Proteomic studies of VEGFR2 in human placentas reveal protein associations with preeclampsia, diabetes, gravidity, and labor.

VEGFR2 (Vascular endothelial growth factor receptor 2) is a central regulator of placental angiogenesis. The study of the VEGFR2 proteome of chorionic villi at term revealed its partners MDMX (Double minute 4 protein) and PICALM (Phosphatidylinositol-binding clathrin assembly protein). Subsequently, the oxytocin receptor (OT-R) and vasopressin V1aR receptor were detected in MDMX and PICALM immunoprecipitations. Immunogold electron microscopy showed VEGFR2 on endothelial cell (EC) nuclei, mitochondria, and Hofbauer cells (HC), tissue-resident macrophages of the placenta. MDMX, PICALM, and V1aR were located on EC plasma membranes, nuclei, and HC nuclei. Unexpectedly, PICALM and OT-R were detected on EC projections into the fetal lumen and OT-R on 20-150 nm clusters therein, prompting the hypothesis that placental exosomes transport OT-R to the fetus and across the blood-brain barrier. Insights on gestational complications were gained by univariable and multivariable regression analyses associating preeclampsia with lower MDMX protein levels in membrane extracts of chorionic villi, and lower MDMX, PICALM, OT-R, and V1aR with spontaneous vaginal deliveries compared to cesarean deliveries before the onset of labor. We found select associations between higher MDMX, PICALM, OT-R protein levels and either gravidity, diabetes, BMI, maternal age, or neonatal weight, and correlations only between PICALM-OT-R (p < 2.7 × 10-8), PICALM-V1aR (p < 0.006), and OT-R-V1aR (p < 0.001). These results offer for exploration new partnerships in metabolic networks, tissue-resident immunity, and labor, notably for HC that predominantly express MDMX.

Patterns of neural activity in response to threatening faces are predictive of autistic traits: modulatory effects of oxytocin receptor genotype.

Autistic individuals generally demonstrate impaired emotion recognition but it is unclear whether effects are emotion-specific or influenced by oxytocin receptor (OXTR) genotype. Here we implemented a dimensional approach using an implicit emotion recognition task together with functional MRI in a large cohort of neurotypical adult participants (N = 255, male = 131, aged 17-29 years) to establish associations between autistic traits and neural and behavioral responses to specific face emotions, together with modulatory effects of OXTR genotype. A searchlight-based multivariate pattern analysis (MVPA) revealed an extensive network of frontal, basal ganglia, cingulate and limbic regions exhibiting significant predictability for autistic traits from patterns of responses to angry relative to neutral expression faces. Functional connectivity analyses revealed a genotype interaction (OXTR SNPs rs2254298, rs2268491) for coupling between the orbitofrontal cortex and mid-cingulate during angry expression processing, with a negative association between coupling and autistic traits in the risk-allele group and a positive one in the non-risk allele group. Overall, results indicate extensive emotion-specific associations primarily between patterns of neural responses to angry faces and autistic traits in regions processing motivation, reward and salience but not in early visual processing. Functional connections between these identified regions were not only associated with autistic traits but also influenced by OXTR genotype. Thus, altered patterns of neural responses to threatening faces may be a potential biomarker for autistic symptoms although modulatory influences of OXTR genotype need to be taken into account.

Are there sex differences in oxytocin and vasopressin V1a receptors ligand binding affinities?

