Amyda Japonica Research Articles

EFFECTS OF ACETYLCHOLINE, VAGAL STIMULATION AND TYRAMINE ON THE ISOLATED ATRIA OF THE TORTOISE.

Abstract In the vagus-atrial preparation of the tortoise (Amyda japonica), a cold-blooded animal, acetylcholine and vagal stimulation, in the presence of atropine, produced positive inotropic effects which were inhibited by dichloroisoprenaline or pronethalol. Hexamethonium blocked the excitatory effect of acetylcholine but only slightly inhibited that of vagal stimulation. Tyramine exerted a positive inotropic action in this preparation, but had little effect on atria prepared from reserpine-treated animals. Tyramine was shown to release a substance from the tortoise atria which caused contraction of aortic strips taken from reserpine-treated rabbits, and which appeared to be a catecholamine.

Localization of Receptors for Natriuretic Peptide and Endothelin in the Duct of the Epididymis of the Freshwater Turtle

The presence of receptor subtypes for natriuretic peptides (NPs) and endothelin (ET) in the epididymis of the freshwater turtle, Amyda japonica, was examined by quantitative in vitro autoradiography using iodinated mammalian-type atrial NP (125I-ANP(1–28)), phylogenically conserved C-type NP (125I-[Tyr0]-CNP(1–22)), and ET-1 (125I-ET-1) as radiolabeled ligands. To characterize NP receptor (NPR) subtypes, we also performed an activation of particulate guanylyl cyclase (GC) in membranes of the epididymis by NPs. Specific 125I-ANP(1–28) and 125I-[Tyr0]-CNP(1–22) bindings were localized in surrounding smooth muscle cell layer of the duct of the epididymis with an apparent dissociation constant (Kd) of 0.84 ± 0.15 and 1.74 ± 0.39 nM and a maximal binding capacity (Bmax) of 0.47 ± 0.11 and 0.08 ± 0.01 fmol/mm2, respectively. Bindings of 125I-ANP(1–28) and 125I-[Tyr0]-CNP(1–22) to these sites were also displaced by des[Gln18,Ser19,Gly20, Leu21,Gly22]ANF(4–23), a specific ligand of the NP clearance receptor. Production of 3′,5′-cyclic guanosine monophosphate by particulate GC in membranes of the epididymis was stimulated by ANP(1–28), BNP(1–26), and CNP(1–22). Receptor subtypes for ET in the epididymis were characterized by competition with BQ 123 and BQ 788 as specific antagonists for ET receptors, type A (ETA) and type B (ETB) subtypes, respectively. Specific 125I-ET-1 bindings were localized in the smooth muscle cell layer of the duct of the epididymis with Kd and Bmax of 0.21 ± 0.03 nM and 0.52 ± 0.05 fmol/mm2, respectively. These specific bindings were potently inhibited in a dose-dependent manner by BQ 123, whereas BQ 788 (10 μM) was not in competing for specific 125I-ET-1 bindings in this structure. Therefore, these results indicate that specific NP and ET receptors are localized in surrounding smooth muscle cells of the duct of the epididymis of the freshwater turtle. It is also suggested that biological and clearance NPR-like subtypes coexist in these cells, and the predominant ET receptor subtype in this tissue is the ETA-like receptor. The localization of specific receptors for NPs and ET in the epididymis may be involved in the control of the transport of sperm in the freshwater turtle.

Receptor subtypes for endothelin in the kidney of the freshwater turtle (amyda japonica)

The distribution of receptor subtypes for endothelin (ET) in the kidney of the freshwater turtle, Amyda japonica, was examined by quantitative in vitro receptor autoradiography using iodinated mammalian type ET‐1 (125I‐ET‐1) as a radiolabeled ligand. Specific 125I‐ET‐1 bindings were localized to renal tubules, renal arteries and ureter with binding densities of 111.21 ± 19.14, 182.13 ± 10.57 and 219.46 ± 12.83 amol/mm2, respectively. Binding dissociation constants in renal tubules, renal arteries and ureter were 1.05 ± 0.63, 2.03±0.56 and 1.70±0.47nM, respectively. Receptor subtypes for ET in the kidney were characterized by competition with BQ 123 and BQ 788 as specific antagonists for ET receptors, type A (ETA), and type B (ETB) subtypes, respectively. Specific 125I‐ET‐1 bindings in renal arteries and ureter were potently inhibited by BQ 123 in a dose‐dependent manner, whereas BQ 788 was not in competing for specific 125I‐ET‐1 bindings in this structure. However, specific 125I‐ET‐1 bindings in renal tub...

