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Abdominal pain presents an onerous reality for millions of people affected by gastrointestinal disorders such as irritable bowel syndrome (IBS) and inflammatory bowel diseases (IBD). The oxytocin receptor (OTR) has emerged as a new analgesic drug target with OTR expression upregulated on colon-innervating nociceptors in chronic visceral hypersensitivity states, accessible via luminal delivery. However, the low gastrointestinal stability of OTR's endogenous peptide ligand oxytocin (OT) is a bottleneck for therapeutic development. Here, we report the development of potent and fully gut-stable OT analogues, laying the foundation for a new area of oral gut-specific peptide therapeutics. Ligand optimisation guided by structure-gut-stability-activity relationships yielded highly stable analogues (t 1/2 >24 h, compared to t 1/2 <10 min of OT in intestinal fluid) equipotent to OT (~3 nM) and with enhanced OTR selectivity. Intra-colonic administration of the lead ligand significantly reduced colonic mechanical hypersensitivity in a concentration-dependent manner in a mouse model of chronic abdominal pain. Moreover, oral administration of the lead ligand also displayed significant analgesia in this abdominal pain mouse model. The generated ligands and employed strategies could pave the way to a new class of oral gut-specific peptides to study and combat chronic gastrointestinal disorders, an area with substantial unmet medical needs., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Oxytocin (OXY), a peptide hormone and neurotransmitter essential for biological processes with nine distinct amino acid residues, has received significant attention due to its illegal use in food adulteration and stimulating milk ejection in cattle. Herein, for the first time, electrochemical detection of oxytocin (OXY) is reported using a novel nanocomposite consisting of a Zn-Cu metal-organic framework (Zn-Cu MOF) decorated on reduced graphene oxide (rGO). An octahedral surface morphology with a crystalline structure of 45 nm in size, formation of a metal-oxygen bond, an enhanced pore diameter of 6.8 nm, a specific surface area of 70.8 m 2 g -1 , and a pore volume of 0.08 cm 3 g -1 were revealed by different characterization techniques. The electro-catalytic activity of the Zn-Cu MOF/rGO nanocomposite increased substantially owing to the synergistic effect, which is evident from cyclic voltammetry (CV) when compared to those of the Zn-MOF, the Cu-MOF, Zn MOF/rGO, Cu MOF/rGO, and Zn-Cu MOF, keeping other parameters the same. Moreover, the electrochemical impedance spectroscopy (EIS) spectra reveal the excellent conductivity of the nanocomposite. The experimental parameters, viz. electrolyte pH (5), supporting electrolyte (0.1 M ABS), and volume of coating (12 μL), were optimized. The differential pulse voltammetry (DPV) technique was adopted to determine the OXY with the lowest limit of detection (LOD) of 1.1 nM (S/N = 3) with a linear range of 40-400 nM. The analytical application of the modified electrode was examined by spiking OXY in pasteurized toned milk, skimmed powder milk, animal milk, and RO water, with a good recovery range of 95-106%.

Bioelectrodes with low carbon footprint can provide an innovative solution to the surmounting levels of e-waste. Biodegradable polymers offer green and sustainable alternatives to synthetic materials. Here, a chitosan-carbon nanofiber (CNF) based membrane has been developed and functionalized for electrochemical sensing application. The surface characterization of the membrane revealed crystalline structure with uniform particle distribution, and surface area of 25.52 m 2 /g and pore volume of 0.0233 cm 3 /g. The membrane was functionalized to develop a bioelectrode for the detection of exogenous oxytocin in milk. Electrochemical impedance spectroscopy was employed to determine oxytocin in a linear concentration range of 10 to 10 5 ng/mL. The developed bioelectrode showed an LOD of 24.98 ± 11.37 pg/mL and sensitivity of 2.77 × 10 -10 Ω / log ng mL -1 /mm 2 for oxytocin in milk samples with 90.85-113.34 percent recovery. The chitosan-CNF membrane is ecologically safe and opens new avenues for environment-friendly disposable materials for sensing applications., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Oxytocin (OT) and vasopressin (AVP) are conserved peptide signaling hormones that are critical for diverse processes including osmotic homeostasis, reproduction, lactation and social interaction. OT acts through the oxytocin receptor (OTR), a magnesium-dependent G protein-coupled receptor that is a therapeutic target for treatment of postpartum hemorrhage, dysfunctional labor and autism. However, the molecular mechanisms that underlie OTR activation by OT and the dependence on magnesium remain unknown. Here we present the wild-type active-state structure of human OTR bound to OT and miniG q/i determined by cryo-EM. The structure reveals a unique activation mechanism adopted by OTR involving both the formation of a Mg 2+ coordination complex between OT and the receptor, and disruption of transmembrane helix 7 (TM7) by OT. Our functional assays demonstrate the role of TM7 disruption and provide the mechanism of full agonism by OT and partial agonism by OT analogs. Furthermore, we find that the identity of a single cation-coordinating residue across vasopressin family receptors determines whether the receptor is cation-dependent. Collectively, these results demonstrate how the Mg 2+ -dependent OTR is activated by OT, provide essential information for structure-based drug discovery efforts and shed light on the molecular determinants of cation dependence of vasopressin family receptors throughout the animal kingdom., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

In peptide production, oxidative sulfitolysis can be used to protect the cysteine residues during purification, and the introduction of a negative charge aids solubility. Subsequent controlled reduction aids in ensuring correct disulfide bridging. In vivo, these problems are overcome through interaction with chaperones. Here, a versatile peptide production process has been developed using an angled vortex fluidic device (VFD), which expands the viable pH range of oxidative sulfitolysis from pH 10.5 under batch conditions, to full conversion within 20 min at pH 9-10.5 utilising the VFD. VFD processing gave 10-fold greater conversion than using traditional batch processing, which has potential in many applications of the sulfitolysis reaction.

It is known since the fifties that oxytocin is a neurohormone synthesized in the brain and released in blood circulation to trigger uterus contraction during delivery. It is also involved in milk ejection during breast-feeding. Over the past 25 years, many other central and peripheral functions have been discovered, in particular for attachment between child and parents as well as between individuals and interaction between a human being and its social group. Over this period, we have studied the functional supramolecular architecture of the hormone bound to its receptor. This information was used to design pharmacological probes and drug candidates. This led to the discovery of the first non-peptide oxytocin receptor full agonist. This molecule, LIT-001, restores social interaction in an animal model of autism and paves the way for a treatment of this neurodevelopmental disorder., (© Société de Biologie, 2023.)

Objective: To establish the storage conditions of oxytocin in a health facility in a low-income country with a tropical climate, as suboptimal storage may lead to ineffectiveness of drugs essential to prevent and treat postpartum hemorrhage., Methods: At Mulago National Referral Hospital (28 000-33 000 deliveries/year) in Kampala, Uganda, temperature logging Safe-Rx cards were placed in boxes of oxytocin and in every known storage location. The route of the boxes through the hospital was tracked for 54 days, and storage conditions were observed., Results: Oxytocin was stored within the recommended temperature range (2°C-8°C) 24% of the time. The average temperature measured within the oxytocin boxes was 18.2°C, with a minimum of -2.3°C and maximum of 30.4°C. Six out of twelve known storage places had a refrigerator, but not one location stored medication at the recommended temperature constantly. The average temperature in the storage places ranged from 9.5°C to 27.6°C, with a minimum temperature of 2.3°C and maximum of 30.9°C., Conclusion: Oxytocin is not stored in the recommended temperature range for the majority of time. The presence of refrigerators does not ensure adherence to advised temperature storage conditions., (© 2020 International Federation of Gynecology and Obstetrics.)