BACKGROUND: There is substantial evidence for sex differences in the functioning of one of the most common receptor systems; G protein-coupled receptors (GPCRs). There are many points along the GPCR-mediated molecular signaling pathway at which males and females may differ, one of the first of which, chronologically, is in the stability of the interaction between the ligand and the receptor, or its binding affinity. Here we investigate the binding affinities of oxytocin (OT) and vasopressin (AVP) at the oxytocin receptor (OTR) and the vasopressin V1a receptor (V1aR), both of which are present in numerous in brain regions associated with social behavior. METHOD: In order to investigate sex- and estrous cycle-dependent differences in ligand-receptor binding affinity, male (n = 6) Syrian hamsters (Mesocricetus auratus), females on the day of estrus (E females, n = 6), and females on the second day of diestrus (D2 females n = 6) were chosen for study. Brains from hamsters were mounted on slides and competition and saturation binding assays were conducted. RESULTS: We report a remarkable similarity in the binding affinities of OT and AVP in males and females. Small differences were detected, however, in receptor and ligand specificity in females depending on whether they were in the estrous or diestrous stage of their ovulatory cycle. CONCLUSION: These data suggest that sex differences in binding affinity are not a likely source of the many sex differences that have been observed in the effects of OT and AVP in hamsters and other species.

Hypothalamic Paraventricular Stimulation Inhibits Nociceptive Wide Dynamic Range Trigeminocervical Complex Cells via Oxytocinergic Transmission.

Oxytocinergic transmission blocks nociception at the peripheral, spinal, and supraspinal levels through the oxytocin receptor (OTR). Indeed, a neuronal pathway from the hypothalamic paraventricular nucleus (PVN) to the spinal cord and trigeminal nucleus caudalis (Sp5c) has been described. Hence, although the trigeminocervical complex (TCC), an anatomical area spanning the Sp5c, C1, and C2 regions, plays a role in some pain disorders associated with craniofacial structures (e.g., migraine), the role of oxytocinergic transmission in modulating nociception at this level has been poorly explored. Hence, in vivo electrophysiological recordings of TCC wide dynamic range (WDR) cells sensitive to stimulation of the periorbital or meningeal region were performed in male Wistar rats. PVN electrical stimulation diminished the neuronal firing evoked by periorbital or meningeal electrical stimulation; this inhibition was reversed by OTR antagonists administered locally. Accordingly, neuronal projections (using Fluoro-Ruby) from the PVN to the WDR cells filled with Neurobiotin were observed. Moreover, colocalization between OTR and calcitonin gene-related peptide (CGRP) or OTR and GABA was found near Neurobiotin-filled WDR cells. Retrograde neuronal tracers deposited at the meningeal (True-Blue, TB) and infraorbital nerves (Fluoro-Gold, FG) showed that at the trigeminal ganglion (TG), some cells were immunopositive to both fluorophores, suggesting that some TG cells send projections via the V1 and V2 trigeminal branches. Together, these data may imply that endogenous oxytocinergic transmission inhibits the nociceptive activity of second-order neurons via OTR activation in CGRPergic (primary afferent fibers) and GABAergic cells.

Cinnamic acid promotes elongation of hair peg-like sprouting in hair follicle organoids via oxytocin receptor activation.

Considerable global demand exists for the development of novel drugs for the treatment of alopecia. A recent report demonstrated that oxytocin promotes hair growth activity in human dermal papilla (DP) cells; however, its application in drugs or cosmetic products is challenging because rapid degradation and relatively large molecular weight prevent long-term topical administration on the scalp. Here, we examined cinnamic acid, a small molecule activator for oxytocin receptor (OXTR) expression. Treatment with cinnamic acid led to upregulation of OXTR and trichogenic gene expression in human DP cells. Furthermore, inhibition of OXTR with an antagonist, L-371,257, suppressed hair growth-related gene expression in DP cells. These findings suggest that cinnamic acid enhances the hair growth ability of DP cells via oxytocin signaling. Additionally, we tested the hair growth-promoting effects of cinnamic acid using hair follicle organoids in vitro and observed that cinnamic acid significantly promoted the growth of hair peg-like sprouting. These promising results may be useful for developing hair growth-promoting products targeting oxytocin.

Adolescent social isolation disrupts developmental tuning of neuropeptide circuits in the hypothalamus to amygdala regulating social and defensive behavior.

