, allowing smooth muscle relaxation and, most probably, penile erection.     

The comparative distribution of forebrain receptors for neurohypophyseal peptides in monogamous and polygamous mice.

Several recent studies have suggested that the neurohypophyseal peptide oxytocin may have a role within the brain to mediate various forms of affiliative behavior. As the regulation of oxytocin function may be largely determined by the number and distribution of its membrane bound receptor, we investigated oxytocin receptor distribution in two Peromyscus species selected for differences in affiliative behavior. Using in vitro receptor autoradiography with the selective oxytocin receptor ligand [125I]d(CH2)5[Tyr(Me)2,Tyr-NH9(2)]OVT ([125I]OTA), we compared Peromyscus maniculatus, a polygamous species, to Peromyscus californicus, a monogamous species. Marked species differences in the distribution of [125I]OTA were apparent in several brain areas, including olfactory pathways, bed nucleus of the stria terminalis, amygdala, dorsal lateral septum, and several cortical regions. In addition, gender differences in the binding pattern were evident in several regions, mostly due to sexually dimorphic patterns in the polygamous species, P. maniculatus. To further compare these species, the binding of a [3H]arginine-vasopressin antagonist was assessed in alternate sections from those used for [125I]OTA. Relative to oxytocin receptors, binding to arginine-vasopressin receptors showed fewer species differences, although the monogamous species appeared to have more arginine-vasopressin receptors in the neocortex and lateral septum. The striking differences in oxytocin receptor distribution are consistent with earlier studies in other rodents, suggesting that oxytocin may have an important role for mediating species-typical patterns of social affiliation.     

Oxytocin and same-sex social behavior in female meadow voles

The neuropeptide oxytocin (OT) has been implicated in a range of mammalian reproductive and social behaviors including parent-offspring bonding and partner preference formation between socially monogamous mates. Its role in mediating non-reproductive social relationships in rodents, however, remains largely unexplored. We examined whether OT facilitates same-sex social preferences between female meadow voles—a species that forms social nesting groups in short, winter-like day lengths. In contrast to results from studies of opposite-sex attachment between prairie vole mates, we found that neither OT nor dopamine neurotransmission was required for baseline levels of social partner preference formation or expression. OT enhanced preference formation beyond baseline levels—an effect that was counteracted by treatment with an oxytocin receptor antagonist (OTA). Oxytocin receptor (OTR) density correlated with social behavior in brain regions not known to be associated with opposite-sex affiliation, including the lateral septum and central amygdala. In addition, voles housed in short day lengths (SD) exhibited higher levels of OTR binding in the central amygdala, and voles exposed to high concentrations of estradiol exhibited less binding in the nucleus accumbens (NAcc) and increased binding in the ventromedial nucleus of the hypothalamus. These results suggest that same-sex social behavior shares common elements with other mammalian social behaviors affected by OT, but that the specific neural pathways through which OT exerts its influence are likely distinct from those known for sexual attachments.     

Desensitization of oxytocin receptors in human myometrium

In the present study, we investigated the possible mechanisms by which oxytocin might regulate oxytocin receptor (OTR) density. Exposure of cultured myometrial cells to oxytocin for a prolonged period caused desensitization: the steady-state level of oxytocin binding was 210 x 10(3) binding sites/cell, but this was time-dependently reduced to 20.1 x 10(3) sites/cell by exposing the cells to oxytocin for up to 20 h. In contrast, Western blotting data showed that the total amount of OTR protein was not affected by oxytocin treatment for up to 24 h. Flow cytometry experiments demonstrated that OTRs were not internalized during this treatment. However, RNase protection assays and Northern analysis showed that in cultured myometrial cells OTR mRNA was reduced by oxytocin treatment to reach a new low steady-state concentration. Analysis of this mRNA in myometrial biopsies from 17 patients undergoing emergency Caesarean section showed how it decreased with advancing labour. Samples obtained after 12 h of labour contained approximately 50 times less OTR mRNA than samples obtained from patients in labour for less than 12 h. We speculate that this decrease in OTR mRNA represents in-vivo OTR desensitization.     

