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Starfish and horseshoe crab egg factors cause elevations of cyclic nucleotide concentrations in spermatozoa from starfish and horseshoe crabs.

Factors collected from the eggs of the starfish (Pisaster giganteus) and the horsehoe crab (Limulus polyphemus) caused significant increases in the sperm cyclic nucleotide concentrations of the respective species. Sea urchin egg factors, at concentrations that resulted in maximal cyclic nucleotide elevations in sea urchin spermatozoa, had no effect on those of starfish or horseshoe crab, suggesting a species specificity with respect to egg factor-induced changes in sperm cyclic nucleotide metabolism.

Animals

The effect of myosin antibody on the division of starfish blastomeres.

Antiserum against starfish egg myosin was produced in rabbits. Antibody specificity to myosin was demonstrated by Ouchterlony's immunodiffusion test and by immunoelectrophoresis in the presence of sodium dodecylsulfate (SDS). The latter technique showed that the antibody binds to both heavy and light chains of egg myosin. Furthermore, the antibody reacted with starfish sperm mysosin and starfish adult muscle myosin at both the heavy and light chains. It did not react with bovine platelet mysosin or rabbit skeletal muscle myosin in Ouchterlony's test; however, a weak reaction was observed in the presence of SDS between the antibody and these myosin heavy chains. Ca- and Mg-ATPase activities of egg myosin were not affected by the antibody, but it did inhibit actin-activated ATPase activity of egg myosin. Microinjection of the antibody into blastomeres of starfish eggs at the two-cell stage was carried out. Anti-egg myosin gamma-globulin inhibited the subsequent cleavages at an amount of more than 0.3 ng when injected at interphase. The inhibition was reduced when the injection was carried out near the initiation of cleavage. At the onset of the second cleavage the antibody was not inhibitory; however, an appropriate amount inhibited the third cleavage. Although the disappearance of the nuclear membrane was observed in the presence of the antibody, the formation of the mitotic apparatus was more or less disturbed. However the formation of daughter nuclei seemed to be scarcely affected by the antibody except that the distance between the nuclei was significantly smaller than normal.

Actins

The starfish (Patiria and Pisaster) oocyte: a model system for the study of hormone-surface interaction in oogenesis.

Studies were made to determine if the starfish (Patiria and Pisaster) oocyte could serve as a model system for investigations of hormone-surface interactions in eucaryotic cells. Treatment of starfisch oocytes with maturation inducing hormone, 1-methyladenine, was observed by scanning electron microscopy to induce surface alteration. The action of the hormone was dependent on the presence of the cation Ca++. Although caffeine, theophylline, and theobromine supposedly inhibit maturation of oocytes, studies using the starfish oocyte showed that theobromine does not inhibit maturation and the inhibition caused by caffeine and theophylline is reversible. Other studies using the starfish oocyte and tritiated 1-methyladenine showed that the hormone does not dissociate from the oocyte once bound, and that the oocyte surface may have multiple receptors for 1-methyladenine. From these studies it was concluded that the starfish oocyte is a useful model for studies of hormone-surface interactions in eucaryotic cells.

Adenine

Purification of a carboxypeptidase B-like enzyme from the starfish Dermasterias imbricata.

A carboxypeptidase B-like enzyme which catalyses the hydrolysis of synthetic esters of lysine and arginine has been isolated from the starfish Dermasterias imbricata. This carboxypeptidase B-like enzyme has a molecular weight of approximately 34 000 and shares this and other properties with bovine pancreatic carboxypeptidase B. The existence of zymogen for this activity in the pyloric caeca of the starfish is demonstrated. This zymogen has a molecular weight near 40 000 and appears to be analogous to other monomeric procarboxypeptidases B. The zymogen possesses an intrinsic low-level activity toward synthetic substrates of carboxypeptidase B and is activated by trypsin.

Animals

Nucleotide sequence of starfish initiator tRNA.

The nucleotide sequence of starfish ovary initiator tRNA was determined to be pA-G-C-A-G-A-G-U-m1G-m2G-C-G-C-A-G-U-G-G-A-A-G-C-G-U-G-C-U-G-G-G-C-C-C-A-U-t6A-A-C-C-C-A-G-A-G-m7G-D-m5C-C-G-A-G-G-A-psi-C-G-m1A-A-A-C-C-U-C-G-C-U-C-U-G-C-U-A-C-C-AOH. The sequence was determined by a combination of the two different post-labeling techniques. Two-dimensional cellulose thin-layer chromatography was adopted for analysis of 5'-terminal nucleotides of tRNA fragments produced by formamide treatment. The nucleotide sequence of starfish initiator tRNA is very similar to that of mammalian cytoplasmic initiator tRNAs, but has seven different nucleotide residues and two modifications: residue 55 is psi instead of U, and residue 26 is unmodified G instead of m2G.

Amino Acyl-tRNA Synthetases

Studies on the glycosphingolipids of the starfish, Asterina pectinifera. II. Isolation and characterization of a novel ganglioside with an internal sialic acid residue.