Engaging in positive social (i.e., prosocial) interactions during adolescence acts to modulate neural circuits that determine adult adaptive behavior. While accumulating evidence indicates that a strong craving for prosocial behavior contributes to sustaining neural development, the consequences of social deprivation during adolescence on social neural circuits, including those involving oxytocin (OXT) and vasopressin (AVP), are poorly characterized. We evaluated adaptive behaviors in socially isolated mice, including anxiety-like, social, and defensive behaviors, along with OXT and AVP neural profiles in relevant brain regions. Social isolation from postnatal day (P-)22 to P-48 induced enhanced defensive and exploratory behaviors, in nonsocial and social contexts. Unlike OXT neurons, AVP+ cell density in the paraventricular nucleus of the hypothalamus increases with age in males. Social isolation also modulated gene expression in the medial amygdala (MeA), including the upregulation of OXT receptors in males and the downregulation of AVP1a receptors in both sexes. Socially isolated mice showed an enhanced defensive, anogenital approach toward a novel adult female during direct social interactions. Subsequent c-Fos mapping revealed diminished neural activity in restricted brain areas, including the MeA, lateral septum, and posterior intralaminar nucleus of the thalamus, in socially isolated mice. These data indicate that neural signals arising from daily social interactions invoke region-specific modification of neuropeptide expression that coordinates with altered defensiveness and neural responsivities, including OXT- and AVP-projecting regions. The present findings indicate an involvement of OXT and AVP circuits in adolescent neural and behavioral plasticity that is tuned by daily social interaction.

Oxytocin alleviates periodontitis in adult rats.

OBJECTIVE: The objective of the study was to evaluate the expression of oxytocin receptors in normal and inflamed gingiva, as well as the effects of systemic administration of oxytocin in bone loss and gum inflammatory mediators in a rat model of experimental periodontitis. BACKGROUND DATA: Current evidence supports the hypothesis of a disbalance between the oral microbiota and the host's immune response in the pathogenesis of periodontitis. Increased complexity of the microbial biofilm present in the periodontal pocket leads to local production of nitrogen and oxygen-reactive species, cytokines, chemokines, and other proinflammatory mediators which contribute to periodontal tissue destruction and bone loss. Oxytocin has been suggested to participate in the modulation of immune and inflammatory processes. We have previously shown that oxytocin, nitric oxide, and endocannabinoid system interact providing a mechanism of regulation for systemic inflammation. Here, we aimed at investigating not only the presence and levels of expression of oxytocin receptors on healthy and inflamed gingiva, but also the effects of oxytocin treatment on alveolar bone loss, and systemic and gum expression of inflammatory mediators involved in periodontal tissue damage using ligature-induced periodontitis. Therefore, anti-inflammatory strategies oriented at modulating the host's immune response could be valuable adjuvants to the main treatment of periodontal disease. METHODS: We used an animal model of ligature-induced periodontitis involving the placement of a linen thread (Barbour flax 100% linen suture, No. 50; size 2/0) ligature around the neck of first lower molars of adult male rats. The ligature was left in place during the entire experiment (7 days) until euthanasia. Animals with periodontitis received daily treatment with oxytocin (OXT, 1000 µg/kg, sc.) or vehicle and/or atosiban (3 mg/kg, sc.), an antagonist of oxytocin receptors. The distance between the cement-enamel junction and the alveolar bone crest was measured in stained hemimandibles in the long axis of both buccal and lingual surfaces of both inferior first molars using a caliper. TNF-α levels in plasma were determined using specific rat enzyme-linked immunosorbent assays (ELISA). OXT receptors, IL-6, IL-1ß, and TNF-α expression were determined in gingival tissues by semiquantitative or real-time PCR. RESULTS: We show that oxytocin receptors are expressed in normal and inflamed gingival tissues in male rats. We also show that the systemic administration of oxytocin prevents the experimental periodontitis-induced increased gum expression of oxytocin receptors, TNF-α, IL-6, and IL-1ß (p < .05). Furthermore, we observed a reduction in bone loss in rats treated with oxytocin in our model. CONCLUSIONS: Our results demonstrate that oxytocin is a novel and potent modulator of the gingival inflammatory process together with bone loss preventing effects in an experimental model of ligature-induced periodontitis.

