bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence suggesting that L‐fucose is part of a recognition signal for sperm‐zona pellucida attachment in mammals
Authors: Huang Thomas T. F. , Ohzu Eiji , Yanagimachi R.
Published: February 2005

Experiments were designed to test the effects of simple sugars and complex polysaccharides on the attachment of mammalian spermatozoa with the zona pellucida. In the guinea pig, L‐fucose was a twofold better inhibitor of the attachment compared to other sugars at 50 mM. Fucoidin, an algal polysaccharide rich in sulfated L‐fucose, was a very potent inhibitor, completely blocking attachment at a concentration of 100 μg/ml. Several other highly sulfated glycosaminoglycans showed no inhibitory activity, suggesting the fucoidin effect was not simply due to its charge or sulfate. In addition, fragments of fucoidin, generated by partial hydrolysis and isolated using Biogel P‐2, were nearly as inhibitory as the native molecule on a weight basis. Fucoidin also inhibited sperm‐zona attachment in the hamster and human; thus, its effect is not species specific. The data suggest that L‐fucose may be part of a recognition signal between mammalian gametes.
Sperm tail entry into the mouse egg in vitro
Authors: Gaddum‐Rosse P. , Blandau R. J. , Langley L. B. , Sato K.
Published: February 2005

Cumulus‐free mouse eggs were placed on microscope slides and inseminated with capacitated mouse spermatozoa. Fertilization could then be observed through the phase contrast microscope and recorded by time‐lapse cinematography. Following the penetration of the fertilizing spermatozoon through the zona pellucida and the fusion of the sperm head with the vitelline membrane, the entire sperm tail gradually entered the vitellus. The time required for tail incorporation into the vitellus as measured in 49 eggs varied from 3 h 3 min to 5 h 49 min, with a mean time of 4 h 23 min. When tail incorporation began, the greater part of the flagellum was still outside the zona pellucida; occasionally it slipped into the perivitelline space, but generally it remained outside the zona and shortened by degrees as incorporation proceeded. The motility of the fertilizing spermatozoon declined abruptly very soon after fusion of the sperm head with the vitellus and remained at a very low level during the 3–6 h required for tail incorporation. Sperm motility, therefore, does not appear to be the main determinant in tail incorporation and the primary mechanism responsible for it remains unclear. As the sperm tail slowly entered the vitellus, the second meiotic division was completed with concomitant extrusion of the second polar body. Key stages in second polar body formation were correlated with events in tail incorporation. Differences between fertilization in vitro and in vivo are discussed.
Biochemical and ultrastructural characterizations of nucleoprotamine in human sperm heads treated with micrococcal nuclease and salt
Authors: Tanphaichitr Nongnuj , Sobhon Prasert , Chalermisarachai Panpaka , Chutatape Chutima
Published: February 2005

Preparation of human sperm heads containing mainly nucleoprotamines as their chromatin constituent was described. Acrosome‐depleted human sperm heads were treated with micrococcal nuclease followed with 2 M NaCl. The treatment specifically removed nucleohistone as small chromatin fragments from the human sperm heads. The nuclease‐NaCl‐treated sperm head pellets primarily contained nuclease‐resistant nucleoprotamine as their chromatin. Transmission electron microscopic study showed that there were two populations of the nuclease‐NaCl‐treated human sperm head pellets. One population had thick nucleoprotamine cords of ˜650–1,200 Å in diameter, and the other had thin nucleoprotamine cords of ˜330–420 Å.
A morphologic, cytochemical, and chromatographic analysis of lipid yolk formation in the oocytes of the dog
Authors: Tesoriero John V.
Published: February 2005

The oocytes of carnivorous mammals are distinguished by the presence of large amounts of a lipid, yolk like material. In the oocytes of the dog, lipid yolk formation marks one of the earliest indications of occyte maturation. In early primary oocytes, the yolk bodies are scattered within the ooplasm, while in later stages they are in discrete clusters. Lipid yolk material appears to be formed by at least two mechanisms. Throughout most of oogenesis the oocyte contains scattered dense granular bodies that become vacuolated by droplets of lipid material and may be transformed, by this process, into lipid yolk bodies. These granular bodies are highly reactive for acid phosphatase and are positive for glycoprotein with the PA‐CA‐methenamine technique. In addition, other glycoprotiein‐rich yolk bodies appear to arise from many of the small dictyosomes. In secondary follicles these two mechanisms often appear to act conjointly with the dense vacuolated granules coalesing with the larger yolk bodies. Small yolk bodies are intensely reactive for glycoprotein, becoming less reactive as they enlarge and mature. The developing yolk bodies are often associated with the acid phosphatase‐positive granules. The peripheral portions of the larger yolk bodies are faintly reactive for both acid phosphatase and glycoprotein. All reactivity is lost in mature yolk bodies. Thin layer chromatography of the total lipids extracted from isolated oocytes reveals a pattern that is consistent among dogs of the same and of different breeds. The most abundant lipid fraction from each dog oocyte extraction stains strongly for glycolipid.
Fine structure of the mature spermatozoon of rhynchocinetes typus, crustacea decapoda
Authors: Dupré E. , Barros C.
Published: February 2005

