pituitrin and isotocin

The brain peptides vasopressin and oxytocin play crucial roles in the regulation of salt and water balance. The genes encoding these neurohormones are regulated by cell-specific and physiological cues, but the molecular mechanisms remain obscure. New strategies, involving the introduction of rat transgenes into rats, are being used to address these issues, but the complexity of the rat genome has hampered progress. By contrast, the pufferfish, Fugu rubripes, has a "junk-free" genome. The oxytocin homologue from Fugu, isotocin, has been introduced into rats and is expressed in oxytocin neurons, where it is upregulated by physiological perturbations that upregulate the oxytocin gene. The Fugu and rat lineages separated 400 million years ago, yet the mechanisms that regulate the isotocin and oxytocin genes have been conserved. Fugu genome analysis and transgenesis in the physiologically tractable rat host are a powerful combination that will enable the identification of fundamental components of the neural systems that control homeostasis.

Topics: Animals; Animals, Genetically Modified; Body Water; Cattle; Diuresis; Evolution, Molecular; Fishes, Poisonous; Gene Expression Regulation; Genome; Homeostasis; Hypothalamo-Hypophyseal System; Hypothalamus; Kidney Tubules, Collecting; Mice; Natriuresis; Neurons; Osmotic Pressure; Oxytocin; Rats; Repetitive Sequences, Nucleic Acid; Sodium; Species Specificity; Transgenes; Vasopressins; Vasotocin; Water-Electrolyte Balance

Oxytocin (OT) and arginine-vasopressin (AVP) regulate social behavior in mammals. Zebrafish (Danio rerio) allows higher throughput and ease in studying human brain disorders.. This study investigated in zebrafish the effect of non-mammalian homologs isotocin (IT) and vasotocin (AVT) in comparison with OT/AVP on social behavior and fear response to predator. The mechanism was studied using the most human selective OT and AVP receptor antagonists.. Zebrafish were injected i.m. with increasing doses (0.001-40 ng/kg) of the neuropeptides. DesGly-NH(2)-d(CH(2))(5)-[D-Tyr(2),Thr(4)]OVT) for OT receptor, SR 49059 for V1a subtype receptor, and SSR-149415 for V1b subtype receptor were injected i.m. 10 min before each agonist.. All the peptides increased social preference and reduced fear to predator response in a dose-dependent manner interpolated by symmetrical parabolas. AVT/AVP were more potent to elicit anxiolytic than social effect while IT and OT were equally potent. All the antagonists dose-dependently inhibited both the effects induced by the neuropeptides. The ratio between the ED50 obtained for blocking the OT-induced effects on social preference and fear response to predator was very high only for desglyDTTyrOVT (160). SR49059 showed the highest ratio in blocking AVP-induced effects (807). The less selective antagonist appeared to be SSR149415.. For the first time, IT/AVT and OT/AVP were found to modulate in zebrafish, social behavior, unrelated to sex, and fear to predator response through at least two different receptors. Zebrafish is confirmed as a valid, reliable model to study deficit in social behavior characteristic of some psychiatric disorders.

Topics: Animals; Anti-Anxiety Agents; Antidiuretic Hormone Receptor Antagonists; Dose-Response Relationship, Drug; Fear; Indoles; Ornipressin; Oxytocin; Pituitary Hormones, Posterior; Pyrrolidines; Radioligand Assay; Receptors, Oxytocin; Receptors, Vasopressin; Social Behavior; Swimming; Vasopressins; Vasotocin; Zebrafish

