Oxytocin In Rats Research Articles

Short- and long-term consequences of different early environmental conditions on central immunoreactive oxytocin and arginine vasopressin levels in male rats

Numerous studies have provided evidence for an important role for the neuropeptides oxytocin (OT) and arginine vasopressin (AVP) in establishment of social behaviour early in life, such as mother-pup interactions. However, there are few reports examining the consequences of early-life experiences on OT and AVP in male offspring. We have used the maternal separation (MS) model to study the effect of different early environmental conditions in rats. The purpose was to study OT and AVP in rats subjected to prolonged daily MS (360 min, MS360), short daily MS (15 min, MS15) and conventional animal facility rearing (AFR) during postnatal days 1-21. In addition, the influence of the presence or absence of littermates during MS, i.e. litter-wise (l) or individual (i) MS, was assessed. The immunoreactive (ir) peptide levels were measured in the hypothalamus, amygdala and pituitary gland of 3 and 10 weeks old male rats. Assessment in 3-week-old rats revealed that MS15 was associated with low ir OT levels in the hypothalamus and amygdala and high levels in the pituitary gland compared with the MS360 and AFR condition. In the amygdala, differences between groups were also detected in adulthood. MS studies commonly use either MS15 or AFR as a control for prolonged MS. The present results show differences in MS360 rats as compared to MS15 but not AFR rats. Consequently, comparisons between prolonged MS with either short periods of MS or AFR will generate divergent results, hence, making the outcome of MS difficult to compare between studies. Moreover, the different early environments had no effect on ir AVP levels. In conclusion, OT in the amygdala was most sensitive to MS. Besides both short- and long-term consequences, distinct effects were seen after litter and individual separation, respectively. We propose that environmentally induced alterations in OT transmission due to disrupted mother-pup interactions early in life may cause altered susceptibility to challenges later in life.

Effect of intratesticular administration of oxytocin on testicular steroidogenesis in immature rats.

The possible physiological role of testicular oxytocin in testicular steroidogenesis was studied in immature rats. In 5-, 9-, and 25-day-old animals one of the testes was injected with 20, 50, and 200 ng of oxytocin, respectively, then the contralateral gonad was removed. Rats were killed 1 h, 1 day, or 7 days post-surgery. In each age group studied intratesticular injection of oxytocin resulted in a significant increase in basal testosterone secretion in vitro and/or serum testosterone concentration. Data indicate that in immature rats oxytocin of testicular origin might act as a local stimulator of steroidogenesis.

Effects of cholecystokinin and oxytocin in rats trained to discriminate 22 from 2 hours food deprivation

Cholecystokinin and oxytocin have previously been shown to decrease food intake in rats. We tested the effects of cholecystokinin and oxytocin, in rats trained to discriminate 22 hr food deprivation from 2 hr food deprivation in a two‐lever, operant choice task. After rats acquired the discrimination, subjects were food restricted for 22 hr and administered cholecystokinin(1‐10 µg/kg,i.p.) or oxytocin (0.01‐3.2 mg/kg, i.p.) before generalization tests. Food intake was measured for 1 hr after the generalization test. Cholecystokinin dose‐dependently decreased the discriminative stimulus effects of 22 hr deprivation. Oxytocin did not affect significantly affect the discriminative effects of deprivation, but significantly reduced response rate and food intake. The findings support the notion that CCK affects feeding stimulated by energy needs, whereas oxytocin affects feeding via other mechanisms.