Rhynchocinetes typus spermatozoa obtained from the vas deferens have the shape of a round‐headed nail. The head measures 30 μm in diameter and 14 μm of height. At the center of the flat face of the head emerges a single rigid spike of 53 μm in length. Cross sections of this spike show that it has a wall of 0.4 μm in thickness and a core of 0.6 to 0.8 μm. The outer surface of the spike has a longitudinal striation. When the spermatozoa are placed in sea water it is possible to observe the unfolding of rays. The number of rays in different spermatozoa of the same individual varies from 9 to 13. Each ray is formed by a channel‐like sheath that contains a rigid rod that occupies about 1/3 the length of the ray. This rod has a transverse striation with a periodicity of 185A. The rays are bound among them by a thin membranous sheet that is highly folded in vas deferens spermatozoa. At the distal end of each ray there is a rigid spine of 50 μm in length. The nucleus is coplanar to the radial plane and it extends through the rays. The structure and ultrastructure of R typus spermatozoa depart from that reported for spermatozoa of other Caridea species.
Studies on the polymorphic spermatozoa of a marine snail. 2. Genesis of the eupyrene sperm
Authors: Buckland‐Nicks John , Williams Daphne , Chia Fu‐Shiang , Fontaine Arthur
Published: February 2005

Spermiogenesis of the eupyrene sperm in the snail, Fusitriton oregonensis, was studied with light and electron microscopes. Endoplasmic reticulum, which encircles the nucleus in each spermatid, appears to connect with the Golgi body and to interconnect between adjacent spermatids via cytoplasmic bridges. It is suggested that as the Golgi body migrates around the nucleus the endoplasmic reticulum may circulate with it. The alignment of the proacrosome with the nucleus is effected by a 180° rotation of the Golgi body, after which it separates and migrates posteriorly with the residual cytoplasm. Each sperm possesses a well‐developed intracellular digestive system as indicated by multivesicular bodies, residual bodies, and myeloid figures. Autophagy begins in the residual cytoplasm before it is released from the middle piece. Microtubules are found outside the nucleus and mitochondria during the final stages of spermiogenesis, when elongation is almost complete. These microtubules appear to be involved in the final shaping and twisting process, in which torsion is locked in the nucleus and the mitochondria spiral around the axoneme. The annulus attaches the distal centriole to the plasma membrane in the early spermatid and as flagellar production begins they move towards the implantation fossa at the base of the nucleus. There are two centrioles in the early spermatid, the distal centriole and procentriole. The small procentriole fuses with the distal centriole in the intranuclear canal to form the centriolar cap of the basal body. This cap is pushed through the end of the nuclear tube and is separated from the subacrosomal space by only the nuclear membranes.
Development of membrane excitability in mammalian oocytes and early embryos
Authors: Eusebi Fabrizio , Colonna Rosella , Mangla Franco
Published: February 2005

Developmental aspects of the capability to generate an action potential have been studied in mouse oocytes and early embryos, taking the peak level and the rate of rise of the action potential as parameters of cell excitability. We report here that experimentally induced membrane excitability in mouse oocytes appears throughout oogenetic growth, further develops throughout meiotic maturation, does not undergo main changes at fertilization, and slightly decreases throughout early cleavage, at least up to the four‐cell stage of embryonic development. Calcium is involved in the rising phase of action potential throughout oogenesis, maturation, and early embryonic development.
Reproduction in mice: Spermatozoa as factors in the development and implantation of embryos
Authors: Chaykin Sterling , Watson J. Gary
Published: February 2005

The effects on mouse embryo development in vivo of varying the numbers of spermatozoa used in artificial inseminations was studied. The two criteria used in the evaluation of the progress of embryo development were 1) ability to reach the two‐cell stage and 2) success of development from the two‐cell stage through implantation. A 44% reduction in the yield of two‐cell embryos and a 67% reduction in the number of implants was observed when C3HeB/FeJ females were inseminated with one‐twentieth the number of spermatozoa estimated to be present in a typical ejaculate. The reduction in the yield of two‐cell embryos was substantially reversed by a second insemination of a large number of heat‐inactivated spermatozoa 12 hr after the first insemination. The sperm‐dependent reduction in development from the two‐cell stage through implantation was prevented only by normal viable (unheated) spermatozoa. These results were rationalized by the hypothesis that in female C3HeB/FeJ mice spermatozoa serve physiological functions beyond the fertilization of ova and that spermatozoa may act to foster early embryo development through modulation of the environments embryos experience as they move through the reproductive tract.
Comparison of strains and culture media used for mouse in vitro fertilization
Authors: Ackerman Steven B. , Swanson R. James , Adams Peter J. , Wortham J. W. Edward
Published: February 2005

Success rates of superovulation in response to gonadotropic hormone treatment and in vitro fertilization (ie, mitotic cleavage following insemination) of mouse eggs from outbred CD‐1, hybrid CB6Fl, or hybrid B6CBAF1, mice were compared using either a mouse inseminationmedium, modified Krebs‐Ringer‐bicarbonate (m‐KRB), or a human insemination medium, Ham's F10 nutrient mixture. Inseminations were performed in either organ culture dishes or screw‐top, flat‐side tissue culture tubes. Mean superovulation rates (± SD) were 24.2 (5.1) for CD‐1, 33.0 (5.8) for CB6F1, and 16.3 (6.6) for B6CBAF1 mice. For in vitro cleavage the best combination of mouse strain, insemination medium, and culture container was achieved using CB6F1, mice, m‐KRB medium, and culture tubes. However, Ham's medium used with either hybrid mouse strain was shown to be employable for fertilization of mouse eggs in vitro as a quality control assay and/or experimental model system for testing the human in vitro fertilization procedure.
Dithiothreitol, a disulfide‐reducing agent, inhibits capacitation, acrosome reaction, and interaction with eggs by guinea pig spermatozoa
Authors: Yanagimachi R. , Huang T. T. F. , Fleming A. D. , Kosower N. S. , Nicolson G. L.
Published: February 2005