Oxytocin mediates the antidepressant effects of mating behavior in male mice

A significant association between plasma oxytocin (OT) levels and depression has been demonstrated. A recent study found that sexual activity and mating with a female induced the release of OT in the central nervous system of male rats. Here we examined the effect of mating behavior on depression-related behavior in wild-type (WT) and OT receptor-deficient (OTR KO) male mice. The WT males showed a reduction in depression-related behavior after mating behavior, but the OTR KO mice did not. Application of an OTR antagonist inhibited mating behavior-induced antidepressant effect in WT males. OT may mediate the antidepressant effects of mating behavior.     

Developmental exposure to oxytocin facilitates partner preferences in male prairie voles (Microtus ochrogaster)

The authors investigated the effects of postnatal manipulations of oxytocin (OT) on the subsequent tendency to form a partner preference in male prairie voles (Microtus ochrogaster). Neonatally, males received either an injection of OT, an oxytocin antagonist (OTA), 0.9% saline vehicle, or handling without injection. As adults, males were tested for partner preference following 1 hr of cohabitation with a nonestrous female. In a 3-hr preference test, males neonatally exposed to exogenous OT exhibited a significant partner preference, not seen in males receiving OTA or saline. Both OT and OTA voles had significantly higher levels of social contact than saline controls. A single neonatal injection of OT increased both total and selective social behaviors in male prairie voles.     

Increased oxytocinergic system activity in the cardiac muscle in spontaneously hypertensive SHR rats

IntroductionThe present study aimed to determine whether the presence of cardiac hypertrophy due to arterial hypertension is associated with a change in the activity of the oxytocinergic system in cardiomyocytes.Material and methodsThe experiments were performed on male, spontaneously hypertensive rats (SHR, n = 10) and normotensive Wistar-Kyoto rats (WKY, n = 12). Blood samples were collected from both SHR and WKY animals to asses plasma oxytocin (OT) concentration; the rats were sacrificed by decapitation. Samples of the left and right ventricles were harvested for the analysis of the OT and oxytocin receptor (OTR) protein by ELISA, and OT and OTR mRNA expression by RT-PCR. Immunohistopathological studies were performed to confirm the presence of OTR receptors in the cardiac muscle of the ventricles.ResultsPlasma OT concentration did not differ between SHR and WKY rats. In the SHR rats, the expression of OT mRNA and the OT protein level was higher in the left and the right ventricle, while OTR mRNA expression was significantly lower in both the left and the right ventricle. However, the level of OTR protein was higher only in the left ventricle of the SHR rats. The presence of OTR receptors was confirmed by immunohistochemical analysis in the muscle of the right and left ventricle.ConclusionsThe presence of arterial hypertension is associated with increased activity of the oxytocinergic system in the heart, especially in the area of the left ventricle. These findings support the important role of this system in the maintenance of cardiovascular homeostasis.     

Blocking oxytocin receptors inhibits vaginal marking to male odors in female Syrian hamsters

In Syrian hamsters (Mesocricetus auratus), precopulatory behaviors such as vaginal scent marking are essential for attracting a suitable mate. Vaginal marking is dependent on forebrain areas implicated in the neural regulation of reproductive behaviors in rodents, including the medial preoptic/anterior hypothalamus (MPOA-AH). Within MPOA-AH, the neuropeptide oxytocin (OT) acts to facilitate copulation (lordosis), as well as ultrasonic vocalizations towards males. It is not known, however, if OT in this area also facilitates vaginal marking. In the present study, a specific oxytocin receptor antagonist (OTA) was injected into MPOA-AH of intact female Syrian hamsters to determine if oxytocin receptor-dependent signaling is critical for the normal expression of vaginal marking elicited by male, female, and clean odors. OTA injections significantly inhibited vaginal marking in response to male odors compared with vehicle injections. There was no effect of OTA on marking in response to either female or clean odors. When injected into the bed nucleus of the stria terminalis (BNST), a nearby region to MPOA-AH, OTA was equally effective in decreasing marking. Finally, the effects of OTA appear to be specific to vaginal marking, as OTA injections in MPOA-AH or BNST did not alter general locomotor activity, flank marking, or social odor investigation. Considered together, these results suggest that OT in MPOA-AH and/or BNST normally facilitates male odor-induced vaginal marking, providing further evidence that OT generally supports prosocial interactions among conspecifics.     

