Spermatology of Some Pycnogonida (Arthropoda), with Special Reference to a Microtubule-Nuclear Envelope Complex

The sperm cells of Nymphon leptocheles and N. rubrum are of the primitive type, which is a remarkable condition among arthropods. The motile sperm consist of a somewhat elongated head, a kind of midpiece and a long tail. An acrosome is absent. The nucleus is surrounded by longitudinally oriented microtubules running in furrows in the nuclear envelope. These microtubules are not interconnected by links or connected to the nuclear envelope; they persist in the mature sperm. No appreciable chromatin condensation takes place. The midpiece contains some unmodified mitochondria and a centriole. The tail is a simple, free flagellum. The results are in particular discussed in relation to other known microtubule-nuclear envelope complexes in sperm cells. The sperm cells of Pycnogonum littorale are, on the other hand, highly aberrant. They are unmotile, elongated cells containing a very high number (often more than 1000) of longitudinal microtubules arranged in complex patterns. Some folded membranes may represent the nuclear envelope. Other organelles are unidentificable or may be absent.     

Fine Structure of Spermatozoa in Perkinsiana rubra and Pseudopotamilla reniformis (Sabellidae: Polychaeta)

The Perkinsiana acrosome is elongated, pointed and tapering, strengthened anteriorly by cortical rings of longitudinally running units. The Pseudopotamilla acrosome is cap-like and its apical region contains a laminar body with two electron densities, the laminae crossed by occasional lines of dislocation, as in a crystal. These apparently solid regions are closely followed by peripherally situated lucent zones. The zone in Pseudopotamilla contains tubules, which are very like those in Sabella sperm, except that they lack connections with the acrosome base. Both species are probably broadcast spawners and have nuclei, mitochondria, centrioles and flagella of the primitive type. This may, however, be a secondary reversion through neoteny.     

Fine structure of the spermatozoa of Crassostrea gigas (Mollusca,Bivalvia)

We describe sperm ultrastructure and acrosome differentiation during spermiogenesis in Crassostrea gigas (Mollusca Bivalvia). The sperm cell is a uniflagellated cell of the primitive type. The head region contains a rounded or conical nucleus surmounted by small acrosome. This organelle consists of a membrane-bound acrosomal granule, the contents of which have a homogeneous density, except in the anterior region, which is positive for PTA. The acrosome also surrounds the perforatorium, which includes oriented fibrillar elements: this is the axial body. The middle piece contains four mitochondria encircling two perpendicular centrioles. The distal centriole is provided with a system of mechanical fixation to the plasma membrane, consisting of nine fibers in radial arrangement. The tail flagellum, about 50 m?m long, contains the usual microtubular axoneme. © 1993 Wiley-Liss, Inc.     

COMPARATIVE SPERMATOLOGY OF THREE SPECIES OF DONAX (BIVALVIA) FROM SOUTH AFRICA

The fine structure of the sperm and spermatogenesis in threespecies of Donax (D. madagascariensis, D. sordidus and D. serra)are described. Although the morphology of the sperm of all speciesis very similar, each has unique features. Donax madagascariensisand D. sordidus reportedly hybridize in regions of sympatryand their spermatozoa are morphologically closer to one anotherthan to D. serra. All sperm are of the primitive type with ahead(about 2 µmu; long), mid-piece of four mitochondria andtail. The head comprises a barrel-shaped nucleus which is cappedby a small, complex acrosome. The structure of the acrosomeis typical of heterodont bivalves. During spermatogenesis thepattern of nuclear chromatin condensation is granular. Glycogenfirst appears in the cytoplasm of spermatids, and in the maturesperm is sited in the mid-piece and base of the acrosome. (Received 15 May 1989; accepted 25 June 1989)     

The Spermatozoon of Siboglinum (Pogonophora)

The spermatozoon and some spermatid stages of Siboglinum (Pogonophora) have been examined by light and electron microscopy. In the spermatozoon a helical acrosome, a helical nucleus and a “body” with axonema follow each other in normal sequence. Head and tail are joined by a very short neck region containing two modified centrioles. The posterior portion of the nucleus is surrounded by a mitochondrial sheath consisting of three tightly wound mitochondrial helices. In the main portion of the tail the 9+2 unit is sorrounded by a granular sheath of dense material. In the neck region a centriole adjunct develops into a dense substance containing about nine rods. At an early stage, when the centriolar apparatus and flagellum become associated with the nucleus, three large mitochondria with fairly regular cristae are seen at the base of the nucleus. A well developed Golgi apparatus is present in early stages. Rows of microtubules are observed encircling the spermatid nucleus. Compared with the primitive type of spermatozoon the pogonophore sperm shows elongated and specialized nucleus, acrosome and mitochondria. It is concluded that the ancestral form must have had a fairly primitive spermatozoon and that evolution has proceeded towards a modified sperm with complicated spiral structure in connection with the evolution of a modified biology of fertilization, viz. specialized spermatophores. It is not known how the spermatophore discharges the spermatozoa nor how the spermatozoa find their way to the eggs. Two kinds of sperms are produced in the gonads of Siboglinum. The atypical sperm is smaller than the typical one.     