Capacitation of guinea pig spermatozoa in vitro was inhibited by the disulfide‐reducing agent dithiothreitol (DTT). Even a brief treatment with DTT inhibited capacitation unless an oxidizing agent (glutathione disulfide) was present in the posttreatment medium. Precapacitated spermatozoa were unable to undergo the acrosome reaction in the presence of DTT, indicating that this reagent also blocks the acrosome reaction. Acrosome‐reacted spermatozoa were incapable of attaching to and penetrating the zona pellucida in the presence of DTT. Even when acrosome‐reacted spermatozoa were directly brought to the surface of zona‐free eggs, they were unable to bind to and fuse with the egg plasma membrane so long as DTT was present in the medium. These observations suggest that the tertiary and quaternary structures of sperm surface proteins regulated by their thioldisulfide status are of critical importance in the physiology and function of spermatozoa preliminary to and in the process of fertilization.
Isolation of the inner acrosomal‐nuclear membrane complex from rabbit spermatozoa
Authors: Rahi Hamid , Sheikhnejade Gholamreza , Srivastava Prakash N.
Published: February 2005

Rabbit spermatozoa were passed through a Bio‐Glas 2500 (BG) column to yield pure cytoplasmic droplets and consequently the droplet‐free spermatozoa. Rapid decondensation of sperm nuclei was achieved by 1 mM dithiothreitol (DTT) and 1.5 M NaCl. This treatment caused removal of the plasma and the outer acrosomal membranes. The viscosity of decondensed nuclei was reduced by pancreatic DNase. Further elution of the suspension from BG column yielded a membrane complex. On the basis of electron microscopic observations and the enzyme analysis, these membranes appeared to be the inner acrosomal‐nuclear membrane complex (IANC). The IANC was also prepared from isolated sperm head by DTT‐NaCl and Dnasetreatment and low‐speed centrifugation.
Change in phosvitin kinase activity during early development of the sea urchin
Authors: Fujino Yukio , Yasumasu Ikuo
Published: February 2005

Protein kinase, which phosphorylated phosvitin at the expense of ATP but did not phosphorylate casein, protamine, and histone mixture, was obtained by DEAE‐cellulose column chromatography of the extract from the embryos of the sea urchin, Strongylocentrotus intermedius. This enzyme, partially purified by DEAE‐cellulose column, reversibly catalyzed the reaction of phosvitin phosphorylation. This indicates that the sea urchin embryos contain phosvitin kinase. Phosvitin kinase in sea urchin embryos is somewhat different from that found in the other types of cells, which are able to phosphorylate casein as well as phosvitin. In unfertilized eggs, the activity of this enzyme was found only in the supernatant fraction obtained by centrifuging the homogenate at 10,000g for 20 min. The activity in the embryos at the swimming and the mesenchyme blastula stage was higher than in unfertilized eggs, and was localized in the sedimentable fraction obtained by centrifuging the homogenate of the embryos at 10,000g for 20 min. The highest activity of phosvitin kinase was observed in the embryos at the mesenchyme blastula stage, and the enzyme activity became quite low at the late gastrula stage. The activity and the intracellular distribution of phosvitin kinase changed during the development. The enzyme in this sedimentable fraction was not solubilized with 1% Triton X‐100 but was extracted by 1 M NaCl.
A modified column for sperm isoelectric focusing
Authors: Rahi Hamid , Srivastava Prakash N.
Published: February 2005

A modified U‐shaped column is described for efficient isoelectric focusing of spermatozoa, other cells, and protein. The washed spermatozoa of the rabbit showed a PI of 4.4. After sonication, heads and tails focused at same pH, indicating similar and equal charge.
Effect of compounds structurally related to taurine and of taurine uptake inhibitors on the motility of hamster sperm in vitro
Authors: Gwatkin Ralph B. L.
Published: February 2005

Several taurine‐related compounds, taurine antagonists and taurine uptake inhibitors were tested for their effect on hamster sperm motility in vitro. Hypotaurine was approximately three times more effective than taurine. N‐methyltaurine and taurocyamine were less effective. Inactive taurine‐related compounds were not effective blockers of taurine's spermtimulating activity. However, 1‐(4‐nitrophenyl)‐2‐dimethylaminomethyl‐l‐propenone, a taurine uptake inhibitor, completely suppressed sperm motility at a molar concentration equal to, or less than, that of the taurine added to the incubation medium and also suppressed the motility‐sustaining action of the cumulus oophorus.
Effects of selenium deficiency on the shape and arrangement of rodent sperm mitochondria
Authors: Wallace Edith , Cooper George W. , Calvin Harold I.
Published: February 2005

