Search

Thirst has evolved for vertebrate terrestrial adaptation. We previously showed that buccal drying induced a series of drinking behaviours (migration to water-taking water into the mouth-swallowing) in the amphibious mudskipper goby, thereby discovering thirst in ray-finned fish. However, roles of dipsogenic/antidipsogenic hormones, which act on the thirst center in terrestrial tetrapods, have remained unclear in the mudskipper thirst. Here we examined the hormonal effects on the mudskipper drinking behaviours, particularly the antagonistic interaction between angiotensin II (AngII) and atrial natriuretic peptide (ANP) which is important for thirst regulation in mammalian 'forebrain'. Expectedly, intracerebroventricular injection of ANP in mudskippers reduced AngII-increased drinking rate. ANP also suppressed the neural activity at the 'hindbrain' region for the swallowing reflex, and the maintenance of buccopharyngeal water due to the swallowing inhibition may attenuate the motivation to move to water. Thus, the hormonal molecules involved in drinking regulation, as well as the influence of buccopharyngeal water, appear to be conserved in distantly related species to solve osmoregulatory problems, whereas hormonal control of thirst at the forebrain might have been acquired only in tetrapod lineage during evolution.

Thirst aroused in the forebrain by angiotensin II (AngII) or buccal drying motivates terrestrial vertebrates to search for water, whereas aquatic fish can drink surrounding water only by reflex swallowing generated in the hindbrain. Indeed, AngII induces drinking through the hindbrain even after removal of the whole forebrain in aquatic fish. Here we show that AngII induces thirst also in the amphibious mudskipper goby without direct action on the forebrain, but through buccal drying. Intracerebroventricular injection of AngII motivated mudskippers to move into water and drink as with tetrapods. However, AngII primarily increased immunoreactive c-Fos at the hindbrain swallowing center where AngII receptors were expressed, as in other ray-finned fish, and such direct action on the forebrain was not found. Behavioural analyses showed that loss of buccal water on land by AngII-induced swallowing, by piercing holes in the opercula, or by water-absorptive gel placed in the cavity motivated mudskippers to move to water for refilling. Since sensory detection of water at the bucco-pharyngeal cavity like 'dry mouth' has recently been noted to regulate thirst in mammals, similar mechanisms seem to have evolved in distantly related species in order to solve osmoregulatory problems during terrestrialization.

The crab-eating frog Fejervarya cancrivora inhabits mangrove swamps and marshes in Southeast Asia. In the present study, circulating angiotensin II (Ang II), aldosterone (Aldo), arginine vasotocin (AVT), and corticosterone (Cort) concentrations as well as various blood parameters were studied under osmotically stressful conditions. Following acclimation to hyperosmotic seawater and dry condition for 5days, body weight was significantly decreased. Under both conditions, plasma Na(+), Cl(-), and urea concentrations, hematocrit values (Ht; blood volume indicator), and osmolality were significantly increased. Dehydration associated with hypovolemic and hyperosmotic states of body fluids was induced during acclimation to hyperosmotic seawater and dry condition in the crab-eating frogs. Ang II, Aldo, AVT, and Cort were maintained within relatively narrow concentration ranges in the control frogs; however, in frogs under dry and hyperosmotic seawater conditions, large variations were observed among individuals in each group. Mean plasma Ang II and Aldo concentrations significantly increased in hyperosmotic seawater-acclimated and desiccated frogs. Although mean plasma AVT concentrations in dehydrated frogs of both the groups were approximately 2.0-3.5 times higher than those in the control frogs, the differences were not significant because of the variation. There was a significant correlation between plasma osmolality and AVT as well as Ang II but not Aldo. A significant correlation was also observed between Ht and AVT as well as Ang II. Plasma Ang II was significantly correlated with plasma Aldo. These results indicate that the crab-eating frogs may exhibit similar physiological responses to both seawater-acclimated and dry conditions. It appears that under dehydrated conditions, osmoregulatory mechanisms participate in stabilization of the situation. The renin-angiotensin system may have pivotal roles in body fluid regulation under volemic and osmotic stress in the Fejervarya species with unique osmoregulation., (Copyright © 2013 Elsevier Inc. All rights reserved.)