Oxytocin is a cyclic nonapeptide used to induce labor and prevent bleeding after childbirth. Due to its instability, storage and transport of oxytocin formulations can be problematic in hot/tropical climates. The aim of this study was to investigate the effect of trehalose and select antioxidants (uric acid, butylated hydroxytoluene, and l-ascorbic acid) on oxytocin stability in solution. The effect of buffer composition and acetate buffer concentration was also studied. Acetate buffer was found to work better than citrate/phosphate buffer for the oxytocin stability. Lower acetate buffer concentrations (0.025 M or less) were also found to yield improved oxytocin stability compared to higher concentrations. Although known degradation pathways of oxytocin include oxidation, the antioxidants uric acid and butylated hydroxytoluene had negligible effect on the oxytocin stability while l-ascorbic acid led to significantly faster degradation. Despite trehalose's reputation as a great stabilizer for biomolecules, it also had small to negligible effect on oxytocin stability at concentrations up to 1 M in acetate buffer. These results were surprising given the present literature on trehalose as a stabilizer for various biomolecules, including proteins and lipids., (© 2021 European Peptide Society and John Wiley & Sons, Ltd.)

Oxytocin/vasopressin-related neuropeptides are highly conserved and play major roles in regulating social behavior across vertebrates. However, whether their insect orthologue, inotocin, regulates the behavior of social groups remains unknown. Here, we show that in the clonal raider ant Ooceraea biroi, individuals that perform tasks outside the nest have higher levels of inotocin in their brains than individuals of the same age that remain inside the nest. We also show that older ants, which spend more time outside the nest, have higher inotocin levels than younger ants. Inotocin thus correlates with the propensity to perform tasks outside the nest. Additionally, increasing inotocin pharmacologically increases the tendency of ants to leave the nest. However, this effect is contingent on age and social context. Pharmacologically treated older ants have a higher propensity to leave the nest only in the presence of larvae, whereas younger ants seem to do so only in the presence of pupae. Our results suggest that inotocin signaling plays an important role in modulating behaviors that correlate with age, such as social foraging, possibly by modulating behavioral response thresholds to specific social cues. Inotocin signaling thereby likely contributes to behavioral individuality and division of labor in ant societies., Competing Interests: The authors have declared that no competing interests exist.

Peptides are commonly used as biosensors for analytes such as metal ions as they have natural binding preferences. In our previous peptide-based impedimetric metal ion biosensors, a monolayer of the peptide was anchored covalently to the electrode. Binding of metal ions resulted in a conformational change of the oxytocin peptide in the monolayer, which was measured using electrochemical impedance spectroscopy. Here, we demonstrate that sensing can be achieved also when the oxytocin is non-covalently integrated into an alkanethiol host monolayer. We show that ion-binding cause morphological changes to the dense host layer, which translates into enhanced impedimetric signals compared to direct covalent assembly strategies. This biosensor proved selective and sensitive for Zn 2+ ions in the range of nano- to micro-molar concentrations. This strategy offers an approach to utilize peptide flexibility in monitoring their response to the environment while embedded in a hydrophobic monolayer.

Oxytocin (OT) is a developmentally important neuropeptide recognized to play a dominant role in social functioning and stress-related behaviors, in a sex-dependent manner. Nonetheless, the underlining factors driving OT and OT receptor (OTR) early brain development remain unclear. Recent evidence highlight the critical influence of gut microbiota and its bidirectional interaction with the brain on neurodevelopment via the gut microbiota-brain axis. Therefore, we aimed to determine the impact of gut microbiota on the OTR system of the rat brain at different developmental stages in a pilot study. Quantitative OTR [ 125 I]-OVTA autoradiographic binding was carried out in the forebrain of male and female conventional (CON) and germ-free (GF) rats at postnatal days (PND) 8, 22, and 116-150. OTR binding was also assessed in the eyes of PND 1 and PND 4 GF female rats. Significant "microbiota × sex × region" interaction and age-dependent effects on OTR binding were demonstrated. Microbiota status influenced OTR levels in males but not females with higher levels of OTR observed in GF versus CON rats in the cingulate, prelimbic, and lateral/medial/ventral orbital cortex, and septum across all age groups, while sex differences were observed in GF, but not in CON rats. Interestingly, OTRs present in the eyes of CON rats were abolished in GF rats. This is the first study to uncover a sex-specific role of gut microbiota on the central OTR system, which may have implications in understanding the developmental neuroadaptations critical for behavioral regulation and the etiology of certain neurodevelopmental disorders., (© 2021 The Authors. Developmental Neurobiology published by Wiley Periodicals LLC.)

Oxytocin (OXT) is an important peptide that is mainly used as a therapeutic drug to induce labor or strengthen uterine contractions, or to control bleeding after childbirth. OXT has also been reported as a biomarker linked to emotion, and as a potential biomarker for cancer diagnosis. The accurate purity characterization of OXT calibrators is critical for quality control of pharmaceuticals and the development of reference measurement systems for this analyte in laboratory medicine. OXT possesses the particular analytical measurement challenge of a disulfide bond. Accurate value assignment of the purity of oxytocin calibrators can be carried out by applying the mass balance approach or alternative approaches such as amino acid analysis, quantitative nuclear magnetic resonance spectrometry, and nitrogen determination. In order to avoid biases, all these approaches require a correction for structurally related peptide impurities. Structurally related peptide impurities present in a synthetic OXT material have been identified and quantified by a newly developed and in-house-validated liquid chromatography-high-resolution mass spectrometry (LC-hrMS) method. This method was adopted for the measurement of the study material used for an international comparison evaluating the competencies of laboratories to perform peptide characterization. Eighteen structurally related impurities were identified, confirmed, and accurately quantified in the OXT study material by using LC-hrMS. The study material contained a total mass fraction of 31.1 mg/g structurally related OXT impurities with an associated expanded uncertainty of 1.7 mg/g.

Oxytocin is a hormone that is increasingly being used for welfare evaluation in animals. Although several types of samples have been used for oxytocin measurement, saliva can be a suitable option for pigs producing less stress than blood sampling. In this study, 3 different methods for oxytocin measurements, 2 based on alphaLISA technology (one with a monoclonal and other with a polyclonal antibody) and one commercially available kit, were compared in saliva of pigs. These methods were used in saliva samples obtained from female pigs at 3 different days during gestation and lactation, with and without a reduction/alkylation (R/A), which is a procedure for breaking the links between oxytocin and proteins of the sample. The assays showed a different behavior after the R/A procedure, with no significant changes in the oxytocin results in case of the alphaLISA monoclonal method, a significant decrease with the alphaLISA polyclonal method, and a significant increase with the commercial kit. Although all assays showed a similar tendency in detecting the changes in oxytocin during gestation and lactation, they showed changes of different magnitude and statistical signification. This report indicates that different assays can measure different forms of oxytocin present in saliva and can have a different behavior after R/A of the sample and when are used to measure oxytocin in gestation and lactation., (Copyright © 2020 Elsevier Inc. All rights reserved.)

G protein-coupled receptors (GPCRs) are essential signaling proteins and tractable therapeutic targets. To develop new drug candidates, GPCR drug discovery programs require versatile, sensitive pharmacological tools for ligand binding and compound screening. With the availability of new imaging modalities and proximity-based ligand binding technologies, fluorescent ligands offer many advantages and are increasingly being used, yet labeling small molecules remains considerably more challenging relative to peptides. Focusing on recent fluorescent small molecule studies for family A GPCRs, this review addresses some of the key challenges, synthesis approaches and structure-activity relationship considerations, and discusses advantages of using high-resolution GPCR structures to inform conjugation strategies. While no single approach guarantees successful labeling without loss of affinity or selectivity, the choice of fluorophore, linker type and site of attachment have proved to be critical factors that can significantly affect their utility in drug discovery programs, and as discussed, can sometimes lead to very unexpected results.

