Oxytocin antagonist blocks the vasodepressor but not the vasopressor effect of neurohypophysial peptides in chickens.

Cockerels with permanent cannulas in the brachial artery and vein were put into isolated slings. Arterial pressure and heart rate were continuously recorded. Following habituation, tests were initiated. In each cockerel 2 nmol/kg of the tested neurohypophysial peptide (NPs) or analogue was IV injected six times at 6-min intervals. Arginine vasotocin (AVT) caused an immediate vasodepressor (VDP) effect and tachycardia. These subsided within 20-30 s and were followed by a vasopressor (VP) response and bradycardia. On repeated injections of AVT, the VDP response declined and bradycardia intensified. Arginine vasopressin (AVP), oxytocin (OT), and mesotocin (MT) had short-lasting VDP effect in the following order of potency: OT = MT > AVT > AVP. Only AVT and, more effectively, AVP, caused a VP response. The VDP effect of MT and OT declined on repeated injections. When AVT was injected after three injections of MT, it had mostly an immediate VP effect. Although the V1 agonist is VP in chickens, at the dose used the V1 antagonist, [d(CH2)5,O-Me-Tyr2]AVP, had no effect on cardiovascular responses to AVT. Pretreatment with OT antagonist, [d(CH2)5-O-Me-Tyr2-Thr4.Tyr9.Orn8]VT, abolished the VDP effect of all NPs. Thus, MT had no effect on blood pressure, whereas AVP and, more effectively, AVT, had a marked immediate VP action. In chickens the VDP effect of NPs is probably mediated by an OT/MT-like receptor, wherein the peptide's ring structure, shared by AVT, OT, and MT, is important.(ABSTRACT TRUNCATED AT 250 WORDS)

Distribution of immunoreactive mesotocin and vasotocin in the brain and pituitary of chickens.

The distribution of vasotocin and mesotocin in the pituitary and central nervous system in male chickens was determined using radioimmunoassays. Neither peptide was detected in the pineal. Mesotocin, but not vasotocin, was detected in the cerebellum. Both peptides were found in the septal area, archistriatum, paleostriatum, optic lobe, anterior, medial and posterior hypothalamus, midbrain, pons, medulla oblongata, and the anterior and posterior pituitary. Equal amounts of the 2 peptides were present in the septal area, archistriatum and anterior hypothalamus whereas vasotocin was more abundant (2- to 10-fold) in the paleostriatum, optic lobe, midbrain, and pituitary. The amount of mesotocin was about twice that of vasotocin in the medulla oblongata and the medial and posterior hypothalamus. The wide distribution of vasotocin and mesotocin in extrahypothalamic sites in the central nervous system suggests that the peptides may, as in mammals, have a role in a variety of autonomic and endocrine regulatory processes in chickens.

Induction of V2 receptors in renal medulla of homozygous Brattleboro rats by arginine vasopressin.

Homozygous Brattleboro rats display pronounced diabetes insipidus and when treated continuously with arginine vasopressin (AVP) acquire the ability to produce concentrated urine. In this study, the effects of continual AVP replacement on the pharmacological properties of the renal medullary V2 receptor and coupling to adenylate cyclase were examined. Osmotic minipumps that delivered AVP at four different rates were implanted into male homozygous Brattleboro rats. At the end of the 14 day treatment period, urine osmolalities were 280 +/- 24, 474 +/- 105, 1777 +/- 304 and 2202 +/- 175 mOsm/kg H2O for the 0, 31.25, 62.5 and 125 ng/hr treatment groups, respectively. Plasma AVP levels were below the level of detection for the 0 and 31.25 ng/hr treatment groups, and were 2.5 +/- 0.5 and 6.5 +/- 1.8 pg/ml for the 62.5 and 125 ng/hr treatment groups. Saturation experiments using [3H] AVP and renal medullary membranes revealed binding site concentrations of 57 +/- 9, 84 +/- 23, 164 +/- 17 and 150 +/- 18 fmol/mg protein for the 0, 31.25, 62.5 and 125 ng/hr treatment groups, respectively. AVP-stimulated cyclic AMP accumulation was enhanced in renal medullary membranes prepared from the 62.5 and 125 ng/hr treatment groups when compared to that in the 0 and 31.25 ng/hr treatment groups. From these results, it appears that circulating AVP is necessary for expression of functional V2 receptors in the homozygous Brattleboro rat renal medulla.