Selenium‐deficient male mice were obtained by feeding a selenium‐deficient diet for three successive generations to Swiss‐Webster mice. Examination of epididymal sperm by transmission electron microscopy revealed progressively increasing alterations in the shape and arrangement of mitochondria within the midpiece. Other midpiece anomalies included acute bends, disorientation of the axoneme and dense fibers, and cytoplasmic masses at atypical locations. Some cross sections showed both the principal piece and midpiece within the same plasma membrane. Negatively stained whole mounts of mitochondrial ghosts prepared from epididymal sperm of normal and first‐generation selenium‐deficient mice and rats indicated that the selenium‐deficient ghosts were smaller, less curved, and more fragile than those of normal sperm mitochondria. Thus, selenium appears to be required for the normal development or stabilization of mitochondria1 shape during spermiogenesis in these rodents.
Binding to spermatozoa of a major soluble protein secreted by the epididymis of the lizard Lacerta vivipara
Authors: Depeiges Annie , Dufaure Jean‐Pierre
Published: February 2005

A major soluble protein occurring in two forms, a monomer (protein L) and a polymer, has been identified in the voluminous secretory granules produced by the epithelium of Lacerta vivipara epididymis. An antiserum was raised against protein L and used as an immunohistochemical probe. Indirect immunofluorescence microscopy has indicated that protein L is able to bind to the heads of the spermatozoa. By incubating spermatozoa with buffers of increasing ionic strength and by using nonionic detergents it was not possible to remove completely the protein L. Therefore it was concluded that the binding of protein L to the spermatozoa was not labile and might play an important role in the physiology of the spermatozoa, which is presently under investigation with this convenient model.
Structural changes in bovine oocytes during final maturation in vivo
Authors: Kruip T. A. M. , Cran D. G. , van Beneden T. H. , Dieleman S. J.
Published: February 2005

On the basis of structural observations bovine oocytes were grouped into four successive classed: 0, those before the luteinizing hormone (LH) surge; 1, those up to 8 h following the LH peak level; 2, those between 8 and 19 h after the LH peak level; and 3, those between 19 h after the LH peak level and ovulation.

Oocytes in class 0 had mitochondria located in a generally peripheral position. Interior to the mitochondria were elements of rough endoplasmic reticulum (RER) and numerous membrane‐bound vesicles which bore ribosome‐like particles on their outer surface. The first visible changesater the LH peak level as seen in class 1 were the formation of the periviteline space with loss of contact between the cumulus cells and the oocyte, and ruffing of the nuclear envelope. These changes were followed b the resumption of meiosis as defined by germinal‐vesicle breakdown (GVBD), the disappearance of RER, and the formation fo clusters of mitochondria in association with lipid droplets and elementrs of smooth endolasmic reticulum (SER).

The period between 8 and 19 h following LH peak level (class 2) was characterized by intensive clustering of mitochoncria in association with lipid droplets and elements of SER, conversion of lipid, fusion of vesicles, and the appearance of ribosomes in the cytoplasm. During the final stage (class 3), the polar body was extruded, the mitochondria dispersed, and the majority of the organelles became located toward the center of the cell. The relatively organelle‐free cortical region contained cortical granules immediately adjacent to the plasma membrane together with aggregates of tubular SER. The structural changes are discussed in the context of follicular steroidogenesis and oocyte developmental competence.
Cytoplasmic and sperm nuclear transformations in fertilized ammonia‐activated sea urchin (Arbacia punctulata) eggs
Authors: Longo Frank J.
Published: February 2005

Studies examining cytoplasmic and sperm nuclear transformations in sea urchin (Arbacia punctulata) eggs inseminated at different periods after ammonia activation have been caried out at the light‐ and electron‐microscopic levels of observation. Arbaca eggs treated with ammonia‐seawater demonstrated chromosome condensation after DNA synthesis and underwent a chromosome cycle similar to that described for Lytechinus [Mazia, 1947]. Cortical granule reaction, fertilization cone formation, and sperm aster development in eggs fertilized at 20 (interphase), 50 (prometaphase), and 180 (interphase) min after ammonia activation were structurally simialr to processes in untreated zygotes. Cyclical changes in the formation of fertilization cones and sperm asters, as reported for eggs fertilized after activation by agents that induce a cortical granule reaction, were not observed. Although sperm nuclear transformations were prolonged (14 vs 18 min), male pronuclei that developed in eggs fertilized 20 min after ammonia activation were morphologically similar to those observed in fertilized, untreated ova and incorporated 3H‐thymidine. Sperm incorporated into eggs at 50 min after ammonia activation underwent nuclear envelope breakdown and chromatin despersion; however, 3H‐thymidine incorporation was not observed, and male pronuclei rarely developed (less than 5% of all specimens examined). Subsequent to dispersion, the paternal chromatin condensed into chromosomes which were associated with an aster. These results demonstrate that although ammonia‐activated eggs inseminated at interphase or prometaphase undergo similar cytoplasmic alterations, sperm nuclear transformations vary with the chromosome cycle of the egg.
cAMP‐dependent phosphorylation associated with activation of motility of Ciona sperm flagella
Authors: Opresko L. K. , Brokaw C. J.
Published: February 2005