Martial arts have become a popular afterschool activity for youths across the globe. Accumulating data suggest that these activities may confer substantial cognitive and psychological benefits, and recent efforts have been made to introduce martial arts training into educational and rehabilitation settings. However, few studies have examined the potential mechanisms that may underlie these benefits. The current study evaluated the reactivity of two hormones, oxytocin (OT) and cortisol (CT), thought to be respectively involved in regulating mammalian social behaviors and responsivity to stress, to a session of intensive martial arts training in samples of at high-risk and low-risk (in regular educational establishments) youths. OT and CT were measured at baseline, during peak training, and following a cool down period. Analyses revealed that high-risk youths had lower OT but similar CT baseline levels, compared to low-risk youths, prior to the martial arts session. A significant group by time interaction indicated that whereas the OT levels among low-risk youths returned to baseline levels following training, OT levels among high-risk youths remained elevated. Finally, unlike low-risk youths for whom CT levels continued to increase throughout the training session, high-risk youths showed no significant CT reactivity. This study suggests that some of the beneficial effects of martial arts may be related to hormonal processes, especially increases in OT levels, and highlights the differing effects that training may have in different populations., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Oxytocin is used for the prevention and treatment of postpartum hemorrhage, the leading cause of maternal mortality in low- and middle-income countries. Because of the high instability of oxytocin, most products are labeled for storage at 2-8°C. Some other products are on the market which are labeled for non-refrigerated storage, but independent evaluations of their stability hardly exist. In the present study, seven brands (nine batches) of oxytocin were purchased from wholesalers and medical stores in Malawi and Rwanda and investigated by accelerated stability testing according to the ICH/WHO guidelines. Two oxytocin brands approved by a stringent regulatory authority (SRA) or by the WHO Prequalification of Medicines program and purchased in Europe were used as comparison. All investigated brands which were either produced in countries with SRAs, or were WHO-prequalified products, were labeled for storage at 2-8°C, and all of them passed stability testing with very good results. Even exposure to 25°C or 30°C for several months hardly affected their oxytocin content. However, two other investigated brands were labeled for non-refrigerated storage, and both of them had been produced in countries without SRAs. These two preparations showed not higher but lower stability than the brands labeled for storage at 2-8°C, and, for both of them, noncompliance with pharmacopoeial specifications was found after accelerated stability testing. At 40°C, and in forced degradation studies at 80°C, chlorobutanol showed a remarkable stabilizing effect on oxytocin, which may deserve further investigation. The results of the present study support the policy "Buy Quality Oxytocin, Keep It Cool."

Oxytocin is a pleiotropic, peptide hormone with broad implications for general health, adaptation, development, reproduction, and social behavior. Endogenous oxytocin and stimulation of the oxytocin receptor support patterns of growth, resilience, and healing. Oxytocin can function as a stress-coping molecule, an anti-inflammatory, and an antioxidant, with protective effects especially in the face of adversity or trauma. Oxytocin influences the autonomic nervous system and the immune system. These properties of oxytocin may help explain the benefits of positive social experiences and have drawn attention to this molecule as a possible therapeutic in a host of disorders. However, as detailed here, the unique chemical properties of oxytocin, including active disulfide bonds, and its capacity to shift chemical forms and bind to other molecules make this molecule difficult to work with and to measure. The effects of oxytocin also are context-dependent, sexually dimorphic, and altered by experience. In part, this is because many of the actions of oxytocin rely on its capacity to interact with the more ancient peptide molecule, vasopressin, and the vasopressin receptors. In addition, oxytocin receptor(s) are epigenetically tuned by experience, especially in early life. Stimulation of G-protein-coupled receptors triggers subcellular cascades allowing these neuropeptides to have multiple functions. The adaptive properties of oxytocin make this ancient molecule of special importance to human evolution as well as modern medicine and health; these same characteristics also present challenges to the use of oxytocin-like molecules as drugs that are only now being recognized. SIGNIFICANCE STATEMENT: Oxytocin is an ancient molecule with a major role in mammalian behavior and health. Although oxytocin has the capacity to act as a "natural medicine" protecting against stress and illness, the unique characteristics of the oxytocin molecule and its receptors and its relationship to a related hormone, vasopressin, have created challenges for its use as a therapeutic drug., (Copyright © 2020 by The Author(s).)

Conopressin, a nonapeptide disulfide CFIRNCPKG amide present in cone snail venom, undergoes a facile cleavage at the Cys6-Pro7 peptide bond to yield a disulfide bridged b 6 ion. Analysis of the mass spectral fragmentation pattern reveals the presence of a major fragment ion, which is unambiguously assigned as the tripeptide sequence IRN amide. The sequence dependence of this unusual fragmentation process has been investigated by comparing it with the fragmentation patterns of related peptides, oxytocin (CYIQNCPLG amide), Lys-vasopressin (CYFQNCPKG amide), and a series of synthetic analogues. The results establish the role of the Arg4 residue in facilitating the unusual N-C α bond cleavage at Cys6. Structures are proposed for a modified disulfide bridged fragment containing the Cys1 and Cys6 residues. Gas-phase molecular dynamics simulations provide evidence for the occurrence of conformational states that permit close approach of the Arg4 side chain to the Cys6 C β methylene protons.

Oxytocin is a hormone of interest in reproduction, but also in the field of psychology and behavior, being considered as a biomarker of positive emotions. Saliva can be a noninvasive way to measure oxytocin, which is very useful in species such as the pig where blood collection can produce a high degree of stress. In this study, a new assay for oxytocin measurement was developed, analytically validated, and used to measure possible changes in oxytocin in saliva of female pigs at different days after farrowing. The assay showed an adequate accuracy and precision and does not need a previous extraction step. In addition, oxytocin concentrations were significantly higher at day 1 of lactation than at day 9 after farrowing, but levels increased at day 20 again. This assay can contribute to a wider use of oxytocin measurements in pigs as it is a noninvasive sampling procedure that minimizes stress., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Positive reinforcement training (PRT) is associated with increases in species-typical behavior and decreases in stereotypic and abnormal behavior in participating animals. Physiological changes following PRT, for example, increases in oxytocin (OXT) and/or decreases in cortisol (CORT), may facilitate these behavioral changes. This study evaluated salivary OXT and salivary CORT concentrations in two adult male western lowland gorillas (Gorilla gorilla gorilla) following PRT with their primary animal care staff. Following PRT, no change in OXT was observed. CORT decreased in one subject following PRT. Changes in endogenous OXT are related to affiliative interactions and interact with strongly bonded conspecifics. PRT may not activate the oxytocinergic system because PRT is not a species-specific affiliative interaction and/or animal care staff are not viewed as conspecifics. Regardless, PRT may still be viewed as a positive interaction resulting in stress reduction via a decrease in CORT. Relationships are unique, thus these results only apply to these two gorillas and one animal caregiver. Larger population-level studies are needed to understand overall trends in human-animal interactions, and ultimately human-animal relationships. Further evaluation of physiological changes following human-animal interactions should be informative for understanding the human-animal relationship in zoos., (© 2019 Wiley Periodicals, Inc.)

The neuropeptides oxytocin (OT) and vasopressin (VP) and their G protein-coupled receptors OTR, V 1a R, V 1b R, and V 2 R form an important and widely-distributed neuroendocrine signaling system. In mammals, this signaling system regulates water homeostasis, blood pressure, reproduction, as well as social behaviors such as pair bonding, trust and aggression. There exists high demand for ligands with differing pharmacological profiles to study the physiological and pathological functions of the individual receptor subtypes. Here, we present the pharmacological characterization of an arthropod (Metaseiulus occidentalis) OT/VP-like nonapeptide across the human OT/VP receptors. I8-arachnotocin is a full agonist with respect to second messenger signaling at human V 2 R (EC 50 34 nM) and V 1b R (EC 50 1.2 µM), a partial agonist at OTR (EC 50 790 nM), and a competitive antagonist at V 1a R [pA 2 6.25 (558 nM)]. Intriguingly, I8-arachnotocin activated the Gα s pathway of V 2 R without recruiting either β-arrestin-1 or β-arrestin-2. I8-arachnotocin might thus be a novel pharmacological tool to study the (patho)physiological relevance of β-arrestin-1 or -2 recruitment to the V 2 R. These findings furthermore highlight arthropods as a novel, vast and untapped source for the discovery of novel pharmacological probes and potential drug leads targeting neurohormone receptors.

