Antagonistic effects of vasotocin and isotocin on the upper esophageal sphincter muscle of the eel acclimated to seawater

The effects of isotocin (IT) and vasotocin (VT), which are fish analogues of mammalian oxytocin and vasopressin respectively, were examined in the isolated upper esophageal sphincter (UES) muscle. IT relaxed and VT constricted the UES muscle in a concentration-dependent manner. The relaxation by IT and the contraction by VT were completely blocked by H-9405 (an oxytocin receptor antagonist) and by H-5350 (a V₁-receptor antagonist), respectively, suggesting that the eel UES possesses both IT and VT receptors. Truncated fragments of VT did not show any significant effects, indicating that all nine residues are essential for the VT and IT actions. IT may relax the UES muscle through enhancing cAMP production, since similar relaxation was also observed after treatment with 3-isobutyl-1-methylxantine, forskolin and 8-bromoadenosine, 3′, 5′-cyclic mono-phosphate (8BrcAMP). Although 8-bromoguanosine, 3′, 5′-cyclic monophosphate also relaxed the UES, its effect was less than 1/3 of that 8BrcAMP, suggesting minor contribution of nitric oxide (NO) in the relaxation of the UES muscle. Both peptides seem to act directly on the UES muscle, not through release of other substances from the epithelial cells, since similar relaxation and contraction were observed even in the scraped UES preparations. When IT and VT were intravenously administrated (in vivo experiments), the drinking rate of the seawater eel was enhanced by IT and was inhibited by VT. These effects correspond to the in vitro results described above, relaxation by IT and contraction by VT in the UES muscle. The significance of the relaxing effect by IT is discussed with respect to controlling the drinking behavior of the eel.     

The vertebrate social behavior network: evolutionary themes and variations

Based on a wide variety of data, it is now clear that birds and teleost (bony) fish possess a core "social behavior network" within the basal forebrain and midbrain that is homologous to the social behavior network of mammals. The nodes of this network are reciprocally connected, contain receptors for sex steroid hormones, and are involved in multiple forms of social behavior. Other hodological features and neuropeptide distributions are likewise very similar across taxa. This evolutionary conservation represents a boon for experiments on phenotypic behavioral variation, as the extraordinary social diversity of teleost fish and songbirds can now be used to generate broadly relevant insights into issues of brain function that are not particularly tractable in other vertebrate groups. Two such lines of research are presented here, each of which addresses functional variation within the network as it relates to divergent patterns of social behavior. In the first set of experiments, we have used a sexually polymorphic fish to demonstrate that natural selection can operate independently on hypothalamic neuroendocrine functions that are relevant for (1) gonadal regulation and (2) sex-typical behavioral modulation. In the second set of experiments, we have exploited the diversity of avian social organizations and ecologies to isolate species-typical group size as a quasi-independent variable. These experiments have shown that specific areas and peptidergic components of the social behavior network possess functional properties that evolve in parallel with divergence and convergence in sociality.     

Post- and pre-synaptic action of isotocin in the upper esophageal sphincter muscle of the eel: its role in water drinking

Isotocin is a fish analogue of the mammalian hormone oxytocin. To elucidate sites of action of isotocin (IT) in the upper esophageal sphincter (UES) muscle, a key muscle in swallowing, IT was applied after treatment with tetrodotoxin (TTX). Even after blocking nerve activity with TTX, IT relaxes the UES muscle in a concentration-dependent manner, suggesting that IT receptor(s) is present on the muscle cells. Similar relaxation was also obtained by application of 3-isobutyl-1-methylxanthine (IBMX), forskolin (FSK) and 8-bromo-adenosine, 3′,5′-cyclic monophosphate (8BrcAMP) after pretreatment with TTX, suggesting that the relaxing effect (postsynaptic action) of IT may be mediated by cAMP. In contrast to such relaxing effect, IT enhanced the UES contraction induced by repetitive electrical field stimulation (EFS). Such enhancement was blocked by an IT receptor antagonist, suggesting that this effect is also mediated by IT receptor(s). Similar enhancement was also induced by IBMX, FSK and 8BrcAMP, suggesting the enhancing effect is also mediated by cAMP. However, no enhancing effect of IT was observed when the muscle was stimulated by carbachol, or after treatment with curare or TTX, denying the postsynaptic modulatory action of IT and suggesting presynaptic action for IT, i.e., accelerating acetylcholine release. Summarizing these results, role of IT in precisely regulating the drinking rate in the seawater eel is discussed.     

Do circulating plasma AVT and/or cortisol levels control pulsatile urea excretion in the gulf toadfish (Opsanus beta)?