In the brains of teleosts, angiotensin II (ANG II), one of the main effector peptides of the renin-angiotensin system, is implicated in various physiological functions notably body fluid and electrolyte homeostasis and cardiovascular regulation, but nothing is known regarding the potential action of ANG II and other angiotensin derivatives on ventilation. Consequently, the goal of the present study was to determine possible ventilatory and cardiovascular effects of intracerebroventricular injection of picomole doses (5-100 pmol) of trout [Asn(1)]-ANG II, [Asp(1)]-ANG II, ANG III, ANG IV, and ANG 1-7 into the third ventricle of unanesthetized trout. The central actions of these peptides were also compared with their ventilatory and cardiovascular actions when injected peripherally. Finally, we examined the presence of [Asn(1)]-ANG II, [Asp(1)]-ANG II, ANG III, and ANG IV in the brain and plasma using radioimmunoassay coupled with high-performance liquid chromatography. After intracerebroventricular injection, [Asn(1)]-ANG II and [Asp(1)]-ANG II two ANG IIs, elevated the total ventilation through a selective stimulatory action on the ventilation amplitude. However, the hyperventilatory effect of [Asn(1)]-ANG II was threefold higher than the effect of [Asp(1)]-ANG II at the 50-pmol dose. ANG III, ANG IV, and ANG 1-7 were without effect. In addition, ANG IIs and ANG III increased dorsal aortic blood pressure (P(DA)) and heart rate (HR). After intra-arterial injections, none of the ANG II peptides affected the ventilation but [Asn(1)]-ANG II, [Asp(1)]-ANG II, and ANG III elevated P(DA) (50 pmol: +80%, +58% and +48%, respectively) without significant decrease in HR. In brain tissue, comparable amounts of [Asn(1)]-ANG II and [Asp(1)]-ANG II were detected (ca. 40 fmol/mg brain tissue), but ANG III was not detected, and the amount of ANG IV was about eightfold lower than the content of the ANG IIs. In plasma, ANG IIs were also the major angiotensins (ca. 110 fmol/ml plasma), while significant but lower amounts of ANG III and ANG IV were present in plasma. In conclusion, our study suggests that the two ANG II isoforms produced within the brain may act as a neurotransmitter and/or neuromodulator to regulate the cardioventilatory functions in trout. In the periphery, two ANG IIs and their COOH-terminal peptides may act as a circulating hormone preferentially involved in cardiovascular regulations.

Angiotensinogen belongs to family A serine protease inhibitors (SERPIN family) and we have cloned and characterized SERPIN genes in two lamprey species, which possess all the properties of angiotensinogen. The putative angiotensinogens in lampreys can be considered as an evolutionary link between SERPIN and other angiotensinogen according to the phylogenetic analyses. The inferred sea lamprey angiotensinogen gene was expressed abundantly in liver and to a lesser extent in other tissues. The predicted lamprey angiotensin I (Ang I) sequence was unique and different from the teleost-type Ang I identified previously by the incubation of lamprey plasma with its kidney extract. Therefore, we characterized and compared the biochemical and physiological properties of this native lamprey Ang II (LpAng II) (EEDYDERPYMQPF) with teleost-type Ang II (NRVYVHPF). Using a newly developed RIA for LpAng II, plasma levels in Japanese lamprey were measured (157.4 ± 35.2 fmol/ml, n=6), but teleost-type Ang II was undetectable. In conscious cannulated lamprey, LpAng II at 100 pmol/kg elicited a transient vasodepressor effect. At doses higher than 300 pmol/kg, a biphasic cardiovascular response with an initial vasodepressor effect followed by a transient rebound vasopressor effect was observed in a dose-dependent manner. However, teleost-type Ang II was not vasoactive up to 1 nmol/kg. In Japanese eel, LpAng II injection up to 3 nmol/kg did not alter the cardiovascular parameters. Our results suggested that the renin-angiotensin system first appeared in cyclostomes, and LpAng II could be important for the regulation of cardiovascular dynamics in lampreys because of its potent and acute vasoactive effect.