In silurid fishes, semen collection is practically impossible, even after hormonal stimulation. Instead, males are killed and testes macerated to obtain sperm. To understand the endocrine control of semen release in catfishes, we investigated the role of smooth muscle contractors in semen release and semen quality of African catfish (Clarias gariepinus). In in vitro experiments, testis slices were incubated with oxytocin (1 and 10 IU), isotocin (2 and 20 ug), vasopressin (0.2 and 2 ug), epinephrine (1 and 10 ug), PGF2alpha (1 and 10 ug), purified Clarias LH (300 ng) and partly purified Clarias pituitary extract (containing 300 ng LH). Only oxytocin increased sperm concentration of the medium (assessed by optical density measurements) compared to control incubations. Oxytocin was then tested in vivo in two groups of fish: normal males, and males that had been treated with 17alpha-methyltestosterone during larval stages to inhibit seminal vesicle development (MT males). Both groups of fish received two doses of carp pituitary suspension (8 and 10 mg/kg, respectively i.m.) with or without subsequent oxytocin treatment (5 IU/kg i.v.; cPS-OT treatment and cPS treatment, respectively). There was no effect of oxytocin on the number of strippable males. Of cPS and cPS-OT treated fish, 87% MT males and 60% normal males were strippable. The stripped semen volume was low in both groups but MT males produced higher (P < 0.001) hatching rates (63.1%) than did normal males (2.1%).

Topics: Animals; Catfishes; Dinoprost; Epinephrine; Larva; Luteinizing Hormone; Male; Methyltestosterone; Oxytocin; Pituitary Gland; Semen; Sperm Count; Testis; Tissue Extracts; Vasopressins

Neurohypophysial hormone receptors and second messengers were studied in trout (Oncorhynchus mykiss) hepatocytes. Arginine vasotocin (AVT) and isotocin (IT) elicited a concentration-dependent inhibition of cAMP accumulation in the presence of 5x10(-8) M glucagon (maximal effect for 4.5x10(-7) M and 1.4x10(-7) M, half-maximal effect for 2.1x10(-8) M and 0.7x10(-8) M, AVT and IT respectively). The effect of glucagon was inhibited up to 90% by AVT and 80% by IT. While AVT inhibited (up to 50%) the basal cAMP production, IT had no such action. Specific V(1) or V(2) analogues (with reference to vasopressin in mammals) were used for pharmacological characterization of the type of neurohypophysial hormone receptor involved in this inhibition. The V(1) agonist [Phe(2), Orn(8)]-oxytocin inhibited the glucagon-stimulated cAMP production with a maximal effect for 6x10(-7) M and a half-maximal effect for 0.9x10(-8) M concentrations of the analogue. While the V(1) agonist reduced the glucagon-stimulated cAMP level by 70%, it showed only a tendency to reduce the basal level. The V(2) agonist [deamino(1), Val(4),d -Arg(8)]-vasopressin had no effect either on basal or on glucagon-stimulated cAMP production. The V(1) antagonist [d(CH(2))(5)(1), O-Me-Tyr(2), Arg(8)]-vasopressin totally reversed the 10(-8) M AVT-induced inhibition of 5x10(-8) M glucagon-stimulated cAMP production, whereas the V(2) antagonist [d(CH(2))(5)(1),d -Ile(2), Ile(4), Arg(8), Ala(9)]-vasopressin had no such effect. In this particular case, maximal and half-maximal effects of the V(1) antagonist were obtained for 2.3x10(-6) M and 1. 2x10(-6 )M respectively. Changes in intracellular calcium content were measured using the fluorescent probe FURA-2/AM. AVT and IT elicited a concentration-dependent increase in Ca(2+) accumulation. The comparison of the effect of 10(-8) M agonists versus AVT showed the following order of potency: AVT=IT>V(1) agonist>V(2) agonist. The V(1) antagonist reversed the AVT-induced Ca(2+) accumulation whereas the V(2) antagonist had no such effect. These results are taken as evidence for the presence in trout hepatocytes of neurohypophysial hormone receptors functionally close to the V(1a)-type linked to cAMP production and Ca(2+) mobilization.

Topics: Animals; Calcium; Cell Culture Techniques; Cyclic AMP; Dose-Response Relationship, Drug; Glucagon; Hepatocytes; Oncorhynchus mykiss; Oxytocin; Pituitary Gland, Posterior; Receptors, Vasopressin; Second Messenger Systems; Vasopressins; Vasotocin