Effects of neonatal handling on social memory, social interaction, and number of oxytocin and vasopressin neurons in rats

Early-life environmental events can induce profound long-lasting changes in several behavioral and neuroendocrine systems. The neonatal handling procedure, which involves repeated brief maternal separations followed by experimental manipulations, reduces stress responses and sexual behavior in adult rats. The purpose of this study was to analyze the effects of neonatal handling on social behaviors of male and female rats in adulthood, as manifest by the results of social memory and social interaction tests. The number of oxytocin (OT) and vasopressin (VP) neurons in the paraventricular (PVN) and supraoptic (SON) nuclei of hypothalamus were also analyzed. The results did not demonstrate impairment of social memory. Notwithstanding, handling did reduce social investigative interaction and increase aggressive behavior in males, but did not do so in females. Furthermore, in both males and females, handling was linked with reduced number of OT-neurons in the parvocellular region of the PVN, while no differences were detected in the magnocellular PVN or the SON. On the other hand, handled males exhibited increased number of VP-neurons in the magnocellular zone of the PVN. We may conclude that the repeated brief maternal separations can reduce affiliative social behavior in adult male rats. Moreover, the disruption of the mother–infant relationship caused by the handling procedure induced long-lasting morphological changes in critical neuroendocrine areas that are involved in social bonding in mammals.

Vasopressin Administration into the Paraventricular Nucleus Normalizes Plasma Oxytocin and Corticosterone Levels in Brattleboro Rats

Adult male rats of the Brattleboro strain were used to investigate the impact of the congenital absence of vasopressin on plasma adrenocorticotropin, corticosterone, and oxytocin concentrations as well as the release pattern of oxytocin within the hypothalamic paraventricular nucleus (PVN), in response to a 10-min forced swimming session. Measurement of adrenocorticotropin in plasma samples collected via chronically implanted jugular venous catheters revealed virtually identical stress responses for vasopressin-lacking Brattleboro (KO) and intact control animals. In contrast, plasma corticosterone and oxytocin levels were found to be significantly elevated 105 min after onset of the stressor in KO animals only. Microdialysis samples collected from the extracellular fluid of the PVN showed significantly higher levels of oxytocin both under basal conditions and in response to stressor exposure in KO vs. intact control animals accompanied by elevated oxytocin mRNA levels in the PVN of KO rats. These findings suggest that the increased oxytocin levels in the PVN caused by the congenital absence of vasopressin may contribute to normal adrenocorticotropin stress responses in KO animals. However, whereas the stressor-induced elevation of plasma oxytocin in KO rats may be responsible for their maintained corticosterone levels, oxytocin seems unable to fully compensate for the lack of vasopressin. This hypothesis was tested by retrodialyzing synthetic vasopressin into the PVN area concomitantly with blood sampling in KO animals. Indeed, this treatment normalized plasma oxytocin and corticosterone levels 105 min after forced swimming. Thus, endogenous vasopressin released within the PVN is likely to act as a paracrine signal to facilitate the return of plasma oxytocin and corticosterone to basal levels after acute stressor exposure.

Catecholaminergic input to the oxytocin neurosecretory system in the human hypothalamus

Previous studies revealed that oxytocin release is increased by various forms of stress. Hypertonic saline injection, immobilization, and several other stressors elevated the blood level of oxytocin in rats. However, the mechanism of the stress-induced oxytocin release in human is not elucidated yet. Although numerous studies indicate that catecholamines play a pivotal role in modulating the release of oxytocin, there is a lack of data regarding the morphological substrate of this phenomenon. In order to reveal putative juxtapositions between tyrosine hydroxylase–immunoreactive (TH–IR) catecholaminergic and the oxytocinergic systems in the human hypothalamus, we utilized double-label immunohistochemistry in the present study. Numerous TH–IR axon varicosities abutted on oxytocin–IR neurons in the supraoptic and paraventricular nuclei, forming synapse-like associations. Close examination of these juxtapositions with high magnification failed to reveal any gaps between the contacting elements. In summary, the intimate associations between the TH–IR and oxytocin–IR elements may be functional synapses and may represent the morphological substrate of stress-influenced oxytocin release. The finding that several oxytocin–IR perikarya did not receive apparent TH innervation suggests that additional mechanisms may play significant roles in the oxytocin modulation by stressors.