Ghrelin is a potent, centrally acting orexigenic hormone. Recently, we showed that centrally administered ghrelin is a potent antidipsogenic hormone in 24-h water deprived rats. In this study, we examined the effect of intracerebroventricular (icv) injection of ghrelin on angiotensin II (AII)-induced water intake in rats. We also examined the effects of icv injection of ghrelin on drinking induced by intraperitoneal injection of an isotonic polyethylene glycol (PEG) solution that causes isotonic hypovolemia. Water intake induced by the icv injection of AII or ip injection of PEG was significantly reduced after icv injection of ghrelin, although food intake was stimulated by the hormone. The drinking induced by AII was also inhibited by the icv administration of 4alpha-phorbol 12, 13-didecanoate, an agonist of the osmosensitive TRPV4 channel. This study showed that ghrelin is a potent antidipsogenic peptide by antagonizing general dipsogenic mechanisms including those activated by AII and hypovolemia in rats.

This study examined the effect of transfer to increased environmental salinity on the circulating levels of angiotensin II (ANG II), C-type natriuretic peptide (CNP), and arginine vasotocin (AVT) in the euryhaline elasmobranch, Carcharhinus leucas. Plasma levels of ANG II and CNP were significantly increased in C. leucas chronically acclimated to seawater (SW) in comparison to freshwater (FW) acclimated fish. There was no difference in plasma AVT levels. Acute transfer of FW fish to 75% SW induced an increase in plasma ANG II levels within 12 h, and subsequent transfer from 75 to 100% SW further increased plasma ANG II levels at both 24 and 72 h. No change in plasma CNP was observed during acute transfer to increased salinity. However, a significant increase in plasma AVT levels was observed following 96 h in 75% SW and 24 h in 100% SW. In chronically SW acclimated C. leucas plasma osmolality, sodium, chloride, and urea were all significantly higher than FW acclimated fish but there was no difference in haematocrit. Acute transfer of C. leucas to 75% SW induced a significant increase in plasma osmolality, sodium and urea concentrations within 96 h of transfer. Subsequent transfer from 75 to 100% SW induced a further increase in these variables within 24 h in addition to a significant increase in plasma chloride above control levels. Haematocrit did not differ between the experimental and control groups throughout the acute study. Circulating levels of ANG II were significantly correlated to plasma, sodium, chloride, and urea concentrations during acclimation to SW. Conversely, circulating levels of CNP and AVT did not correlate to plasma osmolytes, however, CNP was significantly correlated to haematocrit during acclimation to seawater.

To clarify the physiological roles of the renin-angiotensin-aldosterone system (RAAS) and arginine vasotocin (AVT) on body fluid regulation in amphibians, we measured plasma concentrations of aldosterone (ALDO), angiotensin II (ANG II), and AVT after various osmotic challenges in the marine toad, Bufo marinus (Bufonidae). Hematocrit value (Ht) as an indicator of plasma volume, plasma osmolality and concentrations of plasma components (Na(+), Cl(-), K(+), and urea) were also measured. The toads were maintained under various osmotic treatments for 7 days. In dehydrated toads, plasma concentrations of ALDO, ANG II, AVT, and all plasma components measured were increased. In toads maintained in 300 mosmol/kg H(2)O NaCl solution, plasma osmolality, Na(+), Cl(-), urea, and plasma AVT concentrations were significantly increased, and Ht and plasma concentrations of ALDO and ANG II were significantly decreased. In toads maintained in tap water, plasma osmolality, and concentrations of Na(+) and ALDO were significantly decreased. We also estimated total body water (TBW), plasma volume (PV) using Evans Blue dye and Ht in the toads under various osmotic treatments. In dehydrated toads, TBW and PV were significantly decreased and Ht was significantly increased in comparison with those of control. In toads maintained in 300 mosmol/kg H(2)O NaCl solution, TBW and PV were significantly increased and Ht was significantly decreased in comparison with those of control. There was a significant negative correlation between Ht and PV or TBW. These results show that dehydration, which induces hypovolemic and hyperosmotic conditions, stimulates increases of plasma ALDO, ANG II, and AVT concentrations, while hypervolemic treatment induces decreases of plasma ALDO and ANG II concentrations. There were significant correlations between plasma osmolality and AVT concentration, between Ht and concentrations of RAAS hormones, and between plasma concentrations of ALDO and ANG II. These results suggest that volumetric and osmometric systems regulated by RAAS hormones and AVT are present in B. marinus.