The effects of oxytocin and vasopressin on partner preferences in male and female prairie voles (Microtus ochrogaster)

This study compared the effects of centrally administered oxytocin (OT) and arginine vasopressin (AVP) on partner preference formation and social contact in male and female prairie voles (Microtus ochrogaster). After 1 hr of cohabitation and pretreatment with either AVP or OT, both males and females exhibited increased social contact and significant preference for the familiar partner. After pretreatment with either an OT receptor antagonist (OTA) or an AVP (V1a) receptor antagonist (AVPA), neither OT nor AVP induced a partner preference. In addition, treatment with OT+OTA or AVP+AVPA was associated with low levels of social contact in both sexes. Either AVP or OT is sufficient to facilitate social contact if either the OT or AVP receptor is available. However, the formation of partner preferences may require access to both AVP and OT receptors.     

Mapping oxytocin receptor gene expression in the mouse brain and mammary gland using an oxytocin receptor-LacZ reporter mouse

The hypothalamic nonapeptide oxytocin (OT) has an established role as a circulating hormone but can also act as a neurotransmitter and as a neuromodulator by interacting with its central OT receptor (OTR). To understand the role of the OTR in the mouse brain we investigated the expression of the OTR gene at the cellular level. We targeted the lacZ reporter gene to the OTR gene locus downstream of the endogenous OTR regulatory elements. Using lactating mouse mammary gland as a control for OTR promoter directed specificity of lacZ gene expression, X-gal histochemistry on tissue sections confirmed that gene expression was restricted to the myoepithelial cells. We also identified for the first time in mice the expression of the OTR gene in neighbouring adipocytes. Further, investigation in the mouse brain identified numerous nuclei containing neurons expressing the OTR gene. Whilst some of these regions had been described for rat or sheep, the OTR-LacZ reporter mouse enabled the identification of novel sites of central OTR gene expression. These regions include the accessory olfactory bulb, the medial septal nucleus, the posterolateral cortical amygdala nucleus, the posterior aspect of the basomedial amygdala nucleus, the medial part of the supramammillary nucleus, the dorsotuberomammillary nucleus, the medial and lateral entorhinal cortices, as well as specific dorsal tegmental, vestibular, spinal trigeminal, and solitary tract subnuclei. By mapping the distribution of OTR gene expression, depicted through histochemical detection of beta-galactosidase, we were able to identify single OTR gene expressing neurons and small neuron clusters that would have remained undetected by conventional approaches.These novel sites of OTR gene expression suggest additional functions of the oxytocinergic system in the mouse. These results lay the foundation for future investigation into the neural role of the OTR and provide a useful model for further study of oxytocin functions in the mouse.     

Developmental effects of oxytocin on stress response: single versus repeated exposure

Both exogenous and endogenous oxytocin (OT) are associated with an attenuated stress response. Increased levels of OT in the early postnatal period have been shown to affect behavior and physiology in rodents, and these effects last into adulthood, suggesting an organizational role for OT during development. We investigated the effects of neonatal exposure to OT on the development of the stress response in male and female prairie voles (Microtus ochrogaster). OT or an OT antagonist (OTA) was administered either on postnatal day 1 (single, D1) or days 1-7 (repeated, D1-7) and then on day 8 the response to social isolation was assessed by quantifying ultrasonic vocalizations (USVs) and measuring plasma corticosterone (CORT). Treatment effects were observed only in females. A single treatment with OTA was associated with a decrease in vocalizations, while repeated treatment produced an increase in vocalizations. A single treatment with either saline or OTA increased basal CORT. The results suggest that endogenous OT may be involved in the development of the stress response in females.     

Could oxytocin administration during labor contribute to autism and related behavioral disorders?--A look at the literature

This literature review summarizes recent potential evidence, most of which is at the molecular/mechanistic level, in support of Hollander's hypothesis that excess oxytocin (OT), possibly through OT administration at birth, could contribute to the development of autistic spectrum disorders and related syndromes by proposed down regulation of the OT receptor (OTR). In this review, recent molecular evidence for OTR internalization by excess OT is related to OT's reported effects on animal social behavior, favoring social bondage, notably in sheep, voles, rats and especially mice. Adding indications for OT's capability of crossing the maternal placenta and OT's possibility of crossing an underdeveloped or stressed infantile blood brain barrier at birth, a causal connection between OT excess and behavioral disorders such as autism can be supported from a molecular perspective. Possible strategies such as a thorough statistical analysis of numerous birth records as well as molecular studies such as radiotracing using labeled OT are proposed to test this hypothesis.