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Cytochemical demonstration of hydroxysteroid dehydrogenase activity in mouse oocytes and preimplantation embryos
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149
SPERM-EGG INTERACTIONIN THE ASCIDIANCIONA I_NTESTtNALISBIOCHEMICALCHARACTERIZATIONOF THE MOLECULESINVOLVEDIN THE BINDING M,R
VITELLOGENESIS IN XENOPUS AND BUFO: ULTRASTRUCTURE OF YOLK-PLATELET MEMBRANES H . - P e t e r R~chter, Dept. of Physiology, Univ. Saorland, D-6650 Homburg, F.R.Germony The architecture of yolk-plotelet membranes has been studied by thin section and the first time by freeze-fracture electron microscopy. In Bufo morinus oocytes a novel type of yolk formation is found. During vitellogenesis numerous lipoid droplets are attached to the periphery of the crystalline plate]et core, indicating on intense uptake of lipids. The entire lipo-proteinyolk complex is enveloped by a surprisingly smooth membrane which shows ring-like arrangements of IMPs only at the areas of underlying liposomes. In Xenopus laevis oocytes throughout vite]logenesis large yolk platelets ore in tight contact with smaller prlmordial yolk precursors that con be completely incorporated into the main core of the plotelet. Fusion of endoplosmic vesicles with the membrane of nascent yolk is neither observed on thin sections nor on freeze-fracture replicas, indicating no uptake of yolk-containing vesicles into the yolk orgonelle. This yolk plotelet membrane represents IMPs of various size and densities of 200-500 IMPs/)Jm~ at P-faces and 1200-2000 IMPs/jJm2 at E-faces.
Pinto'. G, Casazza* and R De Santis* "Institute of Protein Biochemistry and Enzymology, CNR. Arco Felice. Napoli and *Stazione Zoologica di Napoli, Villa Comunale, Napoli, Italy In Ciona intestinalis the species-specific binding of the spermatozoa to the egg is a prerequisite for fertilization This interaction occurs between the vitelline coat of the egg and the plasma membrane of the tip of the sperm head Fucosyl-proteins with sperm- receptor activity have been isolated from the vitelline coat these glycoproteins are able to inhibit the binding of the spermatozoa to the vitelline coat and to elicitsperm activation and the acrosome reaction A family of high molecular weight glycopetides, obtained by exhaustive pronase digestion of the fucosyl-protoins, retains the receptor activity, but fails to activate spermatozoa. The specificity of monoclonal antibodies raised against these glycopeptides has been checked The plasma membrane of the sperm head that is involved in the interaction with the vitelline coat.
shows binding sites for the lectin. Concanavalin A Proteins from the spermatozoa plasma m e m b r a n e have been isolatedand the Concanavalin A-binding proteins have been identified Their role in the interaction with the egg has been studied
151 PH IN THE JELLY LAYER OF STARFISH OOCYTES. A. de Santis, C.Ciccarelli I, P. Russo and B. Dale. Stazione Zoologica, Villa Comunaie, 8-~21 Naples, IIstituto di Istologia, Universit~ di Roma, Via Scarpa 4, OOIOO Roma Italy.
CYTOCH~AICAL DEMONSTRATION OF HYDROXYSTEROID DEHYDROGENASE ACTIVITY IN MOUSE OOCYTES AND PREIMPLANTATION EMBRYOS G.G.De Schepper and C.J.F.Van Noorden. Academic Medical Centre, University of Amsterdam, The Netherlands 3~ -Hydroxysteroid dehydrogenase (3~ -HSD) is a key enzyme in steroidogenesis. Steroids are supposed to play an important role in morula to blastocyst transformation and in implantation. Until now 3~ -HSD activity has been demonstrated qualitatlvely by cytochemical means in preimplantatlon embryos from rabbit, rat, hamster and mouse. The present study reveals that 3~ -HSD activity, as determined cytochemlcally, can be quantified in units activity per embryo. With the use of dehydroeplandrosteron as suhstrate, activity was not demonstrated in preovulatory oocytes, but a sharp rise in activity was found in ovulated (unfertilized) eggs and a steady decline from the two-cell stage onwards.
Using liquid ion exchanger micro-electrodes we have studied the pH in the vicinity of starfish oocytes and eggs and also in their jelly layers. Although highly variable, the pH in these zones was consistently~wer (6.3-7.6) than the pHof sea water (7.8-8.2). Aged oocytes/eggs presented more acid environments than fresh oocytes. The pH indicator Phenol Red shows that the ovarian fluid is more acid than seawater, ranging from 6.0 to 7.5. Our data suggests that starfish oocytes, from their time in the ovary until several hours after their release, are surrounded by an acid environment. This micro-environment may play a role in metabolic repression in the ovary and in the regulation of sperm-e~g interaction following spawning.