The effects of angiotensin II (ANG II) and natriuretic peptides (NPs) of the eel (ANP, atrial natriuretic peptide; CNP, C-type natriuretic peptide; and VNP, ventricular natriuretic peptide) on prolactin (PRL(188) and PRL(177)) and growth hormone (GH) release from the organ-cultured tilapia pituitary were examined. Eel ANG II at concentrations greater than 1 nM stimulated the release of PRL(188) and PRL(177) in a dose-related manner during the first hour of incubation. Significant stimulation by 100 nM ANG II on PRL(177) release was observed until 4h of incubation, and on PRL(188) release until 12 h. No effect of ANG II was seen on GH release. None of the NPs altered the release of PRLs at any time point. On the other hand, eel VNP at concentrations greater than 1 nM stimulated GH release in a dose-related manner after 4 h, and significant stimulation was observed until 48 h. Eel CNP was less effective than eel VNP; significant stimulation of GH release was observed at 1 and 10 nM during 24-48 h of incubation. No significant effect of eel ANP on GH release was seen at any concentration. ANG II had no effect on GH release at any time point. There was no change in mRNA levels of PRLs or GH in the pituitaries incubated with ANG II for 8 h or those incubated with the NPs for 48 h. These results indicate rapid and short-lasting stimulation by ANG II on PRL release and slow and long-lasting stimulation by VNP and CNP on GH release from the tilapia pituitary., (Copyright 2003 Elsevier Science (USA))

Dogfish (125)I [Asn(1), Pro(3), Ile(5)] angiotensin II ((125)I dfANG II) was used to establish the specific binding patterns of the different cardiac regions of the elasmobranch Scyliorhinus canicula by in vitro autoradiography. In the ventricular myocardium Scatchard analysis of saturation and displacement binding data revealed two classes of high- and low-affinity dfANG II binding sites (K(d) = 53 +/- 10 and 1300 +/- 900 pM). Two classes of dfANG II binding sites were also detected in the atrium (K(d) = 47 +/- 13 and 4690 +/- 930 pM) and in the outer layer of the conus arteriosus (K(d) = 16 +/- 9 and 398 +/- 83 pM). Conversely, the ventricular endocardium and the inner conal layer were characterized by a single class of dfANG II binding sites with affinity values of 48 +/- 11 and 106 +/- 3.3 pM, respectively. Competition experiments with either cold dfANG II or CV11974 or CGP42112 (specific ligands for mammalian AT(1) and AT(2) receptors, respectively) demonstrated a prevalence of CGP42112-selective dfANG II binding sites in both the inner and the outer conal layers. In the atrium, the ventricular myocardium, and the outer conal layer, dfANG II high-affinity binding sites poorly discriminated among the cold ligands. These results suggest that the dogfish heart may be a target organ of ANG II with distinct ANG II receptor subtype distributions., (Copyright 2001 Academic Press.)