THE SPERMATOZOON OF ARTHROPODA: ZOOTERMOPSIS NEVADENSIS AND ISOPTERAN SPERM PHYLOGENY

In this paper, the ultrastructure of the spermatozoon of Zootermopsis nevadensis (Isoptera, Hodotermitidae) and of some Rhinotermitidae and Termitidae is described. Zootermopsis sperm is rod like, aflagellate, immotile, and without an acrosome; it is composed of a filiform nucleus encircled by a monolayered microtubular manchette, and a few mitochondria. This spermatozoon was previously thought to be flagellate, and therefore the most primitive in Isoptera: our present study suggests a new phylogenetical position for Hodotermitidae. All the species of Rhinotermitidae and Termitidae studied by us show a similar spheroidal sperm model, devoid of acrosome, flagellum and manchette at spermatid stage, and are made up of only a round nucleus, two mitochondria and a centriole. This widely distributed model seems to be the more evolved in the order. The nature of sperm evolution in the Isoptera is considered.     

Ultrastructural studies of spermatozoa in three bivalve species with notes on evolution of elongated sperm nucleus in primitive spermatozoa

Sperm ultrastructure and spermiogenesis of the three bivalve species Musculus discors, Nucula sulcata, and Dreissena polymorpha have been studied. During spermatid differentiation in Musculus discors and Nucula sulcata the nucleus attains an elongated rod-like shape. The spermatozoon from Nucula sulcata was found to have a cup-shaped acrosome and five mitochondria surrounding two centrioles in the middle piece. The spermatozoa from Musculus discors has a long complex acrosome. From the distal centriole striated processes extend and attach to the plasma membrane. The spermatozoon of the fresh water species Dreissena polymorpha agrees in all main features with those of other invertebrate groups with external fertilization. It is thus of the primitive type with barrel-shaped nucleus and four to five mitochondria1 spheres in the middle piece. The acrosome is a prominant, complex structure at the apex of the mature spermatozoon. A comparison of sperm ultrastructure among bivalves indicates that there is a certain correlation between the evolution of the elongated sperm nucleus and large, yolk-rich eggs. In species with an elongated sperm nucleus the increased egg size has often led to a lecithotrophic or direct development. The elongated nucleus is a slight modification of the primitive type. There is a great variation in acrosome structure among bivalve spermatozoa, reflecting diverging functional demands at fertilization of the eggs.     

Ultrastructure of the spermatozoon of Lepidogalaxias salamandroides and its phylogenetic significance

The spermatozoon of Lepidogalaxias salamandroides possesses an acrosome (putative), one or two perforatoria (putative) but no nine-triplet centrioles. Two elongated mitochondria (12 μm long) are situated in parallel between the nucleus (20 μm long) and the axoneme (53 μm long). The above features are unique among other teleosts with internal fertilization. The presence of an “acrosome” in this primitive teleost supports the hypothesis that this structure has been secondarily lost in teleosts during evolution. The uncertainty of phylogenetic placement of this fish is reflected by its unique sperm ultrastructure.     

Sperm ultrastructure in representatives of six bivalve families from Peter the Great Bay, Sea of Japan

The ultrastructure of sperm in seven species of bivalves, the representatives of six families, Arcidae (Anadara broughtonii, Arca boucardi), Anomiidae (Pododesmus macrochisma), Tellinidae (Macoma tokyoensis), Ostreidae (Crassostrea gigas), Myidae (Mya japonica) and Trapezidae (Trapezium liratum) is described. All the studied sperm were typical tail sperm, adapted to external insemination, which, however, had a specific structure. Differences were revealed in the form of head, acrosome structure and number of mitochondria. The studied species of the above families had their specific morphology, the Arcidae species had a bullet- or barrel-shaped head with four or five mitochondria in the middle part; the Anomiidae had conic head, the acrosome with periacrosome material and four mitochondria (a basic feature of sperm is the axial core entering periacrosome material and consisting of bundle of actin filaments); the Myidae had a curved conic head and four mitochondria; in the Tellinidae the head was bullet-shaped, the periacrosome material contained a fibril component and four mitochondria; the Trapezidae had sperm of a conic form with spherical acrosome. The spherical sperm of C. gigas were similar to sperm of Saccostrea commercialis and Crassostrea virginica, but with some distinctions in the acrosome substructure. The morphology of sperm testified to the correct attribution of the Crassostreidae family as a synonym to the Ostreidae family.     