Structure and stereochemistry of the higher bile acid isolated from turtle bile: (22S,25R)-3 alpha,12 alpha,15 alpha,22-tetrahydroxy-5 beta-cholestan-26-oic acid.

The structure and stereochemistry of the higher bile acid, tetrahydroxyisosterocholanic acid (TISA), which was previously isolated from the bile of Amyda japonica (turtle) and proposed as a tetrahydroxyisosterocholanic acid, have been established as (22S,25R)-3 alpha,12 alpha,15 alpha,22-tetrahydroxy-5 beta-cholestan-26-oic acid by X-ray crystallographic analysis of its ethyl ester.

Overlapping Distribution of Receptors for Atrial Natriuretic Peptide and Angiotensin II in the Kidney and the Adrenal Gland of the Freshwater Turtle,Amyda japonica

The distribution of receptors for atrial natriuretic peptide (ANP) and angiotensin II (ANG II) in the kidney and adrenal gland of the freshwater turtle,Amyda japonica,was examined by quantitativein vitroautoradiography using125I-labeled rat (r)ANP1–28and125I-labeled human ANG II as labeled ligands. Receptor subtypes were characterized by competition with des[Gln18,Ser19,Gly20, Leu21, Gly22] ANP(4–23)(C-ANP) as a selective ligand of the clearance receptors to specific125I-rANP(1–28)binding, and with DuP 753 and PD 123319 as nonpeptide antagonists for ANG II receptors type 1 (AT1) and type 2 (AT2) subtypes, respectively, to125I-ANG II binding. Specific125I-rANP(1–28)binding with a single binding site was found overlying glomeruli of the kidney and the outer zone of the adrenal gland with dissociation constants (Kd) of 4.39 ± 0.33 and 6.07 ± 1.36 nM,respectively. C-ANP (10 μM) inhibited about 90% of the glomerular and adrenal125I-rANP(1–28)binding. Specific125I-ANG II binding was also localized in the glomeruli of the kidney and the outer zone of the adrenal gland withKdof 1.02 ± 0.22 and 0.37 ± 0.04 nM,respectively. DuP 753 (10 μM) potently inhibited about 80% of glomerular and 90% of adrenal125I-ANG II binding, whereas PD 123319 (10 μM) was very weak in competing for specific125I-ANG II binding in both tissues. Therefore, these results indicate that specific ANP and ANG II receptors with high affinities have an overlapping distribution in glomeruli of the kidney and the outer zone of the adrenal gland of the freshwater turtle. They also suggest that biological and clearance ANP receptor-like subtypes coexist in both tissues, and the predominant ANG II receptor subtype in these tissues is the AT1-like receptor. The overlapping distribution of specific receptors for both peptides in these tissues provides the basis for possible receptor interactions to exert a functional antagonism between ANP and ANG II in the freshwater turtle.

Specific binding sites for atrial natriuretic peptide in the freshwater turtle, Amyda japonica

Specific finding sites for atrial natriuretic peptide (ANP) were investigated by in vitro autoradiographic techniques in various tissues of the freshwater turtle, Amyda japonica. A high density of binding sites for 200 pM of 125I-labelled rANP (1–28) was located in the glomeruli of the kidney and the cortical portion of the adrenal gland. A moderate density of binding sites was seen in the arachnoid matter and choroid plexus of the third and lateral ventricles of the brain and the epididymis. A low density of binding sites was revealed in lamina propria of the mucosa of stomach and intestine, the seminiferous tubules of testes, and the epithelial layer of oviduct. In the presence of excess unlabelled rANP (1–28) (1 μM), binding to these structures were completely displaced. Therefore, specific ANP receptors exist in the kidney, adrenal gland, stomach, intestine, oviduct, epididymis, seminiferous tubules and brain. The ANP system may be involved in physiological regulatory function in the freshwater turtle, Amyda japonica.

Renal and hormonal responses to atrial natriuretic peptide and turtle atrial extract in the freshwater turtle, Amyda japonica

AbstractThe effects of synthetic atrial natriuretic peptide and turtle atrial extract on cardiovascular and renal functions were examined in the freshwater turtle, Amyda japonica.Both the atrium and ventricle of the turtle heart contained an immunoreactive atrial natriuretic peptide. Synthetic rat atrial natriuretic peptide (atriopeptin III) and turtle atrial extract caused a dose‐dependent decrease in mean arterial blood pressure. In turtles that received either atriopeptin III or turtle atrial extract, no significant changes in renal function were observed. However, furosemide was effective in altering renal function. Levels of aldosterone in plasma decreased after 2 hours and increased after 24 hours following injection of atriopeptin III, although the plasma renin concentration did not change.We suggest that the heart of the freshwater turtle may contain an immunoreactive atrial natriuretic peptide and that the atrial natriuretic peptide is more important in the regulation of blood pressure than in the regulation of renal function.