Arginine vasotocin gene expression in hypothalamic neurons is up-regulated in chickens drinking hypertonic saline: an in situ hybridization study.

Osmotic stress stimulates the release of the avian hypothalamic neuropeptide arginine vasotocin (AVT) into the peripheral circulation. We conducted the present study to investigate the effects of salt-loading on AVT secretion and AVT gene expression in specific hypothalamic nuclei in chickens. White Leghorn chickens were provided food ad lib and either water or 2% NaCl to drink. Both plasma osmolality and plasma AVT levels were significantly increased in chickens that drank 2% NaCl for either two or four days compared to that in chickens that drank water. Results from in situ hybridization analysis demonstrated an increase in the number of neurons expressing AVT mRNA in the supraoptic (SON) and paraventricular nuclei (PVN) in chickens provided 2% NaCl to drink compared to chickens that were provided water to drink. The number of grains per neuron increased in the PVN, but not in the SON of osmotically stimulated birds. Thus, increased osmolality resulting from ingestion of hypertonic saline is an effective stimulus to increase hypothalamic AVT mRNA content in chickens.

An in situ hybridization and immunohistochemical study of vasotocin neurons in the hypothalamus of water-deprived chickens.

The distribution of immunoreactive vasotocin (IR-AVT) and AVT mRNA in the hypothalamus of White Leghorn cocks was determined by immunohistochemistry and in situ hybridization, respectively. In control birds that were provided with water ad lib, AVT mRNA was distributed in the periventricular and lateral regions of the hypothalamus in clusters of neurons that correspond structurally with the mammalian paraventricular (PVN) and supraoptic (SON) nuclei. Although the distribution of AVT, identified by immunohistochemistry of adjacent serial sections within the hypothalamus, was similar to the distribution of AVT mRNA, the possibility that some positive staining was due to mesotocin neurons was not excluded. Water deprivation for 2 and 4 days resulted in both an increase in levels of AVT mRNA per neuron and the number of AVT mRNA-containing cells. Additionally, water deprivation resulted in a decrease in the amount of IR-AVT per neuron. The results indicate that osmotic stimulation increases AVT gene expression not only in individual neurons but also by activating subpopulation of neurons that are not observed in normally hydrated birds.

Arginine vasotocin gene expression in neuroendocrine, reproductive and gastrointestinal tissues of the domestic fowl: detection by reverse transcriptase polymerase chain reaction.

Arginine vasotocin gene transcripts in various tissues of the domestic fowl were detected by reverse transcriptase polymerase chain reaction followed by Southern blot analysis using a 209 bp fragment from the 3'-region of a cDNA encoding chicken arginine vasotocin as the probe. Relatively strong signals were observed with hypothalamic, adenohypophysial and proventricular RNA as the starting material. Lesser signals were obtained from RNA isolated from shell gland, adrenal gland, post-ovulatory follicles and ovarian thecal cells. Arginine vasotocin gene transcripts were undetectable in the posterior pituitary gland, small intestine and large intestine. These results suggest that in addition to its well-known antidiuretic and oxytocic actions, arginine vasotocin may act as a local neuromodulator or mediator and have other important autocrine or paracrine actions in non-hypothalamic tissues.

Absence of sodium appetite in cyclophosphamide and DOCA treated house mice.

Intraperitoneal administration of cyclophosphamide 100 mg/kg and 200 mg/kg significantly lowered plasma sodium and significantly increased plasma potassium but did not result in saline preference in a strain of wild-derived house mice given a choice between water and saline (0.15M) to drink. Deoxycorticosterone acetate treatment in dosages up to 1.5 mg for four days also failed to increase salt intake, The data suggest a possible absence of a sodium appetite mechanism in this species.

Embryonic differentiation of sexual dimorphism in vasotocin and mesotocin levels in chickens.