Demembranated spermatozoa of Ciona do not become motile when provided with MgATP, unless their motility is activated in vivo before demembranation or unless the demembranated spermatozoa are activated in vitro with cAMP or with the catalytic subunit of a cAMP‐dependent protein kinase. CAMP causes a greater than fivefold enhancement of 32P incorporation by demembranated spermatozoa. Analysis by one‐dimensional PAGE and autoradiography shows several axonemal protein bands that become 32P‐labeled during in vitro activation with cAMP and identifies protein bands whose labeling is specifically reduced if motility of the spermatozoa is activated before demembranation, suggesting that these proteins also become phosphorylated during activation of motolity in vivo. These phosphorylated proteins appear to include dynein heavy‐chain components, but axonemal tubulin is not phosphorylated. Partially phosphorylated spermatozoa can be activated by an increase in KCI concentration, which appears to dissociate one phosphorylated component from the axoneme.
The evolution of the nematode spermatozoon
Authors: Baccetti B. , Dallai R. , De Zio S. Grimaldi , Marinari A.
Published: February 2005

The main features of the Nematode sperm cell are the absence of a flagellum and of an acrosome. Transition forms have never been described, as in other animal phyla also reaching the aflagellate condition, like Platyhelminths and Arthropods. The absence of the flagellum must be considered as a definitive acquisition in the group. In addition, centrioles have been demonstrated to be lacking in most cases. The absence of the acrosome is the second general feature of the Nematode sperm cell. Among other features, more or less common to the Nematodes, the most important and general is the presence in the cell periphery of spheroidal membranous vesicles, originated from the Golgi complex but not involved in fertilization or in the production of ascaridin granules. These are absent only in the Ascarid Aspiculuris and the Dorylaimiid Xiphinema, both kinds of sperm having a peculiar shape. These granules are possibly involved in cell motility. Some Nematode sperm have proteinaceous crystalline inclusions originated from the rough endoplasmic reticulum, called ascaridin granules, the role of which remains obscure. A third important feature is the absence of a nuclear envelope, characterizing all described Nematode spermatozoa, the only exception being the Enoplid Mesacanthion, which seems to be for this reason the most primitive model in the group. Other features are the reduced number, or total absence of, the chondriome, an amoeboid movement not owing to an actomyosin system and a dense halo of 10‐nm filaments surrounding the perinuclear cytoplasm.

In this apparently homogeneous picture, three main evolutionary steps can be recognized. The first one, represented by the primitive Enoploid Mesacanthion, is that of a sperm conserving the nuclear envelope, surrounded by a few mitochondria and many membranous vesicles. The second, the most typical of the group, present in high Enoplida, and in Rhabditida, Strongylida, Ascarida, Spimrida, Trichinellida, is that of roundish, amoeboid spermatozoa devoid of a nuclear envelope but containing mitochondria, membranous vesicles, filaments, microtubules, sometimes centrioles, and sometimes ascaridin granules. The third step is apparently a simplification of the second; in fact, in Tylenchida and Dorylaimiida, the sperm is devoid of membranous vesicles, while in Mononchida and Dioctophymatida it is devoid of mitochondria. Aspiculuris, also devoid of membranous vesicles and having a big mitochondrial derivative, can be assigned to the same level. Nematode sperm evolution does not seem therefore to be a progressive acquisition of new characters, but rather a radiation from an already perfect model of some further simplifications occurring in parallel in most of the orders.
Fine structure of the spermatozoa of Chiton marginatus (mollusca: Amphineura), with special reference to nucleus maturation
Authors: Russell‐Pinto F. , Azevedo C. , Barandela T.
Published: February 2005

The spermatozoon of Chiton marginatus is a long uniflagellate cell displaying structural features of “modified sperm.” The nucleus presents a conical shape with a long apical cylindrical extension. The chromatin is homogeneously dense. Scattered inside the condensed nucleus, a few nuclear lacunae are visible. The acrosomal complex is lacking. Some mitochondria are located in a laterofrontal structure side by side with the nucleus. The typical midpiece is absent. The cytoplasm forms a thin layer around the nucleus and the mitochondria. The proximal centriole is in a basal nuclear indent. The distal centriole serves to form the axoneme tail with the usual microtubular pattern.

During nuclear maturation, the early spermatid nucleus is spherical and contains fine granular chromatin patches. The nuclear envelope shows a deposit of dense material at the base of the nucleus, forming a semicircular invagination occupied by a flocculent mass. In middle spermatid stage, the chromatin gets organized in filaments, coiled as a hank, attached over the inner surface of the basal thickening of the nuclear envelope. The nucleus starts to elongate anteroposteriorly. At the pointed apical portion of the spermatid, a group of microtubules is observed seeming to impose external pressure to the nucleus giving rise to the long apical nuclear point. The mitochondria have a basal position. Late spermatids have an elongated conical nucleus. The chromatin filaments are further condensed, and lacunae appear inside the nucleus. Some mitochondria migrate to a lateral position.
Effects of surfactants on the fertilizing capacity and acrosome reaction of sea urchin spermatozoa
Authors: Nakano Eizo , Hino Akiya , Furuse Kazumaro
Published: February 2005

The effects of seven surfactants on spermatozoa of the sea urchin, Hemicentrotus pulcherrimus, were studied. All these surfactants induced the acrosome reaction and inhibited the fertilizing capacity of spermatozoa. There was a statistically significant correlation between the concentrations that induce the acrosome reaction and inhibit fertilization. The critical micelle concentrations (CMC) of surfactants in sea water were almost even and these values, which are inherent physical properties of surfactants, did not provide a direct measure of their inhibitory effect of fertilization. Among seven surfactants, p‐menthanyl‐phenol polyoxyethylene (8.8) ether (TS‐88) with a characteristic hydrophobes was the most potent both in the induction of acrosome reaction and in the inhibition of fertilization. Various ethylene oxide adducts to p‐menthanyl‐phenol were also tested for the purpose of comparison. It is suggested that the effects of surfactants on sea urchin spermatozoa at low concentrations reflect their activity associated with the hydrophobic group inherent in each surfactant.
The attachment of acrosome‐intact sperm to the surface of zona‐free hamster oocytes
Authors: Koehler James K. , Smith Dianne , Karp Laurence E.
Published: February 2005