Circannual changes in renin concentration, plasma electrolytes, and osmolality in the freshwater turtle

Seasonal changes in heart rate, blood pressure, plasma and renal renin concentrations, plasma electrolyte concentrations, and osmolality were studied in freshwater turtles ( Amyda japonica) exposed to the natural environment. Seasonal changes in heart rate and blood pressure showed a marked increase in summer and a decrease in winter, which correlated directly with that of an environmental temperature. Two peaks in plasma renin concentration were observed in November and May and correlated with the preparation for hibernation and arousal periods. Renal renin content exhibited a marked seasonal variation characterized mainly by an increase in summer, fall, and winter and a decrease in spring. Plasma sodium concentration and osmolality were the highest in summer, the lowest in winter, and intermediate during spring and fall. A change in plasma osmolality correlated closely with that of temperature. However, no marked seasonal variations in the concentrations of plasma potassium, chloride, and calcium were observed. On the basis of these data, we suggest that a circannual rhythm of renin concentration may be related to hibernation. We also suggest that seasonal changes in sodium concentration and osmolality relate closely to environmental temperature, although the mechanisms are not clear.

Radioimmunoassay and characterization of renin‐angiotensin system in the fresh‐water turtle

AbstractWe developed a radioimmunoassay (RIA) for plasma renin concentration (PRC) and characterized the renin‐angiotensin system (RAS) in the fresh‐water turtle (Amyda japonica).Among three antisera to human angiotensin I (ANG I) had different specificities to ANG I analogues; the antiserum with the highest specificity was used to measure the PRC. The amount of ANG I generated from incubation of turtle renal extracts with turtle plasma was linear with respect to volume of renal extract and incubation time. Turtle renin cleaved heterologous substrates from nephrectomized rats and rabbits but had a higher affinity for renin substrate from rabbits. The generation of ANG I with heterologous substrate was temperature dependent (optimum‐37°C) and the pH optimum was at 5.0–5.5. Furosemide (5 mg/kg) did not affect PRC. Hemorrhage (1% and 3% of body weight) decreased blood pressure. The PRC did not change with the lower hemorrhage, but increased significantly with massive hemorrhage. Angiotensin II increased blood pressure with no effect in PRC. The PRC of fresh‐water turtles transferred to 4°C increased significantly.The RAS in the fresh‐water turtle seems to be involved in the adaptation of environmental temperature rather than in the acute control of blood pressure although the mechanism is not known.

Characterization of angiotensin I-converting enzyme activity in the freshwater turtle, Amyda japonica

1. 1. Angiotensin I-Converting Enzyme (ACE) activity has been characterized in the freshwater turtle, Amyda japonica. 2. 2. Peak activity of ACE in plasma from the freshwater turtle was shown at pH 9.0, which was more alkaline compared to that of mammals. 3. 3. Chloride requirements for the optimal ACE activity were different from species. 4. 4. ACE inhibitors, EDTA, teprotide (SQ 20,881), Captopril (SQ 14,225) showed dose-dependent inhibitions of ACE activity in plasma from the freshwater turtle as well as mammals. 5. 5. ACE activity was found in several different tissues with a different activity showing the highest activity in kidney homogenate from the freshwater turtle, Amyda japonica.

Electron microscopic study on the hepatic sinusoidal wall of the soft-shelled turtle (Amyda japonica) with special remarks on the smooth muscle cells.

The hepatic sinusoids of the soft-shelled turtle (Amyda japonica) were examined by transmission electron microscopy. The sinusoidal wall was composed of endothelial cells, Kupffer cells and Ito cells. The basal surface of the hepatocyte facing the Disse's space was covered by a continuous basal lamina. In addition to the Ito cells, the Disse's space contains a considerable number of smooth muscle cells. Many of these were distributed sporadically, while others appeared as a sphincter circling the sinusoid. The smooth muscle cells in the Disse's space showed the following features: 1) The nucleus was located eccentrically near one end of the cell. 2) The surface vesicles and pits, mitochondria and dense patches along the myofilament bundles were all sparse as compared with those known from mammalian smooth muscle cells. 3) Cytoplasmic processes or ruffles were protruded into the Disse's space. 4) A weak basal lamina could be recognized. Sinusoidal endothelial cells were characterized by many large electron lucent lysosomes in their perikaryon and by small fenestrae in their attenuated cytoplasm. Ito cells sending out several cytoplasmic processes, possessed a single large lipid droplet on one side of the nucleus. A single cilium budding from the distal centriole into the Disse's space was found in an Ito cell. Extrasinusoidal macrophages were considerably numerous in the soft-shelled turtle liver. Some of the macrophages were apparently migrating into the sinusoid, there to presumably transform into the Kupffer cells.