Oxytocin is a nonapeptide consisting of a cyclic six amino-acid structure and a tail of three amino acids. It was originally known for its ability to induce milk ejection and to stimulate uterine contractions. More recently, oxytocin has been shown to stimulate social behaviors, and exert pain-relieving, anti-stress/anti-inflammatory and restorative effects. We hypothesize that oxytocin is a principal hormone that, in part, exerts its effects after degradation to active fragments with more specific effect profiles. Experimental findings on rats show that administered oxytocin exerts biphasic effects. For example, after an initial increase in pain threshold, a second more long-lasting increase follows. Blood pressure and cortisol levels initially increase and then reverse into a long-lasting decrease in blood pressure and cortisol. Whereas the initial effects are, the second-phase effects are not blocked by an oxytocin antagonist, but by an opioid mu-antagonist and by an alpha 2-adrenoreceptor antagonist, respectively, suggesting that other receptors are involved. Repeated administration of oxytocin induces multiple anti-stress effects, which are mediated by alpha 2-adrenoreceptors. Repeated administration of linear oxytocin and linear oxytocin fragments with a retained C-terminal reduce spontaneous motor activity, a sedative or anti-stress effect, suggesting that alpha 2-adrenoreceptors have been activated. In contrast, linear mid-fragments stimulate motor activity. Low-intensity stimulation of cutaneous nerves in rats, as well as breastfeeding and skin-to-skin contact between mothers and babies, trigger immediate anti-stress effects. Some of these effects are likely caused by open ring/linear C-terminal fragments activating alpha 2-adrenoreceptors. Oxytocin fragments may be pre-formed and released in the brain or created by metabolic conversion of the principal hormone oxytocin in the central nervous system. Oxytocin and its fragments may also be released from peripheral sites, such as peripheral nerves, the gastrointestinal tract, and blood vessels in response to decreased sympathetic or increased parasympathetic nervous tone. Smaller fragments of oxytocin produced in the periphery may easily pass the blood-brain barrier to induce effects in the brain. In conclusion, oxytocin is linked to many different, sometimes opposite effects. The intact cyclic molecule may act to initiate social interaction and associated psychophysiological effects, whereas linear oxytocin and C-terminal fragments may induce relaxation and anti-stress effects following social interaction. In this way, the principal hormone oxytocin and its fragments may take part in a behavioral sequence, ranging from approach and interaction to calm and relaxation. Linear fragments, with an exposed cysteine-residue, may exert anti-inflammatory and antioxidant effects and thereby contribute to the health-promoting effects of oxytocin., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Oxytocin (OXT) is an important neuromodulator of social behaviors via activation of both oxytocin receptors (OXTR) and vasopressin (AVP) 1a receptors (AVPR1a). Marmosets are neotropical primates with a modified OXT ligand (Pro 8 -OXT), and this ligand shows significant coevolution with traits including social monogamy and litter size. Pro 8 -OXT produces more potent and efficacious responses at primate OXTR and stronger behavioral effects than the consensus mammalian OXT ligand (Leu 8 -OXT). Here, we tested whether OXT/AVP ligands show differential levels of crosstalk at primate AVPR1a. We measured binding affinities and Ca 2+ signaling responses of AVP, Pro 8 -OXT and Leu 8 -OXT at human, macaque, and marmoset AVPR1a. We found that AVP binds with higher affinity than OXT across AVPR1a, and marmoset AVPR1a show a 10-fold lower OXT binding affinity compared to human and macaque AVPR1a. Both Leu 8 -OXT and Pro 8 -OXT produce a less efficacious response than AVP at human AVPR1a and higher efficacious response than AVP at marmoset AVPR1a. These data suggest that OXT might partially antagonize endogenous human AVPR1a signaling and enhance marmoset AVPR1a signaling. These findings aid in further understanding inconsistencies observed following systemic intranasal administration of OXT and provide important insights into taxon-specific differences in nonapeptide ligand/receptor coevolution and behavior.

Protease inhibition has led to treating many diseases and has been successful in producing many commercial drugs by pharmaceutical companies. Among many proteases, serine protease has been attractive in treating metabolic disorder diabetes mellitus (DM). Gliptins have been proven to inhibit dipeptidyl peptidase-4 (DPP4), a serine protease, and are an emerging therapeutic drug target to reduce blood glucose levels, but until now there is no natural cyclic peptide proven to inhibit serine protease DPP4. This study demonstrates the potential mechanism of natural cyclic peptide oxytocin (OXT) as a DPP4 inhibitor. To achieve this, initially, activity atlas and field-based models of DPP4 inhibitors were utilized to predict the possible features of positive and negative electrostatic, hydrophobic, and activity shapes of DPP4 inhibition. Oxytocin binding mode, flexibility, and interacting residues were studied using molecular docking simulations studies. 3D-RISM calculations studies revealed that the stability of water molecules at the binding site are favorable. Finally, an experimental study using fluorescence assay revealed OXT inhibits DPP4 in a concentration-dependent manner in a significant way ( p < 0.05) and possess IC 50 of 110.7 nM. These new findings significantly expand the pharmaceutical application of cyclic peptides, and in specific OXT, and implicate further optimization of OXT inhibition capacity to understand the effect of DPP4 inhibition. This work highlights the development of natural cyclic peptides as future therapeutic peptides to reduce glucose levels and treat diabetes mellitus., Competing Interests: The authors declare no conflict of interest.

In the course of our studies of hydrophobic oxytocin (OT) analogues, we newly synthesized lipidated OT (LOT-4a-c and LOT-5a-c), in which a long alkyl chain (C14-C16) is conjugated via a carbonate or carbamate linkage at the Tyr-2 phenolic hydroxy group and a palmitoyl group at the terminal amino group of Cys-1. These LOTs did not activate OT and vasopressin receptors. Among the LOTs, however, LOT-4c, having a C16-chain via a carbonate linkage at the phenolic hydroxyl group of the Tyr-2, showed very long-lasting action for the recovery of impaired social behavior in CD38 knockout mice, a rodent model of autistic phenotypes, whereas the effect of OT itself rapidly diminished. These results indicate that LOT-4c may serve as a potential prodrug in mice., (Copyright © 2019. Published by Elsevier Ltd.)

Oxytocin plays a vital role in the occurrence of preterm birth by interacting with oxytocin receptor (OTR), and thus OTR antagonists provide effective approaches for the treatment of early birth. Presently, for purpose of exploring the structural traits affecting the antagonism potency, the up-to-date largest set of 121 2,5-diketopiperazine derivatives as OTR antagonists was subjected to ligand-based three-dimensional quantitative structure-activity (3D-QSAR) analysis applying comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) methods. The resultant optimal CoMSIA model displays proper validity and predictability with cross-validated correlation coefficient Q 2  = 0.614, non-cross-validated correlation coefficient R 2 ncv  = 0.969 and predicted correlation coefficient R 2 pre  = 0.912 for the test set of compounds, respectively. In addition, docking study was carried out for further elucidating the binding modes of OTR antagonists. The final docking cavity was located among the TM2-TM7 helices of the target protein and proved to be the same as described by other scholars' researches of other type of OTR antagonists. The major amino acids forming the cavity are Q92, Q96, K116, Q119, V120, M123, G196, I201, Q295, W288, F311 and M315. Representative compound 118 forms two H-bonds with Q119 and two H-bonds with Q295, with also a π - π stacking effect with F311, respectively. Meanwhile, as a supplementary tool to study the antagonists' conformation in the binding cavity, molecular dynamics (MD) simulation was also performed to further elucidate the changes in the ligand-receptor complex. We hope that the obtained models and information may help to provide an insight into the interaction mechanism of OTR antagonists and facilitate the design and optimization of novel antagonists for preventing premature birth., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The neurohypophyseal hormone oxytocin (OT) and related modulators of the oxytocin receptor (OTR) have been the subject of intensive research for nearly seven decades. Despite having rather poor drug-like properties, OT is used as a treatment for labor induction, postpartum hemorrhage, and lactation support. The potential use of OT in the treatment of central nervous system (CNS)-related diseases has recently renewed interest in the pharmacology of OT. Oxytocin is one of the most extensively studied cyclic peptides and since the elucidation of its structure in 1953 thousands of peptidic OT analogs with antagonistic and agonistic properties have been synthesized and biologically evaluated. Among them are atosiban, a mixed oxytocin receptor (OTR)/vasopressin 1a receptor (V 1a R) antagonist used as a tocolytic agent approved (in certain countries), and carbetocin, a longer acting OTR agonist on the market for the treatment of postpartum hemorrhage. Many other OT analogs with improved pharmacological properties (e.g., barusiban, Antag III) have been identified. These peptides have been tested in clinical trials and/or used as pharmacological tools. In this chapter, the modifications of the OT molecule that led to the discovery of these compounds are reviewed.