5-HT2C receptor activation is a common mechanism on proerectile effects of apomorphine, oxytocin and melanotan-II in rats

5-Hydroxytryptamine (5-HT), dopamine, oxytocin and melanocortin pathways are known to be involved in the induction of penile erections in rats. Although a dopamine–oxytocin–5-HT link in the central nervous system has been suggested to be important to the control of penile erections, the 5-HT receptor subtype that mediates dopamine–oxytocin–5-HT action and the relationship between the dopamine–oxytocin–5-HT pathway and melanocortin pathway have not been fully elucidated. In this study, in order to clarify these matters, we examined the effects of a selective 5-HT2B/5-HT2C receptors antagonist, 1-(1-methylindol-5-yl)-3-(3-pyridyl)urea (SB200646) and a selective 5-HT2C receptor antagonist, 6-chloro-5-methyl-1-[6-(2-methylpyridin-3-yloxy) pyridin-3-yl carbamoyl] indoline (SB242084) on penile erections induced by a dopamine receptor agonist, 10, 11-dihydroxyaporphine (apomorphine), oxytocin, or a melanocortin receptor agonist, melanotan-II (MT-II) in rats. SB200646 at 10 mg/kg and SB242084 at 3 mg/kg, these doses which completely antagonize penile erections induced by 5-HT2C receptor agonists, m-chlorophenylpiperazine (mCPP) and (S)-2-(7-ethyl-1H-furo[2,3-g]indazol-1-yl)-1-methylethylamine (YM348), significantly inhibited penile erections elicited by apomorphine, oxytocin or MT-II. In addition, in order to clarify further the suggestion that the 5-HT pathway projecting from medulla oblongata to lumbosacral spinal site and lumbosacral 5-HT2C receptor are involved in the induction of penile erection, we also examined the proerectile effect of YM348 in spinal and a 5-HT depletor, p-chlorophenyl alanine (pCPA)-treated rats. YM348 induced intracavernous pressure increase in spinal and pCPA-treated rats as well as normal rats. These results suggest that 5-HT2C receptor in lumbosacral spinal sites mediates not only dopamine–oxytocin–5-HT action but melanocortin action on penile erections, and that the 5-HT pathway is located downstream from melanocortin pathway as well as the dopamine–oxytocin pathway.

The Effect of Amiloride during Infusion of Oxytocin in Male Sprague-Dawley Rats: A Study of a Possible Intrarenal Target Site for Oxytocin

A possible natriuretic mechanism of action of oxytocin was investigated in male Sprague-Dawley rats. The effects of an intravenous bolus injection of amiloride on urine volume, potassium and sodium excretion, and osmolality were measured with and without an intravenous infusion of oxytocin in saline. Control values were obtained during the infusion of saline. Amiloride administered during an oxytocin infusion increased sodium excretion from 0.1 ± 0.0 to 16.6 ± 2.1 μmol/min. In animals treated with amiloride only, the sodium excretion was 4.5 ± 0.8 μmol/min. The administration of oxytocin only resulted in a sodium excretion of 1.2 ± 0.3 μmol/min. After the administration of oxytocin, amiloride increased urinary flow from 4.3 ± 0.6 μl/min to 48.8 ± 6.1 μl/min. In animals treated with amiloride only, the flow after the bolus dose was 17.7 ± 1.8 μl/min. The administration of oxytocin only resulted in a flow of 8.5 ± 1.6 μl/min. The amiloride-caused change in potassium excretion was not inhibited by oxytocin. In summary, the effects of amiloride were not inhibited by the actions of oxytocin. Amiloride administrated after reaching a near steady-state effect of oxytocin was found to give rise to an effect far greater than that after the administration of oxytocin or amiloride alone. It is concluded that the intrarenal natriuretic mechanisms of oxytocin do not emanate from the amiloride-sensitive sodium channels.