Chicken embryos of both sexes were injected on the tenth day of incubation with either estradiol benzoate (EB), aromatase inhibitor [1,4,6-androstatrien-3, 17-dione (ATD)], antiestrogen [tamoxifen (TAM)], antiandrogen [flutamide (FLU)], or the oil vehicle as control (C). At adulthood, at the age of 26 weeks, 10 chickens of each sex were killed and the amounts of immunoreactive arginine vasotocin (AVT) and mesotocin (MT) in the anterior hypothalamus (AHA), posterior hypothalamus (PHA), neurohypophysis (NHP), and pineal gland (PNL) were determined. Control hens had significantly more AVT in PNL and less MT in AHA and NHP than the corresponding roosters. This sexual dimorphism was affected by the embryonic treatments; TAM increased AVT in AHA of cockerels but not of hens. In both sexes, TAM and FLU increased AVT content in NYP. In males, but not in females, ATD also increased AVT content in the NHP. TAM and FLU administration to the female embryo reduced PNL AVT to the amount present in normal males. None of the treatments effected AHA MT in hens, while in cockerels TAM increased it. In females, TAM and FLU significantly increased NHP MT to the level of C males. In roosters, ATD, TAM, and FLU increased NHP MT further. In hens, but not roosters, FLU reduced MT in PNL. These results indicate that embryonic differentiation of the MT and AVT systems is affected by gonadal steroids in chickens.

Distribution of immunoreactive vasotocin and mesotocin in the chicken gastrointestinal tract.

Immunoreactive arginine vasotocin (AVT) and mesotocin (MT) were measured in heart, breast muscle, adrenals, testes, and different parts of the gastrointestinal tract in adult male chickens. Neither of the peptides were detected in liver, testis, heart and breast muscle. The amounts of AVT and MT in the adrenals were 167 +/- 25 and 669 +/- 198 pg/gland, respectively. Considerable amounts of immunoreactive peptides were found in the gastrointestinal tract with the highest concentration in the proventriculus (4.18 +/- 0.31 ng AVT and 16.58 +/- 0.86 ng MT per organ). Dose-response curves of duodenal and proventriculus extracts were parallel with synthetic AVT and MT standards.

Alterations in uterine contractility during the oviposition cycle in domestic hens.

1. The potencies of several neurohypophysial hormones were examined at different times during the oviposition cycle in an in vitro fowl oxytocic assay. 2. Uterine tissues were removed 2 h before (-2h-OP), immediately after (0h-OP) and 5 h after (+5h-OP) spontaneous oviposition. In addition, uterine tissue was removed immediately after oviposition was induced, by administering prostaglandin E2 2 h before an expected oviposition (Induced-OP). 3. The rank order of oxytocic potencies for the peptides was arginine vasotocin = vasopressin greater than oxytocin greater than mesotocin. The sensitivity of the uterus to the hormones was 0h-OP = Induced-OP greater than -2h-OP = +5-OP. 4. These results suggest that uterine sensitivity to neurohypophysial hormones changes during the oviposition cycle in domestic fowls.

Apparent volumes of distribution of 125-I-lothalamate and inulin in chickens.

The suitability of utilizing 125-I-iothalamate to estimate the volume of extracellular fluid was assessed in ureterally ligated chickens. Subsequent to intravenous administration the movement of labeled iothalamate from the plasma compartment follows closed two-compartment kinetics and equilibration between vascular and extravascular phases is attained in about 20 minutes. The volume of distribution of 125-I-iothalamate prior to and following the influsion of 0.15 M NaCl (equal to 15% of the estimated ECFV) averaged 23.6 plus or minus 0.61 and 28.4 plus or minus 0.22% of the body weight, respectively. The observed postsaline labeled iothalamate space did not differ statistically from the expected value. When administered simultaneously inulin penetrates into an apparent volume that is 75% of the labeled iothalamate space after 60 minutes. The content of 125-I-iothalamate is relatively high in liver and kidney tissue and suggests that these are major sites where removal of the indicator from plasma occur. It is suggested that 125-I-iothalamate, under appropriate conditions, could be used to measure the plasma volume and the extravascular fluid with which plasma is in rapid diffusion equilibrium.

Flight effects on plasma levels of neurohypophysial hormones in homing pigeons.

A significant increase in the circulating levels of arginine vasotocin (AVT), without change in plasma osmolality, was observed in homing pigeons immediately after a flight of 48 km, lasting 60-80 min. This increase in plasma AVT is perceived as part of an overall homeostatic mechanism of mobilizing lipid as fuel for the flight muscles, of water conservation and temperature regulation in the body during the homing flight within the specific distance and duration of flight and under normal weather conditions. However, there was no change in plasma levels of mesotocin (MT) which suggests that the birds were under no significant stress during the flight.