Methylation of the Oxytocin, Oxytocin Receptor, and Vasopressin Gene Promoters in Tobacco Use Disorder during Cessation.

INTRODUCTION: Vasopressin (AVP) and oxytocin (OT) exert sex-specific effects on social pair bonding and stress reactions while also influencing craving in substance use disorders. In this regard, intranasal oxytocin (OT) and AVP antagonists present potential treatments for tobacco use disorder (TUD). Since transcription of both hormones is also regulated by gene methylation, we hypothesized sex-specific changes in methylation levels of the AVP, OT, and OT receptor (OXTR) gene during nicotine withdrawal. METHODS: The study population consisted of 49 smokers (29 males, 20 females) and 51 healthy non-smokers (25 males, 26 females). Blood was drawn at day 1, day 7, and day 14 of smoking cessation. Craving was assessed with the questionnaire on smoking urges (QSU). RESULTS: Throughout cessation, mean methylation of the OT promoter gene increased in males and decreased in females. OXTR receptor methylation decreased in females, while in males it was significantly lower at day 7. Regarding the AVP promoter, mean methylation increased in males while there were no changes in females. Using mixed linear modeling, CpG position, time point, sex, and the interaction of time point and sex as well as time point, sex, and QSU had a significant fixed effect on OT and AVP gene methylation. The interaction effect suggests that sex, time point, and QSU are interrelated, meaning that, depending on the sex, methylation could be different at different time points and vice versa. There was no significant effect of QSU on mean OXTR methylation. DISCUSSION: We identified differences at specific CpGs between controls and smokers in OT and AVP and in overall methylation of the AVP gene. Furthermore, we found sex-specific changes in mean methylation levels of the mentioned genes throughout smoking cessation, underlining the relevance of sex in the OT and vasopressin system. This is the first study on epigenetic regulation of the OT promoter in TUD. Our results have implications for research on the utility of the AVP and OT system for treating substance craving. Future studies on both targets need to analyze their effect in the context of sex, social factors, and gene regulation.

A dedicated hypothalamic oxytocin circuit controls aversive social learning.

To survive in a complex social group, one needs to know who to approach and, more importantly, who to avoid. In mice, a single defeat causes the losing mouse to stay away from the winner for weeks1. Here through a series of functional manipulation and recording experiments, we identify oxytocin neurons in the retrochiasmatic supraoptic nucleus (SOROXT) and oxytocin-receptor-expressing cells in the anterior subdivision of the ventromedial hypothalamus, ventrolateral part (aVMHvlOXTR) as a key circuit motif for defeat-induced social avoidance. Before defeat, aVMHvlOXTR cells minimally respond to aggressor cues. During defeat, aVMHvlOXTR cells are highly activated and, with the help of an exclusive oxytocin supply from the SOR, potentiate their responses to aggressor cues. After defeat, strong aggressor-induced aVMHvlOXTR cell activation drives the animal to avoid the aggressor and minimizes future defeat. Our study uncovers a neural process that supports rapid social learning caused by defeat and highlights the importance of the brain oxytocin system in social plasticity.

Melanocortin agonism in a social context selectively activates nucleus accumbens in an oxytocin-dependent manner.