Involvement of cholecystokinin receptor in the inhibition of gastrointestinal motility by oxytocin in ovariectomized rats

The effects of oxytocin on gastric emptying, gastrointestinal transit, and plasma levels of cholecystokinin (CCK) were studied in ovariectomized rats. Gastrointestinal motility was assessed in rats 15 min after intragastric instillation of a test meal containing charcoal and Na251CrO4. Gastric emptying was determined by measuring the amount of radiolabeled chromium contained in the small intestine as a percentage of the initial amount received. Gastrointestinal transit was evaluated by calculating the geometric center of distribution of the radiolabeled marker. Blood samples were collected for CCK radioimmunoassay. After administration of oxytocin (0.2–0.8 mg/kg), gastric emptying and gastrointestinal transit were inhibited, whereas plasma concentration of CCK was increased in a dose-dependent manner. Atosiban, an oxytocin receptor antagonist, effectively attenuated the oxytocin-induced inhibition of gastric emptying and gastrointestinal transit. However, administration of atosiban alone had no effect on gastric emptying and gastrointestinal transit. The selective CCK1 receptor antagonists, devazepide and lorglumide, effectively attenuated the oxytocin-induced inhibition of gastric emptying and gastrointestinal transit. L-365, 260, a selective CCK2 receptor antagonist, did not alter the oxytocin-induced inhibition of gastric emptying and gastrointestinal transit. These results suggest that oxytocin inhibits gastric emptying and gastrointestinal transit in ovariectomized rats via a mechanism involving the stimulation of CCK release and CCK1 receptor activation.

Sertraline, a selective serotonin reuptake inhibitor, affects thirst, salt appetite and plasma levels of oxytocin and vasopressin in rats

We investigated the effects of chronic administration of sertraline (SERT; approximately 20 mg kg(-1) day(-1) in drinking water), a selective serotonin reuptake inhibitor, on water and sodium intake and on plasma levels of oxytocin (OT) and vasopressin (AVP) in basal and stimulated conditions. Basal water intake was reduced in SERT-treated rats. After 24 h of water deprivation, rats treated with SERT for 21 days ingested less water than the control rats (9.7 +/- 0.5 versus 20.0 +/- 0.9 ml, respectively, at 300 min after water presentation, P < 0.0001). Subcutaneous injection of 2 m NaCl or isoproterenol evoked a lower dipsogenic response in rats treated with SERT for 21 days. Fluid and food deprivation also induced a weaker dipsogenic response in SERT-treated rats (1.6 +/- 0.5 versus 10.2 +/- 1.2 ml, at 300 min, P < 0.0001) but had no effect on saline intake. Sodium depletion induced a higher natriorexigenic response in the SERT group (5.6 +/- 1.3 versus 1.2 +/- 0.3 ml, at 300 min, P < 0.0002). Higher urinary density and lower plasma sodium levels were observed after SERT treatment. Sertraline also increased plasma levels of vasopressin and oxytocin (AVP, 2.65 +/- 0.36 versus 1.31 +/- 0.16 pg ml(-1), P < 0.005; OT, 17.16 +/- 1.06 versus 11.3 +/- 1.03 pg ml(-1), P < 0.0009, at the third week post-treatment). These data constitute the first evidence that chronic SERT treatment affects water and sodium intake in rats. These effects seem to be related to the hyponatraemia caused by the higher plasma levels of AVP and OT.

Oxytocin, water intake, and food sodium availability in male rats

This study examined the effect of subcutaneous administration of the neurohormone oxytocin on water intake of ad lib-fed (with or without sodium availability in the diet) and food-deprived animals. Results of the first experiment showed that oxytocin increased water intake and urine excretion in food-deprived but not in ad lib-fed animals. However, oxytocin treatment did not modify the reduced water “balance” (fluid intake minus urine volume) resulting from food deprivation or the daily food intake (Experiment 1). The dose-dependent polydipsic effect of oxytocin on food-deprived rats was always preceded by an increase in sodium and fluid urine excretion (Experiment 2). Oxytocin also increased the water intake of animals fed ad lib with a low sodium diet (Experiment 3). These results suggest that the effect of oxytocin on water intake is dependent on the presence or absence of sodium in the diet and that the excretion of sodium is the main mechanism of oxytocinergic polydipsia in food-deprived male rats.