Currently, there are no established pharmaceutical strategies that effectively treat social deficits in autism spectrum disorder (ASD). Oxytocin, a neurohormone that plays a role in multiple types of social behaviors, has been proposed as a possible therapeutic against social impairment and other symptoms in ASD. However, from the standpoint of pharmacotherapy, oxytocin has several liabilities as a standard clinical treatment, including rapid metabolism, low brain penetrance, and activity at the vasopressin (antidiuretic hormone) receptors. The present studies describe findings from a preclinical screening program to evaluate oxytocin receptor (OXTR) agonists and oxytocin metabolites for potential clinical use as more optimal treatments. We first investigated two synthetic oxytocin analogs, TC-OT-39 and carbetocin, using in vitro cell-based assays for pharmacological characterization and behavioral tests in the BALB/cByJ mouse model of ASD-like social deficits. Although both TC-OT-39 and carbetocin selectively activate the OXTR, neither synthetic agonist had prosocial efficacy in the BALB/cByJ model. We next evaluated two oxytocin metabolites: OT(4-9) and OT(5-9). While OT(5-9) failed to affect social deficits, the metabolite OT(4-9) led to significant social preference in the BALB/cByJ model, in a dose-dependent manner. The increased sociability was observed at both 24 h and 12 days following the end of a subchronic regimen with OT(4-9) (2.0 mg/kg). Overall, these results suggest that the prosocial effects of oxytocin could be mediated by downstream activity of oxytocin metabolites, raising the possibility of new pathways to target for drug discovery relevant to ASD., (Copyright © 2018. Published by Elsevier Ltd.)

The study of energy landscapes has led to a good understanding of how and why proteins and nucleic acids adopt their native structure. Through evolution, sequences have adapted until they exhibit a strongly funneled energy landscape, stabilizing the native fold. Design of artificial biomolecules faces the challenge of creating similar stable, minimally frustrated, and functional sequences. Here we present a biminimization approach, mutational basin-hopping, in which we simultaneously use global optimization to optimize the energy and a target function describing a desired property of the system. This optimization of structure and sequence is a generalized basin-hopping method and produces an efficient design process, which can target properties such as binding affinity or solubility.

Oxytocin (OT) is a hypothalamic nonapeptide that mediates a host of physiological and behavioral processes including reproductive physiology and social attachments. While the OT sequence structure is highly conserved among mammals, New World monkeys (NWMs) represent an unusual "hot spot" in OT structure variability among mammals. At least 6 distinct OT ligand variants among NWMs exist, yet it is currently unclear whether these evolved structural changes result in meaningful functional consequences. NWMs offer a new area to explore how these modifications to OT and its canonical G-protein coupled OT receptor (OTR) may mediate specific cellular, physiological and behavioral outcomes. In this review, we highlight relationships between OT ligand and OTR structural variability, specifically examining coevolution between OT ligands, OTRs, and physiological and behavioral phenotypes across NWMs. We consider whether these evolved modifications to the OT structure alter pharmacological profiles at human and marmoset OTRs, including changes to receptor binding, intracellular signaling and receptor internalization. Finally, we evaluate whether exogenous manipulation using OT variants in marmoset monkeys differentially enhance or impair behavioral processes involved in social relationships between pairmates, opposite-sex strangers, and parents and their offspring. Overall, it appears that changes to OT ligands in NWMs result in important changes ranging from cellular signaling to broad measures of social behavior., (© 2018 International Society of Zoological Sciences, Institute of Zoology/Chinese Academy of Sciences and John Wiley & Sons Australia, Ltd.)

Background: Oxytocin is the gold standard drug for the prevention of postpartum haemorrhage, but limitations in cold chain systems in resource-constrained settings can severely compromise the quality of oxytocin product available in these environments. This study investigated the perspectives and practices of stakeholders in low and lower-middle income countries towards oxytocin, its storage requirements and associated barriers, and the quality of product available., Methods: Qualitative inquiries were undertaken in Ethiopia, India and Myanmar, where data was collected through Focus Group Discussions (FGDs) and In-Depth Interviews (IDIs). A total of 12 FGDs and 106 IDIs were conducted with 158 healthcare providers (pharmacists, midwives, nurses, doctors and obstetricians) and 40 key informants (supply chain experts, program managers and policy-makers). Direct observations of oxytocin storage practices and cold chain resources were conducted at 51 healthcare facilities. Verbatim transcripts of FGDs and IDIs were translated to English and analysed according to a thematic content analysis framework., Findings: Stakeholder awareness of oxytocin heat sensitivity and the requirement for cold storage of the drug was widespread in Ethiopia but more limited in Myanmar and India. A consistent finding across all study regions was the significant barriers to maintaining a consistent cold chain, with the lack of refrigeration facilities and unreliability of electricity cited as major challenges. Perceptions of compromised oxytocin quality were expressed by some stakeholders in each country., Conclusion: Knowledge of the heat sensitivity of oxytocin and the potential impacts of inconsistent cold storage on product quality is not widespread amongst healthcare providers, policy makers and supply chain experts in Myanmar, Ethiopia and India. Targeted training and advocacy messages are warranted to emphasise the importance of cold storage to maintain oxytocin quality., Competing Interests: We have the following interests: VLO, PL and MPM are part of a product development team at Monash University, which is progressing the development of a heat stable oxytocin product for the prevention of PPH in resource-poor settings. Inhaled oxytocin is being developed through a product development collaboration agreement between Monash University and GlaxoSmithKline (GSK). GSK had no role in the funding, design or conduct of this study. AT is the founder and CEO of a for-profit research organisation (MERQ consultancy). For the conduct of this study, MT and AM were contracted by MERQ consultancy to provide services as outlined in the ‘author contribution’ section. MPM is the co-inventor of a worldwide patent application ‘Method and Formulation for Inhalation’ (WO 2013/016754) that covers the delivery of biologically active agents (including oxytocin) in the form of dry powders for inhalation. The authors have no commercial interest in the outcomes of this study or the introduction of the inhaled oxytocin product in low and lower-middle income countries. There are no further patents, products in development or marketed products relevant to this research study to declare. This does not alter our adherence to all the PLOS ONE policies on sharing data and materials."