The contributions of circulating angiotensin II (Ang II) and catecholamines to cardiovascular control in the spiny dogfish were investigated by monitoring the effects of exogenous and endogenous dogfish [Asn1, Pro3, Ile5]-Ang II (dfAng II) on plasma catecholamine levels and blood pressure regulation. Bolus intravenous injections of dfAng II (30-1200 pmol kg-1) elicited dose-dependent increases in plasma adrenaline and noradrenaline concentrations, caudal artery pressure (PCA), and systemic vascular resistance (RS), and a decrease in cardiac output (Q). Similar injections of Ang II in dogfish pre-treated with the alpha-adrenoceptor antagonist yohimbine (4 mg kg-1) also elicited dose-dependent increases in plasma catecholamine levels yet the cardiovascular effects were abolished. Dogfish treated with yohimbine were hypotensive and had elevated levels of plasma Ang II and catecholamines. Intravenous injection of the smooth muscle relaxant papaverine (10 mg kg-1) elicited a transient decrease in PCA and RS, and increases in plasma Ang II and catecholamine levels. In dogfish first treated with lisinopril (10(-4) mol kg-1), an angiotensin converting enzyme inhibitor, papaverine treatment caused a more prolonged and greater decrease in PCA and RS, an attenuated increase in plasma catecholamines, and no change in plasma Ang II. By itself, lisinopril treatment had little effect on PCA, and no effect on RS, plasma Ang II or catecholamines. In yohimbine-treated dogfish, papaverine treatment elicited marked decreases in PCA, RS, and Q, and increases in plasma Ang II and catecholamines. Among the three papaverine treatments, there was a positive linear relationship between plasma Ang II and catecholamine concentrations, and the cardiovascular and hormonal changes were most pronounced in the yohimbine + papaverine treatment. Therefore, under resting normotensive conditions, while Ang II does not appear to be involved in cardiovascular control, catecholamines play an important role. However, during a hypotensive stress elicited by vascular smooth muscle relaxation. Ang II indirectly contributes to cardiovascular control by dose-dependently stimulating catecholamine release.

The individual contributions of, and potential interactions between, the renin-angiotensin system (RAS) and the humoral adrenergic stress response to blood pressure regulation were examined in rainbow trout. Intravenous injection of the smooth muscle relaxant, papaverine (10 mg/kg), elicited a transient decrease in dorsal aortic blood pressure (PDA) and systemic vascular resistance (RS), and significant increases in plasma angiotensin II (Ang II) and catecholamine concentrations. Blockade of alpha-adrenoceptors before papaverine treatment prevented PDA and RS recovery, had no effect on the increase in plasma catecholamines, and resulted in greater plasma Ang II concentrations. Administration of the angiotensin-converting enzyme inhibitor, lisinopril (10(-4) mol/kg), before papaverine treatment attenuated the increases in the plasma concentrations of Ang II, adrenaline, and noradrenaline by 90, 79, and 40%, respectively and also prevented PDA and RS recovery. By itself, lisinopril treatment caused a gradual and sustained decrease in PDA and RS, and reductions in basal plasma Ang II and adrenaline concentrations. Bolus injection of a catecholamine cocktail (4 nmol/kg noradrenaline plus 40 nmol/kg adrenaline) in the lisinopril+papaverine-treated trout, to supplement their circulating catecholamine concentrations and mimic those observed in fish treated only with papaverine, resulted in a temporary recovery in PDA and RS. These results indicate that the RAS and the acute humoral adrenergic response are both recruited during an acute hypotensive stress, and have important roles in the compensatory response to hypotension in rainbow trout. However, whereas the contribution of the RAS to PDA recovery is largely indirect and relies on an Ang II-mediated secretion of catecholamines, the contribution from the adrenergic system is direct and relies at least in part on plasma catecholamines.