Effect of exercise training on cardiac oxytocin and natriuretic peptide systems in ovariectomized rats

Exercise training results in cardiovascular and metabolic adaptations that may be beneficial in menopausal women by reducing blood pressure, insulin resistance, and cholesterol level. The adaptation of the cardiac hormonal systems oxytocin (OT), natriuretic peptides (NPs), and nitric oxide synthase (NOS) in response to exercise training was investigated in intact and ovariectomized (OVX) rats. Ovariectomy significantly augmented body weight (BW), left ventricle (LV) mass, and intra-abdominal fat pad weight and decreased the expression of oxytocin receptor (OTR), atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and guanylyl cyclase-A (GC-A), in the right atrium (RA) and LV, indicating estrogenic control of these genes. These effects of ovariectomy were counteracted by 8-wk-long exercise training which decreased fat pad weight (33.4 +/- 2.3 to 23.4 +/- 3.1 g, n = 8, P < 0.05), plasma free fatty acids (0.124 +/- 0.033 to 0.057 +/- 0.010 mM, n = 8, P < 0.01), and plasma triacylglycerol (0.978 +/- 0.174 to 0.588 +/- 0.115 mM, n = 8, P < 0.05). Chronic exercise tended to decrease BW and stimulated ANP (4- to 5-fold) and OTR gene expression in the LV and RA and BNP and inducible NOS (iNOS) mRNA in the LV. In sham-operated rats, exercise augmented ANP expression in the RA, downregulated GC-A mRNA in the LV and RA, but increased its expression threefold in the RA of OVX animals. Endothelial NOS and iNOS expression was enhanced in the left atrium of sham-operated rats. Altogether, these data indicate that in OVX animals, chronic exercise significantly enhances cardiac OT, NPs, and NOS, thus implicating all three hormonal systems in the beneficial effects of exercise training.

Design and Synthesis of the First Selective Agonists for the Rat Vasopressin V1bReceptor: Based on Modifications of Deamino-[Cys]arginine Vasopressin at Positions 4 and 8

The neurohypophyseal peptides arginine vasopressin (AVP) and oxytocin (OT) mediate a wide variety of peripheral and central physiological and behavioral effects by acting on four different G-protein coupled receptors, termed V1a (vascular), V1b (pituitary), V2 (renal), and OT (uterine). We recently reported that d[Cha4]AVP (A), d[Leu4]AVP (B), d[Orn4]AVP (C), and d[Arg4]AVP (D) have high affinity and are selective agonists for the human V1b receptor. However, peptides A-D were subsequently shown to be potent antidiuretic agonists in the rat and are, thus, not selective V1b agonists in the rat. Peptides A-D served as leads for the studies reported here. They were modified at position 8 by Lys, ornithine (Orn), diaminobutyric acid (Dab), and diaminopropionic acid (Dap) to give d[Cha4,Lys8]VP (1), d[Cha4,Orn8]VP (2), d[Cha4,Dab8]VP (3), d[Cha4,Dap8]VP (4), d[Leu4,Lys8]VP (5), d[Leu4,Orn8]VP (6), d[Leu4,Dab8]VP (7), d[Leu4,Dap8]VP (8), d[Orn4,Lys8]VP (9), d[Orn4,Orn8]VP (10), d[Arg4,Lys8]VP (11), d[Arg4,Orn8]VP (12), and d[Arg4,Dab8]VP (13). All peptides were synthesized by the Merrifield solid-phase method. Their binding and functional properties were evaluated in rat AVP V1a, V1b, and V2 receptors and on the rat OT receptor expressed either in native tissues or in stably transfected cells. They were also examined in rat vasopressor, antidiuretic, and in in vitro (no Mg++) oxytocic assays. Functional studies performed on chinese hamster ovary cells expressing the different AVP/OT receptors confirm that d[Cha4,Lys8]VP (1), d[Cha4,Dab8]VP (3), d[Leu4,Lys8]VP (5), and d[Leu4,Dap8]VP (8) are the first selective agonists for the rat V1b receptor. These selective V1b agonists are promising new tools for studies of the role of the V1b receptor in the rat.