Oxytocin is a promising candidate for the treatment of social-deficit disorders such as Autism Spectrum Disorder, but oxytocin cannot readily pass the blood-brain barrier. Moreover, oxytocin requires frequent dosing as it is rapidly metabolized in blood. We fabricated four polymeric nanoparticle formulations using poly(lactic-co-glycolic acid) (PLGA) or bovine serum albumin (BSA) as the base material. In order to target them to the brain, we then conjugated the materials to either transferrin or rabies virus glycoprotein (RVG) as targeting ligands. The formulations were characterized in vitro for size, zeta potential, encapsulation efficiency, and release profiles. All formulations showed slightly negative charges and sizes ranging from 100 to 278 nm in diameter, with RVG-conjugated BSA nanoparticles exhibiting the smallest sizes. No formulation was found to be immunogenic or cytotoxic. The encapsulation efficiency was ≥75% for all nanoparticle formulations. Release studies demonstrated that BSA nanoparticle formulation exhibited a faster initial burst of release compared to PLGA particles, in addition to later sustained release. This initial burst release would be favorable for clinical dosing as therapeutic effects could be quickly established, especially in combination with additional sustained release to maintain the therapeutic effects. Our size and release profile data indicate that RVG-conjugated BSA nanoparticles are the most favorable formulation for brain delivery of oxytocin., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The nonapeptide oxytocin (OT) is used as a model sulfur-containing peptide to study the damage induced by vacuum UV (VUV) radiations. In particular, the effect of the presence (or absence in reduced OT) of oxytocin's internal disulfide bridge is evaluated in terms of photo-fragmentation yield and nature of the photo-fragments. Intact, as well as reduced, OT is studied as dianions and radical anions. Radical anions are prepared and photo-fragmented in two-color experiments (UV + VUV) in a linear ion trap. VUV photo-fragmentation patterns are analyzed and compared, and radical-induced mechanisms are proposed. The effect of VUV is principally to ionize but secondary fragmentation is also observed. This secondary fragmentation seems to be considerably enabled by the initial position of the radical on the molecule. In particular, the possibility to form a radical on free cysteines seems to increase the susceptibility to VUV fragmentation. Interestingly, disulfide bridges, which are fundamental for protein structure, could also be responsible for an increased resistance to ionizing radiations. Graphical Abstract.

Introduction: Separation after preterm birth is a major stressor for infants and parents. Skin-to-skin contact (SSC) is a method of care suitable to use in the neonatal intensive care unit (NICU) to minimise separation between parents and infants. Less separation leads to increased possibilities for parent-infant interaction, provided that the parents' sleep quality is satisfactory. We aimed to evaluate the effect of continuous SSC on sleep quality and mood in parents of preterm infants born <33 weeks of gestation as well as the quality of parent-infant interaction and salivary cortisol concentrations at the time of discharge., Methods and Analysis: A randomised intervention study with two arms-intervention versus standard care. Data will be collected from 50 families. Eligible families will be randomly allocated to intervention or standard care when transferred from the intensive care room to the family-room in the NICU. The intervention consists of continuous SSC for four consecutive days and nights in the family-room. Data will be collected every day during the intervention and again at the time of discharge from the hospital. Outcome measures comprise activity tracker (Actigraph); validated self-rated questionnaires concerning sleep, mood and bonding; observed scorings of parental sensitivity and emotional availability and salivary cortisol. Data will be analysed with pairwise, repeated measures, Mann Whitney U-test will be used to compare groups and analysis of variance will be used to adjust for different hospitals and parents' gender., Ethics and Dissemination: The study is approved by the Regional Research Ethics Board at an appropriate university (2016/89-31). The results will be published in scientific journals. We will also use conferences and social media to disseminate our findings., Trial Registration Number: NCT03004677., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2018. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Role of estrogen and photoperiod is well-established in avian reproduction. In addition, the distribution and the expression of arginine vasotocin (AVT) and its receptor VT3 to ensure reproductive/breeding conditions in Japanese quail influenced by them has been the main focus of this review. To consider this aspect the mRNA expression of VT3 receptor and its ligand AVT in the shell gland has been emphasized. In birds, AVT performs a dual role as an anti-diuretic hormone and the functions accomplished by oxytocin in mammals. The physiological actions of AVT in birds are mediated through its diverse receptor subtypes VT1, VT2, VT3 and VT4.  Dynamic alteration of VT3 expression during different reproductive and photo-sexual conditions of quail can be modulated by estrogen. In addition to the endocrine effect of AVT, the shell gland is complemented by its paracrine action via its receptors. Evidences indicate that the expression of shell gland AVT modulated by estrogen appears to play a priming role by modulating the availability of VT3 receptor for the required action of neurohypophysial AVT during oviposition.

Postpartum haemorrhage is a leading cause of maternal death worldwide. Oxytocin, currently the drug of choice for prevention of PPH, requires constant refrigeration. In pursuit of an alternative medicine, Ferring Pharmaceuticals have developed a heat-stable formulation of carbetocin, an oxytocin analogue. This study aimed to define that formulation, and to investigate its stability under ICH climate zone IV conditions (30°C/75% relative humidity) for at least 3 years and at extreme temperatures, such as 60°C, for shorter periods of time. The development resulted in a heat-stable carbetocin formulation consisting of 0.1 mg/mL carbetocin in sodium succinate buffer, mannitol, and methionine. The optimum pH was determined to be pH 5.45 (5.25-5.65). The generated stability data of this formulation show that ≥95% purity of the peptide was maintained for a minimum of 3 years at 30°C, 6 months at 40°C, 3 months at 50°C and 1 month at 60°C. In addition, the heat-stable carbetocin formulation was not sensitive to freezing or light. The reported highly stable peptide formulation facilitates the distribution in low and middle-income countries, where maintaining cold chain distribution is difficult. Ferring Pharmaceuticals, the World Health Organization, and MSD for Mothers have established a collaboration to develop this heat-stable formulation of carbetocin for the prevention of post-partum hemorrhage in women after vaginal childbirth, with the aim of making the medicine available in the public sector of developing countries that have a high burden of maternal mortality., (© 2018 The Authors. Journal of Peptide Science published by European Peptide Society and John Wiley & Sons Ltd.)

The goal of this work was to investigate a new nasal carrier for enhanced drug delivery to brain, we call Phospholipid Magnesome. The system contains soft phospholipid vesicles, composed of phospholipid, water, propylene glycol, magnesium salt, and the mucoadhesive polymer, alginate. The carrier was characterized by various methods: electron microscopy, calorimetry, and dynamic light scattering. The ability of the carrier's vesicles to entrap various molecules was studied by CLSM and ultracentrifugation combined with HPLC quantification. Mucoadhesivity of the carrier was tested in vitro using porcine nasal mucosa. The delivery of rohdamine 6G, insulin, and epidermal growth factor was estimated by two methods, multiphoton microscopy and near infrared (NIR) imaging. Pharmacodynamic effects of nasal treatment with oxytocin and insulin incorporated in Phospholipid Magnesome were evaluated in animal models. Results show that the system is composed of soft multilamellar nanosized vesicles with the ability to entrap both lipophilic and hydrophilic molecules. The mucoadhesivity test results indicate a prolonged contact time of the drug with the nasal membrane as compared to control. Multiphoton microscopy and NIR imaging of brain show effective delivery of the tested molecules to brain following nasal administration in Phospholipid Magnesome relative to controls. Moreover, the results of the pharmacodynamic study measuring the antinociceptive effect of oxytocin administrated nasally to an animal model indicate the efficiency of the Phospholipid Magnesome as compared to three control systems. Further, nasal administration of insulin resulted in a strong and prolonged hypoglycemic effect for the drug incorporated in the new carrier but not for control systems. Based on the results of the histopathological test, the carrier is safe for local administration on the nasal membrane. In conclusion, the results of this study suggest that Phospholipid Magnesome nasal carrier is able to improve drug effects, probably by a combined mechanism, absorption enhancement, and prolongation of mucosal contact.