The metabolic actions of the vasoactive peptides vasotocin and isotocin and the alpha-agonist phenylephrine are examined in hepatocytes isolated from three teleost species: brown bullhead, rainbow trout, and American eel. These three compounds influenced hepatic gluconeogenesis and glycogenolysis with significant species differences. Vasotocin and isotocin affected only eel hepatocytes activating gluconeogenesis by 1.7-fold and glycogenolysis by 3-fold. Phenylephrine increased glycogenolysis by 7-fold in bullhead hepatocytes and gluconeogenesis by 1.4-fold in trout cells. Vasotocin and phenylephrine actions were correlated with increases in adenosine 3',5'-cyclic monophosphate (cAMP). The vasotocin effects were unaffected by beta- and alpha-antagonists supporting a V2-type receptor on eel hepatocytes. Phenylephrine effects were abolished by propranolol and reduced by prazosin and yohimbine (alpha 1- and alpha 2-antagonists, respectively). Phenylephrine, therefore, affected fish hepatocyte metabolism either by a mixed alpha/beta-receptor mechanism emphasizing beta-adrenoceptors or the classic alpha/beta agonist/antagonist scheme defined for mammals is not appropriate for these fish preparations.

Topics: 1-Methyl-3-isobutylxanthine; Anguilla; Animals; Cells, Cultured; Gluconeogenesis; Ictaluridae; Kinetics; Liver; Liver Glycogen; Oxygen Consumption; Oxytocin; Phenylephrine; Prazosin; Propranolol; Trout; Vasopressins; Vasotocin; Yohimbine

Arginine vasotocin (AVT) and isotocin (IT) induced direct inhibition of adenylate cyclase activity in gill plasma membranes of the rainbow trout adapted to freshwater (FW) and seawater (SW). The maximal inhibition was obtained with 10(-11)-10(-10) M (50% inhibitory concentration approximately 10(-13) M), values in agreement with the circulating levels of AVT in trout blood. Specific V1 or V2 agonists or antagonists (with reference to vasopressin) were used to define the specificity of the neurohypophysial peptide receptors involved in this inhibition. The V1 agonist [Phe2,Orn8]vasotocin ([Phe2]OVT) inhibited the basal and glucagon-stimulated adenylate cyclase activity, and this effect in SW (20%) was twice more than in FW (10%). The V2 agonist 1-deamino[Val4,Arg8]-vasopressin (dVDAVP), however, produced a stimulation that was of the same amplitude (10%) in both media. The V1 antagonist [(1-beta-mercapto-beta-beta-cyclopentamethylenepropionic acid), 1-(O-ethyl)Tyr2,Orn8]vasotocin (d(CH2)5[Tyr(Et)2]OVT) totally reversed the AVT- or IT-induced inhibition of basal or glucagon-stimulated cyclase activity, whereas the V1/V2 antagonist [(1-beta-mercapto-beta-beta-cyclopentamethylene propionic acid), 1-(O-ethyl)D-Tyr2,Val4,Arg8]vasopressin (d(CH2)5[D-Tyr(Et)2]-VAVP) (previously used as V2 antagonist in amphibians) had no such effect. When active, all analogues had their maximal activity at 10(-11)-10(-10) M (50% maximal activity approximately 10(-13) M), as observed with the natural peptides. These results, together with our previous observations, point to the gill epithelium as a direct target organ for neurohypophysial peptides and suggest that the hormone receptors involved in fish belong predominantly, if not exclusively, to a new type that we propose to designate as fish neurohypophysial hormone (NHF) receptors while awaiting further characterization.

Topics: Adenylyl Cyclases; Animals; Epithelium; Gills; Glucagon; Oxytocin; Pituitary Gland, Posterior; Receptors, Pituitary Hormone; Salmonidae; Trout; Vasopressins; Vasotocin

Two genes each encoding a distinct precursor protein to the hormone isotocin and a neurophysin-related protein are present in the teleost fish Catostomus commersoni. These precursors are referred to as isotocin 1 and 2. As shown by the polymerase chain reaction technique, both genes lack introns in their protein-coding sequences. Both genes are transcribed giving rise to mRNAs of 920 (isotocin 1) and 1020 (isotocin 2) bases, respectively. Based on the nucleotide sequences, the predicted isotocin precursors contain, besides the hormone moiety, a neurophysin-like protein that, in contrast to its mammalian counterpart, is extended at its C-terminus by a peptide which includes a leucine-rich core segment. This segment shows similarities to the copeptin of the mammalian vasopressin precursor that is known to possess prolactin-releasing activity. The data imply that the mammalian copeptin sequence was initially part of a larger ancestral neurophysin molecule.