Neuroendocrine Stress But Not Feeding Responses to Centrally Administered Neuropeptide Y Are Suppressed in Pregnant Rats

Metabolic peptides such as orexin and neuropeptide Y (NPY) exert profound effects on feeding but also act centrally to stimulate the hypothalamo-pituitary-adrenal (HPA) axis. In late pregnancy the HPA axis is hyporesponsive to centrally administered orexin-A, which signals to the HPA axis, in part, via arcuate NPY neurones. We investigated whether reduced HPA axis responses to orexin may be a consequence of down-regulated NPY signaling to the paraventricular nucleus (PVN) in pregnancy. Pregnant (d 21) and virgin rats were blood sampled for ACTH, corticosterone, and oxytocin (also a stress hormone in rats) before and after intracerebroventricular NPY or vehicle. Behavior was monitored. Rats were killed 4 h after NPY and brains removed for in situ hybridization. In another experiment rats were given vehicle or NPY, perfuse fixed 90 min later, and brain sections processed for Fos and oxytocin immunocytochemistry. NPY significantly increased ACTH, corticosterone and oxytocin secretion in the virgins but had no such effect on ACTH or oxytocin in the pregnant rats; the corticosterone response to NPY was markedly attenuated in pregnant rats. NPY increased CRH and vasopressin mRNA expression in the parvocellular PVN and stimulated Fos expression in magnocellular supraoptic and PVN oxytocin neurones of virgin but not pregnant rats. NPY increased food intake and drinking similarly in virgin and pregnant rats. Thus, neuroendocrine stress responses to central NPY are absent in late pregnancy, whereas ingestive behavioral responses are intact. These changes may explain the similarly attenuated HPA response to centrally administered orexin-A and will favor anabolic adaptations in pregnancy.

Bcl-xL and caspase inhibition increase the survival of rat oxytocin and vasopressin magnocellular neurons in organotypic culture

Hypothalamic magnocellular neurons (MCNs) are highly vulnerable to axotomy-induced cell death in vivo and in vitro. In this study, we determined whether the anti-apoptotic agent Bcl-xL, a member of the Bcl-2 family which prevents programmed cell death in the central nervous system, can rescue oxytocin (OT) and vasopressin (VP) MCNs in the supraoptic nucleus (SON) in organotypic culture. We found that the novel, membrane permeant form of Bcl-xL that we employed in these studies protected both OT and VP MCNs from degeneration as long as the Bcl-xL was present in the medium. In contrast, z-VAD-fmk, an inhibitor of caspases that are involved in apoptosis, was less effective in that it significantly rescued OT MCNs (P < 0.01) but not VP MCNs (P > 0.09). Unlike the Bcl-xL, Z-VAD-fmk's effectiveness in reducing MCN cell death was not sustained for the full 15 days in vitro.

An activation of parvocellular oxytocinergic neurons in the paraventricular nucleus in oxytocin-induced yawning and penile erection

Intracerebroventricular (ICV) or PVN local injections of oxytocin induce yawning and penile erection, for which a positive feedback mechanism for the PVN oxytocinergic activation is suggested, but this had not been directly substantiated in vivo. We have assessed the behavioral effects and activity of oxytocinergic neurons with double-staining for c-Fos and oxytocin in the PVN after ICV administration of oxytocin in adult male rats. ICV oxytocin injections (50 and 200 ng) dose-dependently induced yawning and penile erection and significantly increased the percentage of c-Fos positive oxytocin neurons in the medial, dorsal and lateral parvocellular subdivision of the PVN. However, increases in the magnocellular portion were not significant. We also found that lithium chloride (LiCl, 0.5 and l.0 mEq), a compound known to activate oxytocinergic neurons, also significantly increased the percentage of c-Fos positive oxytocin neurons in all PVN portions. However, LiCl did not induce yawning and penile erection, but counteracted the oxytocin-induced yawning and penile erection. These results suggest that if the activation of oxytocinergic neurons in the PVN is important for mediating oxytocin-induced yawning and penile erection, a selective activation of parvocellular oxytocinergic neurons in the PVN is likely to be involved.