Social deficits are debilitating features of many psychiatric disorders, including autism. While time-intensive behavioral therapy is moderately effective, there are no pharmacological interventions for social deficits in autism. Many studies have attempted to treat social deficits using the neuropeptide oxytocin for its powerful neuromodulatory abilities and influence on social behaviors and cognition. However, clinical trials utilizing supplementation paradigms in which exogenous oxytocin is chronically administered independent of context have failed. An alternative treatment paradigm suggests pharmacologically activating the endogenous oxytocin system during behavioral therapy to enhance the efficacy of therapy by facilitating social learning. To this end, melanocortin receptor agonists like Melanotan II (MTII), which induces central oxytocin release and accelerates formation of partner preference, a form of social learning, in prairie voles, are promising pharmacological tools. To model pharmacological activation of the endogenous oxytocin system during behavioral therapy, we administered MTII prior to social interactions between male and female voles. We assessed its effect on oxytocin-dependent activity in brain regions subserving social learning using Fos expression as a proxy for neuronal activation. In non-social contexts, MTII only activated hypothalamic paraventricular nucleus, a primary site of oxytocin synthesis. However, during social interactions, MTII selectively increased oxytocin-dependent activation of nucleus accumbens, a site critical for social learning. These results suggest a mechanism for the MTII-induced acceleration of partner preference formation observed in previous studies. Moreover, they are consistent with the hypothesis that pharmacologically activating the endogenous oxytocin system with a melanocortin agonist during behavioral therapy has potential to facilitate social learning.

Age-dependent effects of oxytocin in brain regions enriched with oxytocin receptors.

Although intranasal oxytocin administration to tap into central functions is the most commonly used non-invasive means for exploring oxytocin's role in human cognition and behavior, the way by which intranasal oxytocin acts on the brain is not yet fully understood. Recent research suggests that brain regions densely populated with oxytocin receptors may play a central role in intranasal oxytocin's action mechanisms in the brain. In particular, intranasal oxytocin may act directly on (subcortical) regions rich in oxytocin receptors via binding to these receptors while only indirectly affecting other (cortical) regions via their neural connections to oxytocin receptor-enriched regions. Aligned with this notion, the current study adopted a novel approach to test 1) whether the connections between oxytocin receptor-enriched regions (i.e., the thalamus, pallidum, caudate nucleus, putamen, and olfactory bulbs) and other regions in the brain were responsive to intranasal oxytocin administration, and 2) whether oxytocin-induced effects varied as a function of age. Forty-six young (24.96 ± 3.06 years) and 44 older (69.89 ± 2.99 years) participants were randomized, in a double-blind procedure, to self-administer either intranasal oxytocin or placebo before resting-state fMRI. Results supported age-dependency in the effects of intranasal oxytocin administration on connectivity between oxytocin receptor-enriched regions and other regions in the brain. Specifically, compared to placebo, oxytocin decreased both connectivity density and connectivity strength of the thalamus for young participants while it increased connectivity density and connectivity strength of the caudate for older participants. These findings inform the mechanisms underlying the effects of exogenous oxytocin on brain function and highlight the importance of age in these processes.

Prenatal allergic inflammation in rats confers sex-specific alterations to oxytocin and vasopressin innervation in social brain regions.

Prenatal exposure to inflammation via maternal infection, allergy, or autoimmunity increases one's risk for developing neurodevelopmental and psychiatric disorders. Many of these disorders are associated with altered social behavior, yet the mechanisms underlying inflammation-induced social impairment remain unknown. We previously found that a rat model of acute allergic maternal immune activation (MIA) produced deficits like those found in MIA-linked disorders, including impairments in juvenile social play behavior. The neuropeptides oxytocin (OT) and arginine vasopressin (AVP) regulate social behavior, including juvenile social play, across mammalian species. OT and AVP are also implicated in neuropsychiatric disorders characterized by social impairment, making them good candidate regulators of social deficits after MIA. We profiled how acute prenatal exposure to allergic MIA changed OT and AVP innervation in several brain regions important for social behavior in juvenile male and female rat offspring. We also assessed whether MIA altered additional behavioral phenotypes related to sociality and anxiety. We found that allergic MIA increased OT and AVP fiber immunoreactivity in the medial amygdala and had sex-specific effects in the nucleus accumbens, bed nucleus of the stria terminalis, and lateral hypothalamic area. We also found that MIA reduced ultrasonic vocalizations in neonates and increased the stereotypical nature of self-grooming behavior. Overall, these findings suggest that there may be sex-specific mechanisms underlying MIA-induced behavioral impairment and underscore OT and AVP as ideal candidates for future mechanistic studies.