1. Three gangliosides, provisionally named Gangliosides 1, 2, and 3, were obtained from the lipid extract of the starfish, Asterina pectinifera by silicic acid, DEAE-Sephadex, and Iatrobeads column chromatography. The most abundant, Ganglioside 3 (37.7 microgram/g wet weight of starfish) was isolated in the pure state and its chemical structure was studied. 2. The sugar composition of Ganglioside 3 consisted of arabinose, glucose, galactose, and sialic acid (as N-glycolylneuraminic acid) in a molar ratio of 1:1:3:1. Three sialic acid-containing oligosaccharide fragments were isolated from partial acid hydrolysates of the ganglioside by Dowex 1 X 8 (acetate form) column chromatography and preparative paper chromatography, and identified as Gal leads to NeuGc, Gal (1 leads to 4)[Gal(1 leads to 8)]NeuGc and Ara-(1 leads to 6)Gal(1 leads to 4)[Gal(1 leads to 8)]NeuGc. 3. The structure of Ganglioside 3 was postulated to be: Araf,p(1 leads to 6)Galpbeta(1 leads to 4)[Galpbeta(1 leads to 8)]NeuGc(2 leads to 3)Galpbeta(1 leads to 4)Glcpbeta(1 leads to 1)-ceramide. This is a unique structure with the sialic acid residue internally located in the sugar chain. 4. The ganglioside contained saturated 2-hydroxy fatty acids ranging in length from C16 to C24, among which C22, C23, and C24 acids were predominant. The long-chain bases consisted exclusively of C16, C17, and C18 phytosphingosines of iso and anteiso types.

Carbohydrates

An RPA-assisted homogeneous electrochemical DNA sensor for on-site eDNA detection toward early warning of crown-of-thorns starfish outbreaks.

Crown-of-thorns starfish (COTS) outbreaks seriously threaten coral reef ecosystems, while conventional monitoring approaches are time-consuming and often lack sufficient sensitivity for early warning. Existing electrochemical DNA sensors usually require complex electrode-surface immobilization procedures, which can lead to uneven probe distribution, significant steric hindrance, and poor stability. Meanwhile, the low concentration of environmental DNA (eDNA) in marine environments further complicates detection. To overcome these challenges, this study developed a homogeneous electrochemical DNA sensor assisted by recombinase polymerase amplification (RPA) for COTS eDNA detection. Target DNA was first amplified by RPA, and the amplification products were then hybridized in solution with capture probe (CP)-modified magnetic beads (MB) and biotin-labeled signal probe (SP) to form sandwich-structured MB complexes. These complexes were subsequently magnetically enriched and immobilized on the electrode surface for electrochemical signal readout. Under optimized conditions, the sensor displayed a linear response to COTS genomic DNA from 3.77 fg/μL to 1 ng/μL, with an LOD of 2.02 fg/μL and an LOQ of 3.77 fg/μL. The sensor was applied to Xisha Islands samples, and the results agreed with droplet digital PCR (ddPCR) (P > 0.05), demonstrating its potential for sensitive and reliable on-site COTS eDNA detection.

Animals

A reversible, hydrogen ion blockade of spontaneous oocyte maturation in the starfish: locus of action.

Starfish (Asterias forbesi) oocytes encased within their follicle cells mature spontaneously during a portion of the normal reproductive period when released from the ovary into seawater. A previous report has shown that oocytes isolated in acidic seawater do not mature spontaneously but retain the capacity to do so when returned to normal seawater. The object of this study was to determine the mechanism by which acidic pH reversibly blocks spontaneous oocyte maturation in isolated follicles. Incidence of spontaneous oocyte maturation in follicles isolated in acidic seawater decreased as pH decreased from 7 to 4. Oocytes in which spontaneous maturation was inhibited (ASW at pH 4.7 TO 5.4) underwent germinal vesicle breakdown with the addition of 1-methyladenine. Oocytes isolated in acidic seawater (pH 4 or 5) with intact follicle cells matured spontaneously when transferred immediately to normal seawater pH 8); after four hours, 60-65% of the follicles incubated in seawater at pH 5 matured spontaneously when returned to normal seawater as compared to less than 10% of the follicles maintained at pH 4. Inhibition of spontaneous maturation was not reversible in the absence of the follicle cells. Oocytes isolated in acidic seawater with their follicle cells did not spontaneously mature when transferred to calcium-free seawater at pH 8. The results obtained support the hypothesis that acidic seawater reversibly inhibits spontaneous oocyte maturation by interfering with the release of meiosis-inducing substance from the follicle cells.

Animals

Voltage clamp analysis of two inward current mechanisms in the egg cell membrane of a starfish.