The effects of eel atrial natriuretic peptide (ANP) on drinking were investigated in eels adapted to freshwater (FW) or seawater (SW) or in FW eels whose drinking was stimulated by a 2-ml hemorrhage. An intra-arterial infusion of ANP (0.3-3.0 pmol . kg-1 . min-1), which increased plasma ANP level 1.5- to 20-fold, inhibited drinking dose dependently in all groups of eels. The drinking rate recovered to the level before ANP infusion within 2 h after infusate was replaced by saline. The inhibition at 3.0 pmol . kg-1 . min-1 was profound in FW eels and hemorrhaged FW eels, whereas significant drinking still remained after inhibition in SW eels. Plasma ANG II concentration also decreased dose dependently during ANP infusion and recovered to the initial level after saline infusion in all groups of eels. The decrease at 3.0 pmol . kg-1 . min-1 was large in FW eels and hemorrhaged FW eels compared with that of SW eels. Thus the changes in drinking rate and plasma ANG II level were parallel during ANP infusion. Plasma sodium concentration and osmolality decreased during ANP infusion in SW and FW eels, and they were restored after saline infusion. In hemorrhaged FW eels, however, ANP infusion did not alter plasma sodium concentration and osmolality. Hematocrit did not change during ANP infusion in any group of eels. Collectively, ANP infusion at physiological doses decreased drinking rate and plasma ANG II concentration in parallel in both FW and SW eels. It remains undetermined whether the inhibition of drinking is caused by direct action of ANP or through inhibition of ANG II, which is known as a potent dipsogen in all vertebrate species, including eels.

Arterial rings were prepared from the branchial artery, coeliac artery and ventral aorta of the Japanese dogfish Triakis scyllia and used to determine arterial contraction in a myograph. Noradrenaline caused a dose-dependent contraction (10(-9)-3 x 10(-6) M) that was completely inhibited by pre-treatment with 10(-7) M phentolamine. Homologous dogfish angiotensin II (ANG II) ([Asn1, Pro3, Ile5]-ANG II) also caused dose-dependent contraction (10(-9)-3 x 10(-6) M), but phentolamine had no effect on this response. Administration of dogfish angiotensin I (ANG-I) ([Asn1, Pro3, Ile5, Gln9]-ANG I) resulted in a contraction similar to that produced by ANG II and the effect could be blocked with 10(-7) M captopril. The mammalian ANG II receptor antagonists [Sar1, Ile8]-ANG II and [Sar1, Ala8]-ANG II caused dose-dependent contractions of coeliac artery rings, but were less potent than homologous ANG I and ANG II. These results show that the contractile effect of [Asn1, Pro3, Ile5]-ANG II is not mediated by the alpha-adrenergic system and contractions of arterial rings by noradrenaline and elasmobranch ANG II are mediated by separate vascular receptors. The elasmobranch ANG II vascular receptor may have co-evolved with the unusual structure of this peptide.

A homologous radioimmunoassay was developed to determine the concentration of angiotensin II (Asn1, Pro3, Ile5)-Ang II) in elasmobranchs. Cross-reactivity with elasmobranch angiotensin I and other heterologous angiotensins was high and therefore all potentially cross-reacting angiotensins were separated by high performance liquid chromatography after prior extraction with Sep-Pak C18 cartridges. The validity of the assay for the determination of elasmobranch Ang II was demonstrated by parallelism with a series of Ang II standards with serially diluted elasmobranch plasma extracts. Overall recovery of elasmobranch Ang II added to a plasma pool was 75.1 +/- 5.2%. Plasma Ang II concentrations measured by our RIA were similar in fish adapted to 70, 100, or 120% SW at 139 +/- 20.1, 109 +/- 15.3, and 119 +/- 16.3 fmol . ml-1, respectively., (Copyright 1998 Academic Press.)

The presence of specific Ang II receptors in membrane fractions was investigated using 125I-labeled homologous Ang II ([Asn1, Pro3, Ile5]Ang II; df Ang II) in Triakis scyllia. Specific binding sites occurred in a variety of tissues, with highest binding in interrenal tissue (17.11 +/- 2.45 fmol Ang II/mg protein) and gill (6.26 +/- 0. 69 fmol Ang II/mg protein) and possible Ang II receptors in rectal gland and other tissues. 125I-[Asn1, Pro3, Ile5]Ang II (10(-10)M) binding to branchial cell membrane fraction (25 microg protein) in 5 mM MgCl2, 125 mM NaCl, 50 mM Tris-HCl, 0.2% bovine serum albumin at 28 degrees (1) is rapid and saturable; (2) increases as a function of membrane concentration and time; and (3) optimally fits to a two-site (high-and low-affinity) model. The equilibrium dissociation constant (0.11 +/- 0.01 nM) and binding site concentration (35.00 +/- 1.16 fmol/mg protein) are similar to those of mammalian and avian vascular Ang II receptors. Bound labeled ligand was not competitively displaced by dogfish Ang I, dogfish C-type natriuretic peptide, bradykinin, or the AT1 receptor antagonist, CV 11974. The AT2 receptor antagonist, CGP 42112, was much less potent at displacing the labeled ligand compared to the unlabeled ligand.