Topics: Amino Acid Sequence; Animals; Base Sequence; Blotting, Northern; Blotting, Southern; Cypriniformes; DNA; Exons; Genes; Introns; Molecular Sequence Data; Neurophysins; Oxytocin; Polymerase Chain Reaction; Protein Precursors; Sequence Homology, Nucleic Acid; Transcription, Genetic; Vasopressins

The release of endogenous oxytocin and vasopressin by rat paraventricular and supraoptic nuclei in vitro during a 10-min period, 30 min after beginning the incubation, was measured radioimmunologically. Mean basal hormone release per 10 min and per pair of nuclei was: 128.4 +/- 12.4 (S.E.M.) pg vasopressin (n = 15) and 39.0 +/- 3.0 pg oxytocin (n = 66) for supraoptic nuclei from male rats; 273.9 +/- 42.6 pg vasopressin (n = 11) and 34.2 +/- 3.5 pg oxytocin (n = 15) for supraoptic nuclei from lactating rats; 70.0 +/- 8.6 pg vasopressin (n = 52) and 21.8 +/- 1.3 pg oxytocin (n = 68) for paraventricular nuclei from male rats; 59.1 +/- 8.6 pg vasopressin (n = 10) and 27.0 +/- 4.6 pg oxytocin (n = 16) for paraventricular nuclei from lactating rats. In male and lactating rats, both nuclei contained and released more vasopressin than oxytocin. For oxytocin alone, the paraventricular nucleus of male rats contained and released significantly less hormone than the supraoptic nucleus. This difference was not apparent in lactating rats. For vasopressin alone, the paraventricular nucleus contained and released significantly less hormone than the supraoptic nucleus in both male and lactating rats. When the hormone released was calculated as a percentage of the total tissue content the release was about 0.9% for oxytocin from both nuclei in male and lactating rats and also for vasopressin in lactating rats, but was only about 0.5% for vasopressin from both nuclei in male rats. The influence of oxytocin and analogues of oxytocin (including one antagonist) upon the release of oxytocin and vasopressin was studied. Adding oxytocin to the incubation medium (0.4-4 nmol/1 solution) induced a dose-dependent rise in oxytocin release from both nuclei of male or lactating rats. A 4 nmol/l solution of isotocin had a similar effect to a 0.4 nmol/l solution of oxytocin, but arginine-vasopressin never affected basal release of oxytocin. In no case was vasopressin release modified. An oxytocin antagonist (1 mumol/l solution) significantly reduced basal oxytocin release and blocked the stimulatory effect normally induced by exogenous oxytocin, as did gallopamil hydrochloride (D600, 10 mumol/l solution), a Ca2+ channel blocker, or incubation in a Ca2+-free medium. These findings are discussed in relation to the literature on the central effects of neurohypophysial peptides. It may be concluded that the regulatory role of endogenous oxytocin in the hypothalamus on the milk-ejection reflex

Topics: Animals; Dose-Response Relationship, Drug; Feedback; Female; Lactation; Male; Oxytocin; Paraventricular Hypothalamic Nucleus; Pregnancy; Radioimmunoassay; Rats; Rats, Inbred Strains; Supraoptic Nucleus; Vasopressins

Vasopressin and related peptides cause short-lasting hypothermia when injected into the lateral ventricle of the rat. In the present study, the structure-activity relationships for the induction of this effect were examined. For the agonist peptides studies, the structural requirements were found to be similar to those required to cause peripheral vasoconstriction ant to induce behavioral excitation in mice. However, an antagonist of the pressor and behavioral effects of vasopressin was ineffective in antagonizing the hypothermic response. Moreover, this analog and another pressor antagonist themselves caused hypothermia. Comparison with the structure-activity relationships for other effects on the central nervous system strongly suggests that the hypothermic response is unrelated to the effects of vasopressin on consolidation of memory, development of tolerance to drugs, and mechanisms of reinforcement.

Topics: Animals; Arginine Vasopressin; Hypothermia, Induced; Lypressin; Male; Oxytocin; Rats; Structure-Activity Relationship; Vasopressins; Vasotocin