Dual Oxytocin Receptor-G Protein Signaling in the Autoregulation of Activities of Oxytocin Neurons.

Oxytocin (OT), a hypothalamic nonaneuropeptide, can extensively modulate mental and physical activities; however, the regulation of its secretion from hypothalamic OT neurons remains poorly understood. OT neuronal activity is generally modulated by neurochemical environment, synaptic inputs, astrocytic plasticity, and interneuronal interactions. By changing intracellular signals and ion channel activity, these extracellular factors dynamically regulate OT neuronal activity and OT release in a microdomain-specific manner. In this process, OT receptor (OTR) and OTR-coupled G proteins are pivotal, typically observed during lactation. Suckling-elicited somatodendritic release of OT causes sequential activation of Gq and Gs proteins to increase the firing rate gradually and trigger burst firing transiently, and then of Gi/o protein to cause post-burst inhibition as a result of potential bolus somatodendritic release of OT during the burst-like discharges. Under chronic social stress like mother-baby separation and cesarean section, excessive somatodendritic secretion of OT and over-excitation of OT neurons cause post-excitation inhibition of OT neuronal activity and reduction of OT secretion. In this process, dominance of G protein that couples to OTR is switched from Gq to Gi/o type because of inhibition of OTR-Gq signaling following negative feedback of downstream Gq signaling or crosstalk of Gq with Gs and Gi signals. This review summarizes our current understandings of OT/OTR signaling in the autoregulation of OT neuronal activity under physiological and pathological conditions.

Stimulating oxytocin receptors in the basolateral amygdala enhances stimulus processing: Differential and consistent effects for stimuli paired with fear versus sucrose in extinction and reversal learning.

Oxytocin (OT) influences a range of social behaviors by enhancing the salience of social cues and regulating the expression of specific social behaviors (e.g., maternal care versus defensive aggression). We previously showed that stimulating OT receptors in the basolateral amygdala of rats also enhanced the salience of fear conditioned stimuli: relative to rats given vehicle infusions, rats infused with [Thr4,Gly7]-oxytocin (TGOT), a selective OT receptor agonist, showed greater discrimination between a cue predictive of danger, and one that signaled safety. In the present series of experiments using male rats, the effects of OT receptor activation in the basolateral amygdala on stimulus processing were examined further using conditioning protocols that consist of changes in stimulus-outcome contingencies (i.e., extinction and reversal), and with stimuli paired with aversive (i.e., foot shock) and appetitive (i.e., sucrose) outcomes. It was revealed that the effects of OTR stimulation diverge for aversive and appetitive learning - enhancing the former but not the latter. However, across both types of learning, OTR stimulation enhanced the detection of conditioned stimuli. Overall, these results are consistent with an emerging view of OT's effects on stimulus salience; facilitating the detection of meaningful stimuli while reducing responding to those that are irrelevant.

Extended Metadynamics Protocol for Binding/Unbinding Free Energies of Peptide Ligands to Class A G-Protein-Coupled Receptors.

A metadynamics protocol is presented to characterize the binding and unbinding of peptide ligands to class A G-protein-coupled receptors (GPCRs). The protocol expands on the one previously presented for binding and unbinding small-molecule ligands to class A GPCRs and accounts for the more demanding nature of the peptide binding-unbinding process. It applies to almost all class A GPCRs. Exemplary simulations are described for subtypes Y1R, Y2R, and Y4R of the neuropeptide Y receptor family, vasopressin binding to the vasopressin V2 receptor (V2R), and oxytocin binding to the oxytocin receptor (OTR). Binding free energies and the positions of alternative binding sites are presented and, where possible, compared with the experiment.

Beyond the binary: Characterizing the relationships between sex and neuropeptide receptor binding density measures in the rat brain.