Position 4 analogues of [deamino-Cys1] arginine vasopressin exhibit striking species differences for human and rat V2/V1b receptor selectivity

Arginine vasopressin (AVP) mediates a wide variety of biological actions by acting on three distinct G-protein coupled receptors, termed V(1a) (vascular), V(1b) (pituitary) and V(2) (renal). It also binds to the oxytocin (OT) receptor. As part of a program aimed at the design of selective agonists for the human V(1b) receptor, we recently reported the human V(1b), V(1a), V(2) and OT receptor affinities of the following position 4 substituted analogues of [deamino-Cys(1)] arginine vasopressin (dAVP)-(1) d[Leu(4)]AVP, (2) d[Orn(4)]AVP, (3) d[Lys(4)]AVP, (4) d[Har(4)]AVP, (5) d[Arg(4)]AVP, (6) d[Val(4)]AVP, (7) d[Ala(4)]AVP, (8) d[Abu(4)]AVP, (9) d[Nva(4)]AVP, (10) d[Nle(4)]AVP, (11) d[Ile(4)]AVP, (12) d[Phe(4)]AVP, (13) d[Asn(4)]AVP, (14) d[Thr(4)]AVP: (15) d[Dap(4)]AVP. With the exception of Nos. 7 and 12, all peptides exhibit very high affinities for the human V(1b) receptor. Furthermore, peptides 1-4 exhibit high selectivities for the human V(1b) receptor with respect to the V(1a), V(2) and OT receptors and, with d[Cha(4)]AVP, in functional tests, are the first high affinity selective agonists for the human V(1b) receptor (Cheng LL et al., J. Med. Chem. 47: 2375-2388, 2004). We report here the pharmacological properties of peptides 1-4, 5 (from a resynthesis), 7, 9-13, 15 in rat bioassays (antidiuretic, vasopressor and oxytocic) (in vitro: no Mg(++)) with those previously reported for peptides 5, 6, 8, 14. We also report the rat V(1b), V(1a), V(2) and OT receptor affinities of peptides 1-5 and the rat V(2) receptor affinities for peptides: 7-15.The antidiuretic activities in units/mg of peptides 1-15, are: 1=378; 2=260; 3=35; 4=505; 5=748; 6=1150; 7=841; 8=1020; 9=877; 10=1141; 11=819, 12=110; 13=996; 14=758; 15=1053. Peptides 1-4 exhibit respectively the following rat and human (in brackets) V(2) receptor affinities: 1=3.1 nm (245 nm); 2=3.4 nm (1125 nm); 3=24.6 nm (11,170 nm); 4=0.6 nm (1386 nm). Their rat V(1b) receptor affinities are 1=0.02 nm; 2=0.45 nm; 3=9.8 nm; 4=0.32 nm. Their rat V(1a) receptor affinities are 1=1252 nm; 2=900 nm; 3=1478 nm; 4=32 nm. Their rat oxytocin (OT) receptor affinities are 1=481 nm; 2=997 nm; 3=5042 nm; 4=2996 nm. All four peptides have high affinities and selectivities for the rat V(1b) receptor with respect to the rat V(1a) and OT receptors. However, in contrast to their high selectivity for the human V(1b) receptor with respect to the human V(2) receptor, they are not selective for the V(1b) receptor with respect to the V(2) receptor in the rat. These findings confirm previous observations of profound species differences between the rat and human V(2) receptors. Peptides 1-4 are promising leads to the design of the first high affinity selective agonists for the rat V(1b) receptor.

Centrally administered adrenomedullin 2 activates hypothalamic oxytocin-secreting neurons, causing elevated plasma oxytocin level in rats