54S
149
SPERM-EGG INTERACTIONIN THE ASCIDIANCIONA I_NTESTtNALISBIOCHEMICALCHARACTERIZATIONOF THE MOLECULESINVOLVEDIN THE BINDING M,R
VITELLOGENESIS IN XENOPUS AND BUFO: ULTRASTRUCTURE OF YOLK-PLATELET MEMBRANES H . - P e t e r R~chter, Dept. of Physiology, Univ. Saorland, D-6650 Homburg, F.R.Germony The architecture of yolk-plotelet membranes has been studied by thin section and the first time by freeze-fracture electron microscopy. In Bufo morinus oocytes a novel type of yolk formation is found. During vitellogenesis numerous lipoid droplets are attached to the periphery of the crystalline plate]et core, indicating on intense uptake of lipids. The entire lipo-proteinyolk complex is enveloped by a surprisingly smooth membrane which shows ring-like arrangements of IMPs only at the areas of underlying liposomes. In Xenopus laevis oocytes throughout vite]logenesis large yolk platelets ore in tight contact with smaller prlmordial yolk precursors that con be completely incorporated into the main core of the plotelet. Fusion of endoplosmic vesicles with the membrane of nascent yolk is neither observed on thin sections nor on freeze-fracture replicas, indicating no uptake of yolk-containing vesicles into the yolk orgonelle. This yolk plotelet membrane represents IMPs of various size and densities of 200-500 IMPs/)Jm~ at P-faces and 1200-2000 IMPs/jJm2 at E-faces.
Pinto'. G, Casazza* and R De Santis* "Institute of Protein Biochemistry and Enzymology, CNR. Arco Felice. Napoli and *Stazione Zoologica di Napoli, Villa Comunale, Napoli, Italy In Ciona intestinalis the species-specific binding of the spermatozoa to the egg is a prerequisite for fertilization This interaction occurs between the vitelline coat of the egg and the plasma membrane of the tip of the sperm head Fucosyl-proteins with sperm- receptor activity have been isolated from the vitelline coat these glycoproteins are able to inhibit the binding of the spermatozoa to the vitelline coat and to elicitsperm activation and the acrosome reaction A family of high molecular weight glycopetides, obtained by exhaustive pronase digestion of the fucosyl-protoins, retains the receptor activity, but fails to activate spermatozoa. The specificity of monoclonal antibodies raised against these glycopeptides has been checked The plasma membrane of the sperm head that is involved in the interaction with the vitelline coat.
shows binding sites for the lectin. Concanavalin A Proteins from the spermatozoa plasma m e m b r a n e have been isolatedand the Concanavalin A-binding proteins have been identified Their role in the interaction with the egg has been studied
151 PH IN THE JELLY LAYER OF STARFISH OOCYTES. A. de Santis, C.Ciccarelli I, P. Russo and B. Dale. Stazione Zoologica, Villa Comunaie, 8-~21 Naples, IIstituto di Istologia, Universit~ di Roma, Via Scarpa 4, OOIOO Roma Italy.
CYTOCH~AICAL DEMONSTRATION OF HYDROXYSTEROID DEHYDROGENASE ACTIVITY IN MOUSE OOCYTES AND PREIMPLANTATION EMBRYOS G.G.De Schepper and C.J.F.Van Noorden. Academic Medical Centre, University of Amsterdam, The Netherlands 3~ -Hydroxysteroid dehydrogenase (3~ -HSD) is a key enzyme in steroidogenesis. Steroids are supposed to play an important role in morula to blastocyst transformation and in implantation. Until now 3~ -HSD activity has been demonstrated qualitatlvely by cytochemical means in preimplantatlon embryos from rabbit, rat, hamster and mouse. The present study reveals that 3~ -HSD activity, as determined cytochemlcally, can be quantified in units activity per embryo. With the use of dehydroeplandrosteron as suhstrate, activity was not demonstrated in preovulatory oocytes, but a sharp rise in activity was found in ovulated (unfertilized) eggs and a steady decline from the two-cell stage onwards.
Using liquid ion exchanger micro-electrodes we have studied the pH in the vicinity of starfish oocytes and eggs and also in their jelly layers. Although highly variable, the pH in these zones was consistently~wer (6.3-7.6) than the pHof sea water (7.8-8.2). Aged oocytes/eggs presented more acid environments than fresh oocytes. The pH indicator Phenol Red shows that the ovarian fluid is more acid than seawater, ranging from 6.0 to 7.5. Our data suggests that starfish oocytes, from their time in the ovary until several hours after their release, are surrounded by an acid environment. This micro-environment may play a role in metabolic repression in the ovary and in the regulation of sperm-e~g interaction following spawning.
54S