Previous work has shown that pulsatile urea excretion at the gills of the gulf toadfish is due to periodic activation of a facilitated diffusion transport system with molecular and pharmacological similarity to the UT-A transport system of the mammalian kidney. In mammals, AVP and glucocorticoids are two important endocrine regulators of this system. The present study focused on the potential role of circulating AVT (the teleost homologue of AVP) and cortisol levels as possible triggers for urea pulses. Long-term (34–84 h) monitoring of plasma levels by repetitive sampling at 2-h intervals from chronic cannulae in individual toadfish demonstrated that circulating AVT concentrations are low (10⁻¹²–10⁻¹¹ M), and show no relationship to the occurrence of natural urea pulses. In contrast, plasma cortisol levels decline greatly prior to natural pulses and rise rapidly thereafter. AVT injections into the caudal artery or ventral aorta elicited pulse events, but these were extremely small (1–10%) relative to natural pulses, and occurred only at unphysiological dose levels (10⁻⁹ M in the plasma). AVP was a partial agonist, but isotocin, insulin-like growth factor-1, and atrial natriuretic peptide were without effect at the same concentration. Artificially raising plasma cortisol levels by cortisol injection tended to reduce responsiveness to AVT. Pharmacological reduction of plasma cortisol levels by metyrapone injection elicited small pulses similar to those caused by AVT. Following such pulse events, AVT was ineffective in inducing pulses. We conclude that decreases in circulating cortisol play an important permissive role in urea pulsing, but that circulating AVT levels are not involved.     

Changes in expression of provasotocin and proisotocin genes during adaptation to hyper- and hypo-osmotic environments in rainbow trout

Summary The physiological roles of neurohypophysial hormones, vasotocin (VT) and isotocin (IT), are not yet clear in teleosts. Since information on responsiveness of hypothalamic neurosecretory neurons to environmental stimuli may contribute to an understanding of their physiological roles, effects of environmental hyper- and hypo-osmotic stimuli on expression of VT and IT precursor (proVT and proIT) genes in rainbow trout were investigated, using an in situ hybridization technique in which 46 mer synthetic oligonucleotides were used as hybridization probes. The probes corresponded to the mRNA loci encoding chum salmon proVT (-5 to 11) and proIT (-5 to 11), and were labeled at the 3-end with ³⁵S. Autoradiographic silver grains which represent the hybridization signals of proVT and proIT mRNAs were localized in both magnocellular and parvocellular neurons in the nucleus preopticus magnocellularis (NPOmg). Localizations of proVT and proIT hybridization signals coincided with those of VT- and IT-immunoreactive neurons in adjacent sections, and showed that proVT and proIT genes are expressed in separate neurons. The intensity of proVT hybridization signals as determined by grain counting in magnocellular neurons in the NPOmg was conspicuously decreased after transfer from fresh water (FW) to 80% seawater (SW). The proVT mRNA levels in SW trout were consistently lower than those of FW trout for up to 2 weeks. After return from 80% SW to FW, the proVT mRNA level increased, attaining the initial FW level. The proIT mRNA levels in SW trout were not statistically different from those in FW trout, except for the 1st day after transfer to SW. These results suggest that synthesis of proVT was elevated by transfer from higher to lower salinity, and that VT may have a physiological role in salmonid osmoregulation, especially in adaptation to a hypo-osmotic environment.Abbreviations ABC avidin-biotin-peroxidase complex - AVP arginine vasopressin - BSA bovine serum albumin - EDTA ethylene diamine tetra-acetic acid - cpm counts per minute - FW fresh water - GFR gromerular filtration rate - ISH in situ hybridization - IT isotocin - mRNA messenger ribonucleic acid - NPOmg nucleus preopticus magnocellularis - OXT oxytocin - PBS phosphate-buffered saline - SSC saline sodium citrate - RT room temperature - SW seawater - VT vasotocin     

Ultrastructural characterization of neurosecretory fibres immunoreactive for vasotocin,isotocin, somatostatin,LHRH and CRF in the pituitary of a teleost fish,Poecilia latipinna

Summary Using the electron-microscopic immunogold method, vasotocin, isotocin, somatostatin (SRIF), gonadotrophin-releasing hormone (LHRH) and corticotrophin-releasing factor (CRF)-like immunoreactivities were localized in separate neurosecretory fibres in the pituitary of a teleost fish Poecilia latipinna. Antigenicities were preserved in sections of conventionally fixed tissue, except in the case of LHRH and CRF-like substances which were sensitive to osmium postfixation. Under the same fixation conditions, ultrastructural differences were observed between the 5 fibre types, and morphometric analysis of their granule sizes revealed significant differences in mean diameter except between vasotocin and isotocin fibres.Terminal-like regions of each type were identified on blood vessels, glial cells or other fibres in the neurohypophysis, on the basement lamina of the adenohypophysis, or directly on adenohypophysial endocrine cells. The fibres containing the two neurohypophysial hormones, originating from separate preoptic perikarya, were intermingled with, and may form endings near all the adenohypophysial cell types except those secreting prolactin. Although both types had similar mean granule diameters, the granules in the vasotocin fibres (mean 135 nm) were markedly less electron dense than those in the isotocin fibres (mean 140 nm). SRIF-immunoreactive fibres (mean 101 nm) appeared to form synapse-like endings on the somatotrophs, and a few thyrotrophs in the proximal pars distalis, and near the pars intermedia cells. An LHRH-positive type (mean 103 nm) contacted only the gonadotrophs of the proximal pars distalis. The rarer CRF-like fibres (mean 116 nm) appeared to project mainly towards the pars intermedia, but a few appeared to terminate rostrally near the adrenocorticotrophic cells.The significance of these observations is discussed in relation to the direct neurosecretory control of adenohypophysial function in teleosts.