We examined the effects of intracerebroventricular (i.c.v.) administration of adrenomedullin 2 (AM2) on plasma oxytocin (OXT) and arginine vasopressin (AVP) levels in conscious rats. Plasma OXT levels were markedly increased 5 min after i.c.v. administration of AM2 (1 nmol/rat) compared with vehicle and remained elevated in samples taken at 10, 15, 30, and 60 min. By contrast, plasma AVP levels were not significantly elevated in samples taken between 5 and 180 min after i.c.v. administration of AM2 except at the 30-min time point. Fos-like immunoreactivity (Fos-LI) was observed in various brain areas, including the paraventricular (PVN) and the supraoptic nuclei (SON) after i.c.v. administration of AM2 (2 nmol/rat) in conscious rats (measured at 90 min post-AM2 infusion). Dual immunostaining for OXT/Fos and AVP/Fos showed that OXT-LI neurons predominantly exhibited nuclear Fos-LI compared with AVP-LI neurons in the PVN and the SON. In situ hybridization histochemistry showed that i.c.v. administration of AM2 (0.2, 1, and 2 nmol/rat) caused marked induction of the expression of the c-fos gene in the PVN and the SON. This induction was significantly reduced by pretreatment with both the calcitonin gene-related peptide (CGRP) antagonist CGRP-(8-37) (3 nmol/rat) and the AM receptor antagonist AM-(22-52) (27 nmol/rat). These results suggest that centrally administered AM2 mainly activates OXT-secreting neurons in the PVN and the SON, at least in part through the CGRP and/or AM receptors with marked elevation of plasma OXT levels in conscious rats.

Differential expression of mesotocin receptors in the uterus and ovary of the pregnant tammar wallaby

Mesotocin, an oxytocin-like peptide, is released in highest concentrations during parturition in macropodid marsupials. In late pregnant wallabies, uterine sensitivity to mesotocin increases markedly in the myometrium of the gravid uterus. This coincides with a significant increase in myometrial mesotocin receptor concentrations 3-4 days before term. To date, there is no information on mesotocin receptor gene expression in female wallaby reproductive tissues. This study aimed to examine mesotocin receptor gene expression in the uterus and ovaries of pregnant tammar wallabies, and to localise mesotocin receptors within the uterus. An RT-PCR strategy produced a consensus nucleotide sequence of 834 bp, which encoded 278 amino acids of transmembrane domains I to VI. This protein sequence has approximately 80% homology with the bovine and rat oxytocin receptor exon 2 region. Only one mesotocin receptor was detected in the tammar genome. The myometrium and mammary gland both expressed a 4.1 kb mesotocin receptor gene transcript. Myometrial mesotocin receptor gene expression increased on day 22 of the 26-day gestation and was significantly higher in the gravid than the non-gravid uterus in late pregnancy. This pattern of mesotocin receptor gene expression paralleled mesotocin receptor concentrations. Mesotocin binding sites were localised only to the myometrium, the highest densities being observed in the gravid uterus. Finally, this study showed high expression of mesotocin receptors in the corpus luteum. The pattern of luteal mesotocin receptor expression differed from the myometrium, with a decrease in mesotocin receptors occurring on the day of expected births.

Oxytocinergic activity is linked to lower blood pressure and vascular resistance during stress in postmenopausal women on estrogen replacement

Estrogen administration results in increased release of the oxytocin (OT) prohormone reflected by increases in oxytocin intermediate peptide (OT Int) in both animal models and humans, and sequential treatment of ovariectomized rats with estrogen/progesterone then progesterone withdrawal leads to increased hypothalamic OT mRNA. Blood pressure (BP) reductions have been related to increased exogenous and endogenous OT in rats and to higher endogenous OT activity in premenopausal women, but not previously in postmenopausal women. Thus, we used plasma obtained at rest and during a speech stressor from 54 postmenopausal women who participated in a 6-month randomized trial of oral conjugated estrogens vs. placebo to examine effects of estrogen replacement therapy (ERT) on plasma OT and OT Int levels and their relationships to changes in BP during the trial. ERT alone and with progesterone (but not placebo) led to significant increases in plasma levels of OT Int, but no change in plasma OT levels. Women showing greater increases in OT Int during treatment showed greater decreases in BP and total vascular resistance during a series of behavioral stressors compared to women with moderate or no increases in OT Int, even after controlling for effects related to treatment condition or to changes in plasma estradiol. The findings suggest that enhanced oxytocinergic activity may contribute to BP decreases associated with ERT in more responsive postmenopausal women.