Sperm ultrastructure in two species of the polychaete genusHarmothoe (Polynoidae)

The structure of spermatozoa is described for two species of polynoid polychaete,Harmothoe imbricata andHarmothoe impar, from material fixed and examined by both scanning and transmission electron microscopy. The two species undergo spermiogenesis within discrete testes. The testis ofH. imbricata is shown to have a layer of epithelial cells which possess an outer cuticular layer and a microvillous inner surface. Spermatocytes of both species are spherical but there are marked differences in the shape and size of the spermatozoa of the two species.H. impar has a classical primitive spermatozoon with a rounded head (2 μm long) and a button-shaped acrosome. Fully differentiated spermatozoa ofH. imbricata are modified from the primitive form by having a long head (10 μm length) with a pointed acrosome about 6 μm in length. Spermatozoa ofH. imbricata have a ring of up to fourteen mitochondria around a centrally inserted flagellum at the posterior whereasH. impar has a ring of four or five spherical mitochondria. Spermiogenesis is well synchronised inH. imbricata but all developmental stages can be found simultaneously in the testis ofH. impar. The differences in sperm structure of the two species may be related to differences in breeding biology which are hitherto unknown.     

Not all sperm are equal: functional mitochondria characterize a subpopulation of human sperm with better fertilization potential

Human sperm samples are very heterogeneous and include a low amount of truly functional gametes. Distinct strategies have been developed to characterize and isolate this specific subpopulation. In this study we have used fluorescence microscopy and fluorescence-activated cell sorting to determine if mitochondrial function, as assessed using mitochondrial-sensitive probes, could be employed as a criterion to obtain more functional sperm from a given ejaculate. We first determined that mitochondrial activity correlated with the quality of distinct human samples, from healthy donors to patients with decreased semen quality. Furthermore, using fluorescence-activated cell sorting to separate sperm with active and inactive mitochondria we found that this was also true within samples. Indeed, sperm with active mitochondria defined a more functional subpopulation, which contained more capacitated and acrosome intact cells, sperm with lower chromatin damage, and, crucially, sperm more able to decondense and participate in early development using both chemical induction and injection into mature bovine oocytes. Furthermore, cell sorting using mitochondrial activity produced a more functional sperm subpopulation than classic swim-up, both in terms of improvement in a variety of functional sperm parameters and in statistical significance. In conclusion, whatever the true biological role of sperm mitochondria in fertilization, mitochondrial activity is a clear hallmark of human sperm functionality.     

Fine structure of the spermatozoon of Marthasterias glacialis (Linnaeus) (Echinodermata; Asteroidea), with special reference to acrosomal morphology

The sperm of Marthasterias glacialis (Linnaeus) was studied by light and electron microscopy. It is a long uniflagellated cell of the “primitive” type. The head has a spherical shape and contains a nucleus with a spheroid acrosome lying in a cup-shaped anterior fossa. The acrosome is formed by an acrosomal vesicle surrounded by the periacrosomal material. The basal specializations of the acrosomal vesicle show a clear differentiation of its constituents resembling the structure of membrane. The midpiece contains a very large annular mitochondrion which encircles two perpendicular centrioles. The distal centriole is in close association with a pericentriolar radial complex. The tail, containing a common microtubular axoneme, is projected to a variable position.     

中国绿螂精子的超微结构

应用透射电镜技术观察了中国绿螂(Glaucomya chinensis)精子的超微结构。精子为典型的原生型,包括头部、中段和尾部三部分。头部由顶体和细胞核组成。顶体呈倒"V"字型。细胞核呈长圆柱形,没有核前窝,具有核后窝。中段由4个线粒体环绕中心粒而成。尾部细长,为典型的"9+2"结构。文中还讨论了双壳类精子形态结构的种属间差异。     

Ultrastructure of the sperm and spermatogenesis and spermiogenesis of Dina lineata (hirudinea,erpobdellidae)