Plasma levels of immunoreactive mesotocin and vasotocin during oviposition in chickens: relationship to oxytocic action of the peptides in vitro and peptide interaction with myometrial membrane binding sites.

Plasma concentrations of immunoreactive vasotocin (AVT) and mesotocin (MT) were measured periodically before and subsequent to spontaneous oviposition in conscious chickens. The concentrations of AVT and MT approximately an hour prior to oviposition were 5.2 +/- 1.1 microU/ml and 14.7 +/- 5.1 pg/ml, respectively. Plasma AVT levels increased abruptly at oviposition (25.1 +/- 3.3 microU/ml) and decreased to 5.0 +/- 0.6 microU/ml within 30 min postoviposition. Significant changes in MT were not observed. The data indicate that AVT is selectively released during oviposition. The uterus was removed immediately after oviposition and the oxytocic potencies of several peptides were tested on muscle strips in vitro. The order of oxytocic potencies was AVT greater than or equal to arginine vasopressin (AVP) much greater than MT = pressinoic acid. Partially purified membranes were prepared from separate portions of the uteri used in the oxytocic assay. [3H]arginine8 vasopressin, [3H]AVP, bound to membranes saturably (Bmax = 17 fmol/mg protein) and with high affinity (Kd = 0.7 nM). The rank order of potency of the peptides in displacing [3H]AVP from the binding sites was the same as in the oxytocic assay which suggests that the [3H]AVP binding sites in uterine membranes represent physiological receptors that interact with AVT during oviposition.

Arginine vasotocin and mesotocin levels in theca and granulosa layers of the ovary during the oviposition cycle in hens (Gallus domesticus).

Neurohypophysial hormones in mammals are synthesized by the ovary and may influence reproductive function. Although arginine vasotocin (AVT) can induce premature oviposition in chickens, the gonadal content of AVT or mesotocin (MT) has not been examined in this species. Ovarian follicles were removed 2 hr before oviposition (-2 hr), immediately after oviposition (0 hr), and 5 hr after oviposition (+5 hr). The theca and granulosa layers of the three largest preovulatory and postovulatory follicles were isolated, extracted, and assayed for immunoreactive AVT and MT. AVT content in the theca and granulosa layers was lowest at -2 hr and increased to maximum levels at +5 hr. Pre- and postovulatory follices contained similar amounts of AVT. Maximum AVT levels in the theca and granulosa layers were 2.5 +/- 0.6 and 1.3 +/- 0.2 ng/tissue, respectively. In contrast, theca content of MT was maximal at 0 hr and the highest peptide levels were observed in the largest preovulatory follicle (0.24 +/- 0.02 ng/tissue). The results demonstrate that, compared with circulating levels of the hormones, the ovary of the chicken contains high concentrations of AVT and MT. The ovarian content of AVT and MT varies in a different manner in relation to the oviposition cycle.

Changes in plasma concentrations of arginine vasotocin after intrauterine injections of prostaglandin F-2 alpha and acetylcholine at various times during the oviposition cycle of the domestic hen (Gallus domesticus).

Prostaglandin F-2 alpha (PGF-2 alpha, 1 microgram) and acetylcholine (10 mg) were injected into the uterus of chickens 23, 21, 16, 8 or 4 h before expected oviposition. Plasma concentrations of immunoreactive arginine vasotocin and PGF were measured in relation to the time of administration of PGF-2 alpha or acetylcholine or to the premature oviposition that was induced. PGF-2 alpha or acetylcholine administration caused premature oviposition and a marked increase in plasma arginine vasotocin levels only when an egg was present in the uterus. Changes in plasma PGF concentrations were not observed. After premature oviposition was induced, plasma values of PGF and arginine vasotocin increased at the expected time of oviposition. Manual stimulation of the uterus 4 h after oviposition also stimulated arginine vasotocin release. During spontaneous oviposition, a rise in plasma PGF concentration preceded increases in uterine contractility and plasma arginine vasotocin concentration. These results suggest that PGF may stimulate uterine contractility which in turn causes the release of arginine vasotocin to provide an additional contractile stimulus during oviposition.