Many peptides with antimicrobial activity and/or therapeutic potential contain disulfide bonds as a means to enhance stability, and their quantitation is often performed using electrospray ionization mass spectrometry (ESI-MS). Disulfides can be reduced during ESI under commonly used instrument conditions, which has the potential to hinder accurate peptide quantitation. We demonstrate that this in-source reduction (ISR) is predominantly observed for peptides infused from acidic solutions and subjected to elevated ESI voltages (3-4 kV). ISR is readily apparent in the mass spectrum of oxytocin-a small, single disulfide-containing peptide. However, subtle m/z shifts due to partial ISR of highly charged (z ≥ 3) peptides with multiple disulfide linkages may proceed unnoticed. Ion mobility (IM)-MS separates ions on the basis of charge and shape in the gas phase, and using insulin as a model system, we show that IM-MS arrival time distributions (ATDs) are particularly sensitive to partial ISR of large peptides. Isotope modeling allows for the relative quantitation of disulfide-intact and partially reduced states of the mobility-separated peptide conformers. Interestingly, hepcidin peptides ionized from acidic solutions at elevated ESI voltages undergo gas-phase compaction, ostensibly due to partial disulfide ISR. Our IM-MS results lead us to propose that residual acid is the likely cause of disparate ATDs recently measured for hepcidin from different suppliers [Anal. Bioanal. Chem. 409, 2559-2567 (2017)]. Overall, our results demonstrate the utility of IM-MS to detect partial ISR of disulfide-bonded peptides and reinforce the notion that peptide/protein measurements should be carried out using minimally activating instrument conditions. Graphical Abstract ᅟ.

Peptides are usually administered through subcutaneous injection. For low potency drugs, this may require high concentration formulations increasing the risk of peptide aggregation, especially for compounds without any intrinsic chargeable groups. Carbetocin was used as a model to study the behavior of uncharged peptides at high concentrations. Manipulation of the aggregation behavior of 70 mg/mL carbetocin was attempted by selecting excipients which interact with hydrophobic groups in carbetocin, and cover hydrophobic surfaces and interfaces. Peptide aggregation was induced by shaking stress and followed over time. Carbetocin solutions showed significant visible particle formation already after 4 h of shaking stress. This particle formation was not due to supersaturation or phase separation but suggested a nucleated aggregation process. None of the excipients prevented carbetocin aggregation, though altered aggregation behavior was observed, such as induction of fibril formation for most, but not all, charged excipients. Sodium dodecyl sulfate was found to accelerate peptide aggregation both below and above the critical micelle concentration in half-filled vials. However, in the absence of an air headspace, sodium dodecyl sulfate above the critical micelle concentration was capable of preventing shaking-induced carbetocin aggregation. Our study highlights the complexity in rational excipient selection to stabilize uncharged peptides at high concentration., (Copyright © 2018 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.)

Oxytocin neurons of the rat hypothalamus project to the posterior pituitary, where they secrete their products into the bloodstream. The pattern and quantity of that release depends on the afferent inputs to the neurons, on their intrinsic membrane properties, and on nonlinear interactions between spiking activity and exocytosis: A given number of spikes will trigger more secretion when they arrive close together. Here we present a quantitative computational model of oxytocin neurons that can replicate the results of a wide variety of published experiments. The spiking model mimics electrophysiological data of oxytocin cells responding to cholecystokinin (CCK), a peptide produced in the gut after food intake. The secretion model matches results from in vitro experiments on stimulus-secretion coupling in the posterior pituitary. We mimic the plasma clearance of oxytocin with a two-compartment model, replicating the dynamics observed experimentally after infusion and injection of oxytocin. Combining these models allows us to infer, from measurements of oxytocin in plasma, the spiking activity of the oxytocin neurons that produced that secretion. We have tested these inferences with experimental data on oxytocin secretion and spiking activity in response to intravenous injections of CCK. We show how intrinsic mechanisms of the oxytocin neurons determine this relationship: In particular, we show that the presence of an afterhyperpolarization (AHP) in oxytocin neurons dramatically reduces the variability of their spiking activity and even more markedly reduces the variability of oxytocin secretion. The AHP thus acts as a filter, protecting the final product of oxytocin cells from noisy fluctuations.

BackgroundHuman milk has a high content of the antimicrobial compound hydrogen peroxide (H 2 O 2 ). As opposed to healthy full-term infants, preterm neonates are fed previously expressed and stored maternal milk. These practices may favor H 2 O 2 decomposition, thus limiting its potential benefit to preterm infants. The goal of this study was to evaluate the factors responsible for H 2 O 2 generation and degradation in breastmilk.MethodsHuman donors' and rats' milk, along with rat mammary tissue were evaluated. The role of oxytocin and xanthine oxidase on H 2 O 2 generation, its pH-dependent stability, as well as its degradation via lactoperoxidase and catalase was measured in milk.ResultsBreast tissue xanthine oxidase is responsible for the H 2 O 2 generation and its milk content is dependent on oxytocin stimulation. Stability of the human milk H 2 O 2 content is pH-dependent and greatest in the acidic range. Complete H 2 O 2 degradation occurs when human milk is maintained, longer than 10 min, at room temperature and this process is suppressed by lactoperoxidase and catalase inhibition.ConclusionFresh breastmilk H 2 O 2 content is labile and quickly degrades at room temperature. Further investigation on breastmilk handling techniques to preserve its H 2 O 2 content, when gavage-fed to preterm infants is warranted.

Diabetes mellitus (DM) is associated with increased plasma levels of arginine-vasopressin (AVP), which may aggravate hyperglycemia and nephropathy. However, the mechanisms by which DM may cause the increased AVP levels are not known. Electrophysiological recordings in supraoptic nucleus (SON) slices from streptozotocin (STZ)-induced DM rats and vehicle-treated control rats revealed that γ-aminobutyric acid (GABA) functions generally as an excitatory neurotransmitter in the AVP neurons of STZ rats, whereas it usually evokes inhibitory responses in the cells of control animals. Furthermore, Western blotting analyses of Cl - transporters in the SON tissues indicated that Na + -K + -2Cl - cotransporter isotype 1 (a Cl - importer) was upregulated and K + -Cl - cotransporter isotype 2 (KCC2; a Cl - extruder) was downregulated in STZ rats. Treatment with CLP290 (a KCC2 activator) significantly lowered blood AVP and glucose levels in STZ rats. Last, investigation that used rats expressing an AVP-enhanced green fluorescent protein fusion gene showed that AVP synthesis in AVP neurons was much more intense in STZ rats than in control rats. We conclude that altered Cl - homeostasis that makes GABA excitatory and enhanced AVP synthesis are important changes in AVP neurons that would increase AVP secretion in DM. Our data suggest that Cl - transporters in AVP neurons are potential targets of antidiabetes treatments., (© 2017 by the American Diabetes Association.)