Structure of the steroidal lactone isolated from turtle bile: (22S,25R)-3α,7α,12α-trihydroxy-5β-cholestano-26,22-lactone

The structure of the steroidal lactone (previously designated as tetrahydroxysterocholanic acid lactone) isolated from the bile of Amyda japonica(turtle) has been characterized as (22S,25R)-3α,7α,12α-trihydroxy-5β-cholestano-26,22-lactone (1). The 22S,25R configuration was determined by 1H n.m.r. comparison with the reference compounds, four possible stereoisomers (with respect to the C-22 and C-25 positions) of 6β-methoxy-3α,5-cyclo-5α-cholestano-26,22-lactones (7)–(10).

Electron microscopic observations of the subcommissural organ in the soft-shelled turtle, Amyda japonica.

The subcommissural organ (SCO) of the soft-shelled turtle was studied with the electron microscope.In the SCO cells, two types of secretory substances are distinguishable; one is represented by secretory vacuoles containing a fine flocculent material which is closely associated with the cisternae of the granular endoplasmic reticulum, and the other by granules with homogeneously dense or finely granular contents which are surrounded by a limiting membrane and believed to be derived from the Golgi apparatus. Both types of secretions are scattered throughout the cytoplasm and are abundant especially in the apical region near the ventricle as well as in the basal processes abutting on capillaries.In the hypendyma large spaces connecting with the perivascular ones are often observed. Their interior is filled with fine flocculent substances similar to the secretions of the SCO cells. In the endothelial cells lining the capillary lumen also, there are vacuoles frequently containing similar dense or flocculent substances.The findings of the perivascular spaces and the endothelial cells described are suggestive of a possible basal secretion from the SCO cells of the soft-shelled turtle.

The relationship between mast cells and histamine in phylogeny with special reference to reptiles and birds.

Various animals from fish to mammals, especially the carp (Cyprinus carpio), turtles (Clemmys japonica and Amyda japonica) and birds (hens, chicks and pigeons) were studied with special reference to the phylogenic relation between mast cells and histamine. Histamine assay (histamine extraction with Code's method and bioassay with guinea pig ileum) in different tissues was combined with a histological study on their mast cell population, and histamine histochemistry with o-phthalaldehyde in p-xylene or ethylbenzene was performed on the spreads and smears.Bioassay revealed an appreciable amount of histamine in tissues rich in mast cells and in the blood of the hen, chick and pigeon, the mast cells, basophiles, thrombocytes and other unknown types of cell distinctly fluoresced with the o-phthalaldehyde method.The blood of the turtles was shown by bioassay to contain a considerable amount of histamine and their blood basophiles exhibited distinct yellow fluorescence. Though the result of the histamine bioassay of turtle tissues was equivocal and only a few mast cells fluoresced, the injection of l-histidine in Clemmys japonica caused an increase in the number of fluorescent mast cells and an increase in the intensity of their fluorescence, suggesting an ability of mast cells to synthesize histamine.Several tissues of the carp contained only a trace of histamine and the mast cells did not fluoresce. Taking into consideration the similar results in the frog (TAKAYA, FUJITA and ENDO, 1967) and newt (TAKAYA, 1968), it is likely that histamine firstly appears in mast cells at the level of reptiles in phylogeny. The amount of histamine in mast cells and basophiles gradually increases in reptiles, birds and mammals.Several histochemical tests for mucopolysaccharides in the granules of mast cells and basophiles showed that the mast cells of all the animals examined, except for the carp whose mast cells were hardly preserved, contained acidic sulfated mucopolysaccharides. The solbuility of the granules in water varied conspicuously from species to species.

スツポンの臓器に於けるVitamin Cの分布

Researches on the distribution of ascorbic acid in animal organs, such as with mammals by PLANT and BULOW, FUJITA and EBIHARA and SVIRBELY, with fish by LUDANY and with invertebrata by OYA and HIROSAWA, and SVIRBELY, were extended to snapping turtle, Amyda japonica. As shown in the annexed table, it was known that among the examined organs, - muscle, lung, pancreas, liver, heart, kidny, spleen and intestine, - spleen is the richest in ascorbic acid, liver and kidny are rather rich, while muscle contains the least.