The attachment of acrosome‐intact mammalian sperm to zona‐free hamster vitelli has been investigated using light and electron microscopy. Uncapacitated sperm from hamsters, stallions, bulls, rabbits, and humans can attach firmly to the oocyte surface by means of microvillus processes. The general appearance of such attachments mimics the events seen during the interaction of acrosome‐reacted sperm with the oolemma. Very close, junction‐like connections develop between apical portions of sperm and oolemma after initial attachment, but no evidence of sperm penetration was observed. Significant species differences were noted with respect to the avidity of the sperm‐egg interaction. Bull, horse, and rabbit sperm showed a high degree of attraction for vitelli; hamster and human sperm interácted at a lower level. Thus, in the species tested, an acrosome reaction is not a prerequisite for sperm attachment to the zona‐free oolemma.
The antigenicity of the rabbit sperm acrosomes and the reversible sterility induced in the immunized rabbits
Authors: Srivastava P. N. , Tandon H. O. , Kumar V.
Published: February 2005

Rabbits were immunized against their alloantigens extracted from the sperm acrosome. The sequential method of extraction, first by MgCl2 then by detergents, provided the plasma and outer acrosomal membranes and their contents in the first step and the inner acrosomal membrane and its components in the next step. Both groups of antigens appear to have produced monospecific antisera. The MgCl2 extract antiserum lost its sperm immobilization and agglutination activities on absorption to its antigen. The detergent extract, under similar treatment, remained unaffected. Therefore, the agglutination and immobilization factors resided on the plasma membrane. The decrease in sperm concentration suggested that the immunogen affected the germinal epithelium. The reduction in the semen volume and seminal enzymes seems to indicate that the accessory sex glands were also affected by the treatment.

The antiserum blocked the dissolution of the hamster and rabbit zona pellucida by the MgCl2 extract. The normal serum had no such effect. Both the normal and immune sera inhibited acrosin equally well while the antisera inhibited other proteinases, arylsulfatase, and β‐N‐acetylhexosaminidase to a significantly greater extent. Immunized rabbits failed to impregnate females as long as the antibody titer remained high. As the titer declined these animals became fertile and the females delivered normal litters.
An ultrastructural study of the establishement of the testicular cysts during spermatogenesis in the sea anemone Actinia fragacea (Cnidaria: Anthozoa)
Authors: Larkman Alan U.
Published: February 2005

Spermatogenesis in the sea anemone Actinia fragacea takes place in numerous testicular cysts located in the mesoglea of the gonads. Prospermatogonia arise among the bases of the gonadal epithelial cells bordering the mesoglea, and later migrate into the mesoglea to establish the cysts. The prospermatogonia arise singly, but soon most are found as small groups within the endoderm. They are small cells, 6–7 μm in diameter, and have relatively large nuclei with a single nucleolus. Their cytoplasm is dense, and contains dense bodies and nuage material as well as Golgi, mitochondria, and individual cisternae of endoplasmic reticulum. Each prospermatogonium bears a flagellum, originating in a groove or channel in the cytoplasm.

A small proportion of prospermatogonia enter the mesoglea singly, but most migrate as elongate groups or “slugs” of cells. As they enter, the groups often become constricted into hour‐glass shapes, and they become covered by the endodermal basal lamina. During the later stages of entry, the last part of the group to enter retains contact with the bases of the epithelial cells, which are dragged into the mesoglea behind the germ cells. This contact between germ cells and endoderm persists throughout spermatogenesis and prevents closure of the mesoglea behind the group. The endodermal cells involved begin specialization to form the trophonema. Once entry is complete, the groups enlarge rapidly to form the testicular cysts. A small number of germ cells appear to remain behind in the endoderm after most have entered the mesoglea, and the possible significance of these cells is discussed.
Normal levels of zinc and sulfhydryls in morphologically abnormal populations of spermatozoa from moderately zinc‐deficient rats
Authors: Wallace Edith , Calvin Harold I. , Salgo Miklos P. , Dennis James E. , Ploetz Karin
Published: February 2005

Zinc is required for spermatogenesis in mammals and is concentrated in the dense outer fibers of the sperm tail, where it is associated with cysteine‐rich protein. To investigate the effects of marginal zinc deficiency upon dense fiber formation and upon sperm quality in general, weanling Sprague‐Dawley rats were administered a commercial low‐zinc diet, supplemented with phytate, for approximately 60 days, and were compared with controls fed the same diet plus 50 ppm zinc in their drinking water. The following characteristics of the zinc‐deficient rats were significantly lower than in the controls: body weight, testis weight, epididymis weight, seminal vesicle weight, sperm content of the cauda epididy‐midis, sperm motility, testis zinc, and hair zinc. By contrast, the levels of sperm zinc and sperm sulfhydryls were the same in the zinc‐deficient and control rats. The zinc‐deficient rats displayed a highly variable spectrum of sperm defects, which included decapitation, disorganized and redundant tail elements, and superfluous cytoplasm. However, abortive dense fiber development was only rarely observed. Apparently, even when availability of zinc is limited and reduced sperm production ensues, elaboration of dense fibers rich in zinc and sulfhydryls continues to be obligatory for the completion of spermiogenesis.
Structural components of sea urchin sperm nuclei
Authors: Kunkle Mel
Published: February 2005