Sex differences exist in numerous parameters of the brain. Yet, sex-related factors are part of a large set of variables that interact to affect many aspects of brain structure and function. This raises questions regarding how to interpret findings of sex differences at the level of single brain measures and the brain as a whole. In the present study, we reanalyzed two datasets consisting of measures of oxytocin, vasopressin V1a, and mu opioid receptor binding densities in multiple brain regions in rats. At the level of single brain measures, we found that sex differences were rarely dimorphic and were largely persistent across estrous stage and parental status but not across age or context. At the level of aggregates of brain measures showing sex differences, we tested whether individual brains are 'mosaics' of female-typical and male-typical measures or are internally consistent, having either only female-typical or only male-typical measures. We found mosaicism for measures showing overlap between females and males. Mosaicism was higher a) with a larger number of measures, b) with smaller effect sizes of the sex difference in these measures, and c) in rats with more diverse life experiences. Together, these results highlight the limitations of the binary framework for interpreting sex effects on the brain and suggest two complementary pathways to studying the contribution of sex to brain function: (1) focusing on measures showing dimorphic and persistent sex differences and (2) exploring the relations between specific brain mosaics and specific endpoints.

A targeted long-read sequencing approach questions the association of OXTR methylation with high-functioning autism.

BACKGROUND: DNA sequence variation and altered epigenetic regulation of the oxytocin receptor gene (OXTR) have been implicated in autism and autistic-like behaviors. While previous studies have examined subsegments of OXTR, nanopore Cas9-targeted sequencing (nCATS) allows deep characterization of entire genes with simultaneous assessment of epigenetic 5-methylcytosine (5mC) modification and without the need for prior DNA amplification or bisulfite conversion. This pilot study uses an nCATS approach to sequence the entire OXTR gene and its regulatory construct and screen for 5mC modification to compare results between individuals with high-functioning autism (HFA) and neurotypical controls (NC). METHODS: Using DNA extracted from peripheral blood, OXTR (Hg38, chr3: 8750381-8770434, 20,054 base pairs) was analyzed by nCATS. 5mC modification probabilities were calculated and visualized across the gene and differential methylation analysis was performed. RESULTS: Twenty adults with HFA (10 males, 10 females) and 20 age- and sex-matched NC (± 5 years) were included. There were no apparent group differences in the entire OXTR gene sequence, except for the intron variant rs918316, which was clustered in the HFA group. However, differential methylation analysis did not reveal a single significant group-dependent differentially methylated site among the 412 CpG sites captured. LIMITATIONS: Limitations of this study include the small number of samples due to the pilot nature of the study, which particularly limits the relevance of the sequence variants found. It should also be noted that the use of peripheral blood material limits the ability to draw conclusions about central processes. CONCLUSIONS: Previous findings of autism-associated OXTR epigenetic alterations were not reproducible with our method. In our opinion, this may lead to a reconsideration of the relevance of altered methylation at individual OXTR CpG positions in autism research. However, given the pilot nature of the study, these results need to be replicated in independent cohorts and with larger sample sizes.

Oxytocinergic projection from the hypothalamus to supramammillary nucleus drives recognition memory in mice.

Oxytocin (OXT) neurons project to various brain regions and its receptor expression is widely distributed. Although it has been reported that OXT administration affects cognitive function, it is unclear how endogenous OXT plays roles in cognitive function. The present study examined the role of endogenous OXT in mice cognitive function. OXT neurons were specifically activated by OXT neuron-specific excitatory Designer Receptors Exclusively Activated by Designer Drug expression system and following administration of clozapine-N-oxide (CNO). Object recognition memory was assessed with the novel object recognition task (NORT). Moreover, we observed the expression of c-Fos via immunohistochemical staining to confirm neuronal activity. In NORT, the novel object exploration time percentage significantly increased in CNO-treated mice. CNO-treated mice showed a significant increase in the number of c-Fos-positive cells in the supramammillary nucleus (SuM). In addition, we found that the OXT-positive fibers from paraventricular hypothalamic nucleus (PVN) were identified in the SuM. Furthermore, mice injected locally with CNO into the SuM to activate OXTergic axons projecting from the PVN to the SuM showed significantly increased percentage time of novel object exploration. Taken together, we proposed that object recognition memory in mice could be modulated by OXT neurons in the PVN projecting to the SuM.