Disulfide bonds are among the most important factors related to correct folding of the proteins. Protein disulfide isomerase (PDI) is the enzyme responsible for the correct formation and isomerization of these bonds. It is rarely studied so far and none of them showed industrial properties. In this study, the gene encoding for a putative PDI from Bacillus subtilis DR8806 was identified, cloned and expressed in Escherichia coli. It was encoded a 23.26kDa protein. The enzyme was purified by GST affinity chromatography with a specific activity of 1227u/mg. It was active and stable over a wide range of temperature (20-85°C) and pH (4.5-10) with an optimum at 65°C and pH 5.5. Its activity was enhanced by Mn 2+ and Co 2+ while Ag + and Zn 2+ decreased it. Some of the known PDI inhibitors such as Tocinoic acid and Bactiracin did not affect its activity. In-silico analysis shows the five amino acids changes in the protein sequence regarding to the consensus sequence of PDIs, have a positive impact toward the protein thermal stability. This was further confirmed by molecular dynamics simulations. By considering the overall results, the enzyme might be a potential candidate for applications in the respective industries., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Oxytocin (OT) is a peptide hormone agonist of the oxytocin receptor (OTR) that has been proposed as a therapeutic to treat a number of social and emotional disorders in addition to its current clinical use to induce labor and treat postpartum bleeding. OT is administered intravenously and intranasally rather than orally, in part because its low passive permeability causes low oral bioavailability. Non-peptidic OTR agonists have also been reported, but none with the exquisite potency of the peptide based agonists. In this report, we describe the OTR agonist activity and exposed polarity of a set of truncated OT analogs as well as hybrid peptide-small molecule analogs of OT. Examples of both truncated analogs and peptide-small molecule hybrid analogs are potent and selective OTR agonists. Hybrid agonist 13, which is 232 Da smaller than OT, still retains subnanomolar potency, full agonist activity, and selectivity over V1a. While these compounds were designed to address the low permeability of OT and other full length analogs, we found that reduction in molecular weight and the removal or replacement of the three amino acid tail of OT did not have a significant effect on passive permeability., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Oxytocin (OT), a neuropeptide produced primarily in the hypothalamus, is associated with both critical physiological and psychological processes, particularly stress and feelings of affiliation. Increasingly, researchers are seeking ways to reliably incorporate OT as an outcome biomarker in clinical research. Previously, OT levels were measured in plasma or urine. Recently, researchers have measured this biomarker in saliva, particularly when conducting research in clinical and community settings. In spite of increased interest in the use of salivary OT in clinical research, procedures for handling, transport, and analysis of specimens vary. It is not known if significant OT protein degradation occurs if samples are initially transported on wet ice before being frozen. The aim of this study is to evaluate the effect of transport media (wet vs. dry ice) on OT levels derived from saliva collected from 12 postpartum women residing in the community. Saliva collected from each participant was divided between two microcentrifuge tubes (MIDSCI, Valley Park, MO), one placed on wet ice and one on dry ice for transport from the participant's home to the laboratory freezer. Time from collection to storage freezer was recorded. Laboratory personnel, blinded to method of transport, batch processed the samples. No significant differences in OT levels were found by transport method. Despite large interperson variations in OT levels, there were negligible intraperson variations. Although further research is required to identify factors (including transport time) related to interperson variation, this study supports the use of wet ice as a means of transporting salivary OT specimens in community-based research.

Objective: To examine changes that occur in infant and parent salivary oxytocin (OT) and salivary cortisol (SC) levels during skin-to-skin contact (SSC) and whether SSC alleviates parental stress and anxiety while also supporting mother-father-infant relationships., Methods: This randomized crossover study was conducted in the neonatal intensive care unit (NICU) with a sample of 28 stable preterm infants and their parents. Saliva samples were collected from infants, mothers, and fathers on Days 1 and 2 (1/parent) for OT and cortisol measurement pre-SSC, during a 60-min SSC session, and a 45-min post-SSC. Parental anxiety was measured at the same time points. Parent-infant interaction was examined prior to discharge on Day 3 via video for synchrony and responsiveness using Dyadic Mutuality Coding., Results: Salivary OT levels increased significantly during SSC for mothers ( p < .001), fathers ( p < .002), and infants ( p < .002). Infant SC levels decreased significantly ( p < .001) during SSC as compared to before and after SSC. Parent anxiety scores were significantly related to parent OT and SC levels. Parents with higher OT levels exhibited more synchrony and responsiveness ( p < .001) in their infant interactions., Conclusion: This study addresses a gap in understanding the mechanisms linking parent-infant contact to biobehavioral responses. SSC activated OT release and decreased infant SC levels. Facilitation of SSC may be an effective intervention to reduce parent and infant stress in the NICU. Findings advance the exploration of OT as a potential moderator for improving responsiveness and synchrony in parent-infant interactions.

Leachables derived from multi-component drug-device syringe systems can result in changes to the quality of drug products. Diphenylguanidine (DPG), a leachable released from styrene butadiene rubber syringe plungers, interacts with Oxytocin to form protein-adducts. This study investigated the mechanism and kinetics of this interaction in both solid and solution states through in-vitro tests and spectroscopic methods For solid state interaction, the protein-adducts with DPG were characterized using SEM, XRD, DSC, FTIR, 13 C ss NMR, and dissolution analysis. For solution state interaction, LC-HRMS was used to assess stability of Oxytocin solutions in presence of various concentrations of DPG at 25°C and 40°C for 4 weeks. Moreover, molecular docking analysis was used to identify possible molecular configurations of the interaction.Results were consistent with the formation of a new solid state with distorted surface morphology for oxytocin-DPG adducts, in which the oxytocin carbonyl group(s) and the secondary amine groups of DPG interact. This interaction was also confirmed by molecular docking analysis through hydrogen bonding (2.31Å) and Van der Waal attraction (3.14Å). Moreover, LC-HRMS analysis revealed an increase in Oxytocin stability and suppression of Oxytocin dimerization by DPG. A potential reduction in the rate of Oxytocin dissolution from the formed adducts was indicative of its strong association with DPG. Hence, the leaching potential of DPG from rubber closures and plungers should be monitored and controlled to maintain the quality and stability of the pharmaceutical product., (Published by Elsevier B.V.)

C-terminally modified peptides are important targets for pharmaceutical and biochemical applications. Known methods for C-terminal diversification are limited mainly in terms of the scope of accessible modifications or by epimerization of the C-terminal amino acid. In this work, we present a broadly applicable approach that enables access to a variety of C-terminally functionalized peptides in either protected or unprotected form. This chemistry proceeds without epimerization of C-terminal Ala and tolerates nucleophiles of varying nucleophilicity. Finally, unprotected peptides bearing nucleophilic side chain groups can be selectively functionalized by strong nucleophiles, whereas macrocyclization is observed for weaker nucleophiles. The potential utility of this method is demonstrated through the divergent synthesis of the conotoxin conopressin G and GLP-1(7-36) and analogs., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The purpose of the study described in this Research Communication was to report the full characterisation of the goat and sheep oxytocin-neurophysin I gene (OXT), their promoters and amino acid sequences. Using the genomic DNA as template, we sequenced and compared the whole OXT gene (3 exons), plus 958/960 nucleotides at the 5' flanking region and 478/477 nucleotides at the 3' flanking region, in 46 sheep and 24 goats belonging to different breeds/genetic types reared in Italy, Greece and Germany. The comparison of the obtained sequences showed a high degree of genetic variability at these loci. In particular, we focused on the SNP g.438T > C as possible example of trans-specific polymorphism. This SNP alters a putative binding site of the transcription factor Oct-1. The set-up of a luciferase assay confirmed that the C variant of this SNP negatively affects the promoter activity of the sheep OXT gene. The results of this study suggest that the SNP g.438T > C might be useful to promote association studies with traits/physiological processes controlled by this hormone.

The article presents an overview of the comparative distribution, structure and functions of the nonapeptide hormones in chordates and non chordates. The review begins with a historical preview of the advent of the concept of neurosecretion and birth of neuroendocrine science, pioneered by the works of E. Scharrer and W. Bargmann. The sections which follow discuss different vertebrate nonapeptides, their distribution, comparison, precursor gene structures and processing, highlighting the major differences in these aspects amidst the conserved features across vertebrates. The vast literature on the anatomical characteristics of the nonapeptide secreting nuclei in the brain and their projections was briefly reviewed in a comparative framework. Recent knowledge on the nonapeptide hormone receptors and their intracellular signaling pathways is discussed and few grey areas which require deeper studies are identified. The sections on the functions and regulation of nonapeptides summarize the huge and ever increasing literature that is available in these areas. The nonapeptides emerge as key homeostatic molecules with complex regulation and several synergistic partners. Lastly, an update of the nonapeptides in non chordates with respect to distribution, site of synthesis, functions and receptors, dealt separately for each phylum, is presented. The non chordate nonapeptides share many similarities with their counterparts in vertebrates, pointing the system to have an ancient origin and to be an important substrate for changes during adaptive evolution. The article concludes projecting the nonapeptides as one of the very first common molecules of the primitive nervous and endocrine systems, which have been retained to maintain homeostatic functions in metazoans; some of which are conserved across the animal kingdom and some are specialized in a group/lineage-specific manner., (Copyright © 2016 Elsevier Inc. All rights reserved.)