The sea urchin sperm nucleus rapidly loses its conoid morphology and becomes more voluminous and spherical upon its entry into the egg cytoplasm during fertilization. This investigation has attempted to determine what are the structural constraints placed upon the sperm nucleus, so that further investigations might determine the egg cytoplasmic factors that are responsible for modifying nuclear morphology. Isolated sperm nuclei were subjected to various extraction procedures in order to remove the majority of the proteins (histones) and also the DNA; subsequently, the residual structures were processed for and examined by electron microscopy. The data presented in this investigation demonstrate the removal of the sperm nuclear histones plus other nonhistone proteins has no effect on the conoid morphology of the sperm nucleus, yet this protein removal has a profound effect on the structure of the nuclear chromatin. It is also shown that removal of the majority of the nuclear DNA has no effect on the shape of the sperm nucleus. These results indicate that there are other components (possibly a nuclear matrix) associated with the sperm nucleus that are responsible for maintaining its conoid morphology.
Sperm head structure in the hydromyinae (rodentia:Muridae): A further evolutionary development of the subacrosomal space in mammals
Authors: Breed W. G.
Published: February 2005

This scanning and transmission electron microscopical study has been performed to determine the extent of the development of the subacrosomal space in the sperm head of various members of the subfamily, the Hydromyinae, which is a diverse group of murid rodents occurring in various habitats in Australia. Sperm of all species from the three tribes of Hydromyini, Uromyini, and Conilurini investigated in this study had a head with three hooks although their absolute and relative lengths varied markedly. The top hook was similar in structure to that present in many other murid rodents and had a typical pseudoperforatorium present. The two ventral hooks had nuclear material basally, apical to which a large extension of the subacrosomal space occurred. Studies of testicular sections showed that the ventral hooks were formed late in spermiogenesis largely after condensation of the nucleus during which time electron dense material accumulated within them. No structure homologous to these hooks has been found in sperm from any other mammalian group, but, as the three tribes of Hydromyinae probably diverged about 20 million years ago, the extension of the subacrosomal space presumably evolved prior to this date.
A separate catalase and peroxidase in sea urchin sperm
Authors: Boldt Jeffrey , Alliegro Mark C. , Schuel Herbert
Published: February 2005

The release of hydrogen peroxide (H2O2) by the fertilized sea urchin egg has been shown to assist in the prevention of polyspermy [Coburn et al, 1981; Boldt et al, 1981]. Physiological data suggested that egg‐derived H2O2 reacts with a phenylhydrazine‐sensitive sperm peroxidase to inactivate sperm, while a 3‐amino‐1,2,4‐triazole‐sensitive catalase acts to protect sperm from H2O2 [Boldt et al, 1981]. Strongylocentrotus purpuratus sperm contain heat and pronase labile catalase and peroxidase activities. Differential extraction of sperm (hypotonic phosphate buffer for catalase and Triton X‐100 at high ionic strength for peroxidase) results in complete separation of these enzyme activities. The catalase is highly sensitive to inhibition by azide and 3‐amino‐1,2,4‐triazole, and less sensitive to inhibition by phenylhydrazine. The peroxidase is highly sensitive to inhibition by phenylhydrazine and relatively insensitive to 3‐amino‐1,2,4‐triazole and azide. These results show that two distinct H2O2 reactive enzymes, catalase and peroxidase, are present in sea urchin sperm, and are consistent with our hypothesis concerning the biological functions of these enzymes in fertilization.
Cyclic nucleotide metabolism in mouse epididymal spermatozoa during capacitation in vitro
Authors: Stein D. M. , Fraser L. R.
Published: February 2005

The role of cyclic nucleotides in sperm capacitation is equivocal. Using conditions known to support mouse sperm capacitation after 120 min incubation in vitro, the cAMP and cGMP contents of epididymal spermatozoa were measured and the cGMP/cAMP ratio determined. The initial high cAMP content detected upon release of spermatozoa decreased within 30 min to a lower plateau, which was then maintained throughout incubation. With the cGMP content remaining approximately constant, the cGMP/cAMP ratio increased over 120 min. In the presence of 2 mM caffeine, an increased cAMP content was noted at 0 and 30 min before a fall to the plateau level. To investigate cyclic nucleotide metabolism, adenylate cyclase and phosphodiesterase activities were compared in two sperm populations, one essentially uncapacitated and the other incubated for 120 min. Adenylate cyclase activity, higher in the presence of 2 mM Mn2+ compared to Mg2+, showed increased activity at 120 min compared to 30 min incubation, while phosphodiesterase activity decreased during this period. The ability of spermatozoa to form adenosine and inosine from cAMP indicated endogenous 5′‐nucleotidase and deaminase, as well as phosphodiesterase, activities. Although the endogenous cAMP content appeared to remain constant during the time that acrosome loss, hyperactivated motility and fertilizing ability can be demonstrated, activities of the enzymes responsible for cAMP metabolism indicate an increased potential for cAMP availability and turnover. The increased cGMP/cAMP ratio may also play a role during capacitation.
Morphological and ultrastructural characterization of mammalian spermatozoa processed for flow cytometric DNA analyses
Authors: Garner D. L. , Johnson L. A. , Lake S. , Chaney N. , Stephenson D. , Pinkel D. , Gledhill B. L.
Published: February 2005