At the Head and Heart of Oxytocin's Stress-Regulatory Neural and Cardiac Effects: A Chronic Administration RCT in Children with Autism.

INTRODUCTION: Intranasal administration of oxytocin presents a promising new approach to reduce disability associated with an autism spectrum disorder diagnosis. Previous investigations have emphasized the amygdala as the neural foundation for oxytocin's acute effects. However, to fully understand oxytocin's therapeutic potential, it is crucial to gain insight into the neuroplastic changes in amygdala circuitry induced from chronic oxytocin administrations, particularly in pediatric populations. OBJECTIVE: We aimed to examine the impact of a 4-week course of intranasal oxytocin on amygdala functional connectivity in children with autism, compared to placebo. Additionally, we investigated whether oxytocin improves cardiac autonomic arousal, as indexed by high-frequency heart rate variability. METHODS: Fifty-seven children with autism aged 8-12 years (45 boys, 12 girls) participated in a double-blind, randomized pharmaco-neuroimaging trial involving twice-daily administrations of intranasal oxytocin or placebo. Resting-state fMRI scans and simultaneous, in-scanner heart rate recordings were obtained before, immediately after, and 4 weeks after the nasal spray administration period. RESULTS: Significant reductions in intrinsic amygdala-orbitofrontal connectivity were observed, particularly at the 4-week follow-up session. These reductions were correlated with improved social symptoms and lower cardiac autonomic arousal. Further, oxytocin's neural and cardiac autonomic effects were modulated by epigenetic modifications of the oxytocin receptor gene. The effects were more pronounced in children with reduced epigenetic methylation, signifying heightened expression of the oxytocin receptor. CONCLUSION: These findings underscore that a 4-week oxytocin administration course decreases amygdala connectivity and improves cardiac autonomic balance. Epigenetic modulators may explain inter-individual variation in responses to oxytocin.

Differential Roles of Oxytocin Receptors in the Prefrontal Cortex and Nucleus Accumbens on Cocaine Self-Administration and Reinstatement of Cued Cocaine Seeking in Male Rats.

BACKGROUND: Little is known about the specific roles of cortical and accumbal oxytocin receptors in drug use disorders. To better understand the importance of the endogenous oxytocin system in cocaine relapse behavior, we developed an adeno-associated viral vector-expressing short hairpin (sh) RNAs to selectively degrade the rat oxytocin receptor (OxyR) mRNA in vivo. METHODS: Male (Sprague-Dawley) rats received bilateral infusions of the shRNA for the oxytocin receptor (shOxyR) or an shRNA control virus into the prefrontal cortex (PFC) or the nucleus accumbens core (NAc). Rats self-administered cocaine on an escalating FR ratio for 14 days, lever responding was extinguished, and rats were tested for cued and cocaine-primed reinstatement of drug seeking. RESULTS: OxyR knockdown in the PFC delayed the acquisition of lever pressing on an fixed ratio 1 schedule of reinforcement. All rats eventually acquired the same level of lever pressing and discrimination, and there were no differences in extinction. OxyR knockdown in the NAc had no effect during acquisition. In both the PFC and NAc, the shOxyR decreased cued reinstatement relative to shRNA control virus but was without effect during drug-primed reinstatement. OxyR knockdown in the PFC increased chamber activity during a social interaction task. CONCLUSIONS: This study provides critical new information about how endogenous OxyRs function to affect drug seeking in response to different precipitators of relapse. The tool developed to knockdown OxyRs in rat could provide important new insights that aid development of oxytocin-based therapeutics to reduce return-to-use episodes in people with substance use disorder and other neuropsychiatric disorders.