The amino acid sequence of angiotensin I (ANG I) from the Japanese quail, Coturnix coturnix japonica, obtained from incubation of homologous plasma and kidney extract was determined as H-Asp-Arg-Val-Tyr-Val-His-Pro-Phe-Ser-Leu-OH. A bolus, intravenous injection of native ANG I or of ANG II induced an immediate vasodepressor response and a subsequent vasopressor response in quail which has been lightly anesthetized with urethane (0.75 g/kg). The values for ED50 for the vasopressor and depressor effects of ANG II were 85 and 113 pmol/100 g body weight, respectively. The extent of the hypotension was dependent on the arterial pressure prior to injection. The effects of ANG I and II on heart rate were variable. Human [Asp1, Ile5, His9] ANG I and II were almost as potent as their quail counterparts with respect to the cardiovascular effects, but eel [Asn1, Val5, Gly9]ANG I and II were less than half as potent. Human ANG III had little effect on arterial pressure in the quail. A bolus injection (100 micrograms/100 g) or infusion (1 micrograms/100 g/min) of [Sar1, Ile8] ANG II almost abolished the cardiovascular effects of ANG I and II, but [Sar1, Ala8] ANG II and [Sar1, Thr8] ANG II, which are effective inhibitors in mammals, had no inhibitory effects. The vasopressor and depressor effects of ANG I were abolished, while those of ANG II were slightly enhanced, after injection (100 micrograms/100 g) or infusion (1 micrograms/100 g/min) of SQ14225, whereas des-Pro2 bradykinin and bradykinin potentiator B, which are effective inhibitors of ANG I converting enzyme in mammals, failed to inhibit the effect of ANG I. These results indicate that vascular ANG II receptors and ANG I converting enzyme in the quail may be different from those in mammals.

Monamine distribution in a septohypothalamic area was investigated in the Japanese quail using a histochemical fluorescence method. This area includes the subfornical organ (SFO) and the preoptic area (POA) which are inferred dipsogenic receptor sites for angiotensin II (AII) in the Japanese quail. Nerve fibers showing yellow-green fluorescence were found between the POA and the SFO. Thwy traversed from the POA to the SFO, and some fibers seemed to terminate on the neurons in the SFO. After a low dose of reserpine, a considerable number of fluorescent perikarya were found in the POA. These fibers and perikarya appeared to be of primary catecholamine judging from the fluorescence color. Following transection of these fibers, fluorescence disappeared from the fibers located on the SFO side of the transection plane, while it became a little more intense on the POA side. After transection, microinjection of AII into the POA was no longer effective in induction of drinking. On the other hand, sham operation or transection in areas other than between the POA and the SFO produced only minute changes in those fluorescent fibers and had little effect on the dipsogenic potency of AII injected into the POA. These results suggest that information of AII perceived at the POA is transferred to the SFO via those primary catecholamine-containing nerve fibers, which effect induced drinking.

Synthetic 5-valine angiotensin II (AII) induced copious drinking when applied directly to the the subfornical organ (SFO) in the Japanese quail. Reliable response was obtained with as little as 1 ng of AII. The amount of water intake increased dose-dependently from 5 ng to 1 microgram. A latent period of 73.0 +/- 11.0 seconds at 100 ng was noted. The electrical destruction of the SFO significantly reduced the amount of water intake induced by both intravenous and intracranial AII injections. The decrease was proportional to the extent of the SFO lesion. It is conceivable, therefore, that the SFO plays an important role in elicitation of drinking by AII in birds as suggested in mammals.