Ionic mechanisms of excitation were studied in the immature egg cell membrane of a starfish, Mediaster aequalis, by analyzing membrane currents during voltage clamp. The cell membrane shows two different inward current mechanisms. One is activated at a membrane potential of -55 approximately -50 mV and the other at -7 approximately -6 mV. They are referred to as channels I and II, respectively. A similar difference is also found in the membrane potential of half inactivation. Currents of the two channels can, therefore, be separated by selective inactivation. The currents of both channels depend on Ca++ (Sr++ or Ba++) but only the current of channel I depends on Na+. The time-course of current differs significantly between the two channels when compared at the same membrane potential. The relationship between the membrane current and the concentration of the permeant ions is also different between the two channels. The result suggests that channel II is a more saturable system. The sensitivity of the current to blocking cations such as Co++ or Mg++ is substantially greater in channel II than in channel I. Currents of both channels depend on the external pH with an apparent pK of 5.6. They are insensitive to 3 muM tetrodotoxin (TTX) but are eliminated totally by 7.3 mM procaine. The properties of channel II are similar to those of the Ca channel found in various adult tissues. The properties of channel I differ, however, from those of either the typical Ca or Na channels. Although the current of the channel depends on the external Na the amplitude of the Na current decreases not only with the Na concentration but also with the Ca concentration. No selectivity is found among Li+, Na+, Rb+, and Cs+. The experimental result suggests that Na+ does not carry current but modifies the current carried by Ca in channel I.

Action Potentials

Isolation of myosin from starfish sperm heads.

Myosin was extracted and partially purified from the head portion of spermatozoa of the starfish, Asterias amurensis. The sperm myosin showed a specific Ca2+-activated ATPase [EC 3.6.1.3] activity of 0.2 mumoles Pi/min/mg at high ionic strength and pH 6.5. It resembled egg myosin in forming thick filaments, becoming attached to actin filaments. subunit composition, and serological properties.

Adenosine Triphosphatases

Studies on the glycosphingolipids of the starfish, Asterina pectinifera. III. Isolation and structural studies of two novel gangliosides containing internal sialic acid residues.

Two gangliosides, provisionally named Gangliosides 1 and 2 in previous studies, were isolated from starfish, Asterina pectinifera by silicic acid, DEAE-Sephadex, and Iatrobeads column chromatography procedures, and preparative thin-layer chromatography, and their structures were established. On the basis of the results of partial acid hydrolysis, methylation and oxidation with chromium trioxide, Gangliosides 1 and 2 were proposed to be Ara p beta(1 leads to 6)Gal p beta(1 leads to 4)8-O-MeNeuGc(2 leads to 3)Gal p beta(1 leads to 4)Glc p beta(1 leads to 1)-ceramide and Ara p beta(1 leads to 6)Gal p beta(1 leads to 4)NeuGc(2 leads to 3)Gal p beta(1 leads to 4)Glc p beta(1 leads to 1)-ceramide, respectively. The ceramide moieties of both gangliosides had similar phytosphingosine and 2-hydroxy fatty acid compositions, and both gangliosides were structurally related to the previously described Ganglioside 3.

Animals

Partial purification and properties of phospholipase A2 from the starfish, Asterina pectinifera.

1. Phospholipase A2 (phosphatide 2-acyl-hydrolase, EC 3.1.1.4) activity was shown to occur in the supernatant fraction of a freshly prepared homogenate from the pyloric caecum of starfish (Asterina pectinifera). 2. The phospholipase A2 has been isolated and purified 130-fold by ultracentrifugation, ammonium sulfate precipitation and column chromatographic procedures. 3. The purified enzyme was stable to heat at low pH values and the optimal pH was observed at approximately 9.0. 4. The enzyme activity was activated by Ca2+ and sodium deoxycholate, and was inhibited by EDTA.

Animals

Relationships between hormone-induced calcium release and 86rubidium uptake stimulation in starfish oocytes.

86Rubidium+ uptake, but not 86Rubidium efflux, is strongly stimulated after addition of the meiosis inducing hormone 1-methyladenine (1-MeAde) to prophase blocked oocytes of the starfish Marthasterias glacialis. This stimulation is a transient process which does not require the continuous presence of 1-MeAde and is elicited within 1 minute of contact. 1-MeAde and its biologically active structural analogs fully stimulate Rb+ uptake at concentrations which are about two orders of magnitude lower than those required to trigger meiosis reinitiation but which already release underthreshold levels of Ca2+ from the inner part of the plasma membrane. External Ca2+ concentrations effective in triggering meiosis reinitiation also stimulate Rb+ influx, while drugs like D600, theophyllin and caffein which suppress the hormone induced Ca2+ release, simultaneously preclude the stimulation of Rb+ uptake. Dithiothreitol (DTT) which mimicks 1-MeAde action in releasing Ca2+ and inducing meiosis acts both on the efflux and on active and passive Rb+ influxes. Ouabain, the classical inhibitor of the Na+, K+ pump does not preclude meiosis reinitiation under the influence of 1-MeAde, its agonists of mimetics. It suppresses the active component of Rb+ uptake both in control or stimulate oocytes. When applied only in preincubation before starting the hormone treatment, it cannot however inhibit the stimulation of Rb+ uptake, while basal pump inhibition is preserved. These results demonstrate that stimulation of the active Rb+ or K+ transport is not indispensable to meiosis reinitiation. They suggest moreover that the hormone induced Ca2+ release from the plasma membrane may be responsible for unmasking new ouabain sensitive transport sites.

Adenine