The morphological and ultrastructural changes that occur during preparation of porcine, bovine, and murine spermatozoa for flow cytometric quantification of the relative DNA content of the X‐ and Y‐chromosome‐bearing sperm populations were examined. Ejaculated spermatozoa from the boar and bull were washed using a series of dimethyl sulfoxide (DMSO) solutions prior to fixation, whereas the epididymal mouse spermatozoa were washed only in phosphate‐buffered saline (PBS). Spermatozoa from all three species were then fixed in ethanol and processed for fluorochrome staining by a treatment regimen consisting of sulfhydryl reduction and proteolysis. The processed sperm nuclei were stained for DNA with the fluorochrome, 4′‐6‐diamidino‐2‐phenylindole (DAPI) before quantification by flow cytometry. Scanning and transmission electron micrographs of sperm heads taken at various steps of the preparation and staining procedures show 1) that the rigorous washing procedure disrupted the plasma and outer acrosomal membranes, 2) that ethanol fixation resulted in removal of the outer membranes and disintegration of the nuclear envelope, and 3) that thiol and proteolysis treatment removed the remaining cellular organelles including the tail and rapidly induced partial decondensation of the tightly packed chromatin. Sequential micrographs showed that the nuclear matrix of all three species increased in thickness about twofold during the preparation and staining. Consequently, the harsh procedures currently used for quantitative staining of DNA for high‐resolution flow cytometric analyses destroy most cellular organelles and thereby prevent simultaneous characterization of DNA content and other sperm cell constituents.
Fate of argyrophilic proteins during sea‐urchin (Hemicentrotus pulcherrimus) oogenesis detected by NOR‐silver staining
Authors: Amikura Reiko M. , Ihnuma Morio
Published: February 2005

NOR‐silver staining technique was applied to paraffin sections of sea urchin ovaries. The fibrillar component and the granular component of the nucleolus were stained dark brown and yellowish tan, respectively. Silver‐stained particles were also observed in germinal vesicles (GV) during oocyte growth. After the breakdown of GV, such particles appeared both in female pronucleus and egg cytoplasm, suggesting dispersion of the stained materials from the GV to the egg cytoplasm in the course of the breakdown of GV. The silver‐stained sites disappeared by the pretreatment with NaOH, trypsin, and pepsin but not with HCl, and RNase, indicating the protein derivation of silver‐stained sites. These results indicate the stained materials are kinds of protein.
Postinsemination changes in the amoeboid sperm of a nematode, nippostrongylus brasiliensis
Authors: Wright E. J. , Sommerville R. I.
Published: February 2005

Nematode sperm undergo a capacitationlike change called activation, after insemination into the female reproductive tract. In this paper we examine the morphological and functional consequences of activation of the sperm of Nippostrongylus brasiliensis by comparing the ultrastructure of sperm in the reproductive tracts of males and females and by analysing locomotion using time‐lapse cinematography. Sperm in males consist of a long tail, composed of highly condensed chromatin, and a cytoplasmic region containing numbrane‐bound secretory products of the Golgi called membranous organelles (MOs) as well as mitochondria. Following activation, the MOs fuse with the plasma membrane, release their electron‐dense contents, and remain connected to the outside by a narrow channel.

A small region at the anterior of sperm from males is amoeboid and can produce small pseudopodia. In sperm from the uterus this region enlarges considerably, the groundplasm changes from a granular to a filamentous from and the sperm become fully motile. During locomotion a cycle of events occurs at the leading edge beginning with the extension of a small pseudopodium. A constriction ring forms across its base and as the sperm progresses the constriction ring moves back along the cell, but remains stationary relative to the substrate. The location of new pseudopodia at the leading edge dictates the direction of movement of the spermatozoon. Since Cytochalasin B had no effect on the translocation of sperm, the polymerization of actin appears not to play a part in locomotion.
Binding of fluorescent lectins to the surface of germ cells from fetal and early postnatal mouse gonads
Authors: Felici Massimo De
Published: February 2005

Fluorescent lectins were used to study the chemical nature of carbohydrate moieties present on the surface of female and male germ cells isolated from mouse gonads during fetal and early posnatal development. Concanavalin A (ConA), lens culinaris agglutinin (LCA), ricinus communis agglutinin (RCAI) and wheat germ agglutinin (WGA) bound intensely to the germ cell plasma membrane at all stages studied. Other lectins such as ulex europaeus agglutinin (UEAI) and agglutinin (SBA) did not bind or bound moderately (SBA to female germ cells only).

Distinct developmental‐related changes were observed when female germ cells were labeled with fluorescein‐conjugated peanut agglutinin (PNA) or dolichos biflorus agglutinin (DBA). DBA and PNA binding was absent or weak in fetal female and male germ cells, but became intensely positive in oocytes in the immediate postnatal period. The percentage of oocytes stained with DBA increased during the first three days after birth, and from day 3–4 onwards all oocytes were strongly labeled.

I suggest that these changes in lectin binding reflect changes in biochemical structure of the oocyte surface related to differentiative events occurring in the mouse ovary immediately after birth.