The Japanese quail drinks water vigorously after water deprivation, haemorrhage and administration of hypertonic saline solution. Most avian species responded to angiotensin II (AII) by drinking, but carnivorous birds and those originating in arid regions were insensitive. The receptive sites for AII were the subfornical organ (SFO) and the preoptic area (POA) in the Japanese quail. Catecholaminergic fibers proceed from the POA to the SFO. Dipsogenic information generated by AII at the POA is transferred to the SFO through the catecholaminergic nerve fibres. Plasma AII increased following dehydration and haemorrhage and returned to a normal level immediately after rehydration. Following dehydration, arginine vasotocin, aldosterone and corticosterone increased in plasma as well as AII. A single intraperitoneal injection of AII induced increases of arginine vasotocin, aldosterone and corticosterone in plasma. It seems that AII functions as a trigger for release of these other hormones during dehydration.

In the domestic fowl, angiotensin II (ANG II) causes an in vivo depressor response and in vitro relaxation of aortic rings which appear to be a direct action of ANG II on the blood vessels. Thus, we determined whether binding sites specific to ANG II exist in the membrane fraction of the fowl aorta. The particulate fraction of aortas from adult female fowl, Gallus gallus, exhibits high specific binding to ANG II ligand. 125I-[Ile5]ANG II (0.5 nM) binding to the above fraction (30 micrograms protein) in 50 mM Tris (pH 7.2), 10 mM MgCl2, and 0.2% bovine serum albumin at 12 degrees (1) is rapid, saturable, and reversible; (2) increases as a function of ligand or membrane concentration, time, and temperature; and (3) optimally fits to a two-site (high and low affinity) model. The equilibrium dissociation constant (0.15 +/- 0.03 nM) and binding site concentration (28.7 +/- 8.1 fmol/mg protein) of the high affinity site as well as association (0.055 nM-1.min-1) and dissociation (0.0122 min-1) rate constants are similar to those of mammalian vascular ANG II receptors. Both 125I-[Ile5]ANG II and 125I-[Val5]ANG II are competitively displaced by unlabeled ANG II. These results suggest that specific ANG II receptors exist in the fowl aorta.

Among 20 species of freshwater fishes examined, Pseudorasbora parva, Rhodeus ocellatus, Cobitis anguillicaudatus, Carassius auratus, Oryzias latipes, Gambusia affinis, and Gyrinocheilus anymonieri were found to drink water like seawater fishes, while 13 remaining species did not drink. For fish species found exclusively in fresh water, angiotensin II (AII) treatment did not induce drinking. In contrast, those freshwater fishes which survive in estuarine brackish water (Leuciscus hakonensis, C. carassius, Parasilurus asotus, G. affinis, Chaenogobius annularis, Tridentiger obscurus, and G. anymonieri responded to AII by drinking. Furthermore, some freshwater fishes which survive either in hypertonic water (C. auratus) or in sea water (Anguilla japonica and O. latipes) also responded to AII by drinking. Of 17 seawater fishes examined, Eptatretus burgeri, Triakis scyllia, and Heterodontus japonicus failed to drink water, and for Trachurus japonicus, Platichthys bicoloratus, and Glossogobius giuris fasciatopunctatus, water intake was minor (similar to freshwater fishes). The 11 remaining seawater fishes drank water. AII did not induce drinking in fishes living exclusively in sea water. However, seawater fishes which survive either in tide pools (Chasmichthys dolichognathus gulosus) or in brackish water (Sillago japonica, Mugil cephalus, G. giuris fasciatopunctatus) responded to AII by drinking. P. bicoloratus, Acanthopagrus schlegeli, and Fugu niphobles were exceptional, in that they survive in brackish water, but did not respond to AII. Although some exceptions exist, it is generally concluded that a drinking response to AII is characteristic of fishes which encounter water more hypertonic than that in which they typically reside. Accordingly, a drinking mechanism induced by AII may be a compensatory emergency reaction to dehydration stress.