The mature sperm of Dina lineata is of the modified type. The sperm are 48 μm long and 0.3 μm wide. The sperm are filiform and helicoidal cells with a distinct head, a midpiece, and a tail. There are two distinct regions in the head: the acrosome and the posterior acrosome, each with its own characteristic morphology. The midpiece is the mitochondrial region and has a single mitochondrion. Two distinct portions can be observed in the tail: the axonematic region and the terminal piece. In the process of spermatogenesis the early spermatogonia divide to form a poliplast of 512 spermatic cells. In the spermiogenesis the following sequential stages can be distinguished: elongation of the flagellum; reciprocal migration of mitochondria and Golgi complex; condensation of chromatin and formation of the posterior acrosome; spiralization of nuclear and mitochondrial regions; and, finally, formation of the anterior acrosome. The extreme morphological complexity of the Dina spermatozoon is related to the peculiar hypodermal fertilization which characterizes the erpobdellid family. Correlation between sperm morphology and fertilization biology in the Annelida is revised.     

Ultrastructure of Sperm in <Emphasis Type="Italic">Mercenaria stimpsoni</Emphasis> and <Emphasis Type="Italic">Mactra chinensis</Emphasis> (Mollusca: Bivalvia) from the Sea of Japan

The ultrastructure of the sperm of the common bivalve species Mercenaria stimpsoni and Mactra chinensis from Peter the Great Bay is described. The sperm structure is typical for animals with external insemination. The sperm consists of a head, middle part, and flagellum. The sperm head of M. stimpsoni has a curved crescent form and includes the nucleus and acrosome; the head length is 9.8 μm. The acrosome is subdivided to the acrosome granule and the periacrosomal material. There are 4 mitochondria of about 0.8 μm in size in the middle part of the spermatozoon. The mitochondria surround the centriolar apparatus, which consists of proximal and distal centrioles located at a right angle. The axoneme originates from the distal centriole. The sperm of M. chinensis is barrel-shaped, with a head length of 3.2 μm. The acrosome is relatively larger, and its height is 1–1.2 μm. There are also 4 mitochondria 0.6–0.8 μm in the middle part of the spermatozoon. The sperm structure of the described species is typical of the families to which the mollusks belong, with insignificant variations.     

Nuclear and cytoplasmic differentiation in developing sperm of the crayfish,Cambaroides japonicus

Summary In the present electron microscopic study of spermatogenesis in the crayfish, Cambaroides japonicus, it was possible to clarify several aspects of the unusual differentiation which leads to the production of an aflagellate sperm. The centriole is followed from the metaphase of the second spermatocyte division to the time at which, in the nearly mature sperm, it appears to disintegrate. It has no connection with the acrosome but in the late spermatid and maturing sperm it is found randomly oriented among the convoluted membranes of the filamentous endoplasmic reticulum.There appears to be a close association of mitochondria with the developing acrosomal vesicle. Typical mitochondria, however, are not present after the late spermatid stage of development. It is suggested that the complex lamellar bodies associated with the nuclear envelope in the late stages of spermatogenesis may be related to mitochondria for these lamellar bodies resemble the complex mitochondria found in the adjacent nutritive cells.The development of the acrosome has been traced from an aggregate of dense granules which first appear in the interzonal spindle region and are later segregated at one side of the cell after the second spermatocyte division. As differentiation proceeds, tubular elements appear and disappear within the acrosome, while somewhat later, fibrous elements appear in the matrix. In the mature acrosome, the fibrous elements remain only adjacent to the granular periphery of the acrosome and the core again becomes homogeneous.No typical Golgi complex is found in these cells at any time during their differentiation.In the maturing sperm the development of the arms of the nucleus was studied. Preceding the differentiation of the arms a coarse fibrous material develops in the periphery of the nucleus. It is shown that the fibrillar material in the matrix of the arms is in continuity with the fibrillar material in the matrix of the nucleus proper.Supported in part by Grant No. B 2314 of the National Institute of Neurological Diseases and Blindness, U.S. Public Health Service.Predoctoral Research Fellow of the National Institute of Neurological Diseases and Blindness, U.S. Public Health Service.     

Use of fluorescent probes to assess membrane integrity in mammalian spermatozoa.

Carboxyfluorescein diacetate and propidium iodide were used as fluorescent stains to assess membrane integrity in sperm populations from ram and boar. The living spermatozoa were immobilized with low concentrations of formaldehyde so that individual stained cells could be observed in a suspension with the aid of a fluorescence microscope. Intracellular esterases liberated impermeant-free carboxyfluorescein from the permeant carboxyfluorescein diacetate and caused the product to accumulate and fluoresce green within the acrosome and the mitochondria as well as within the cytoplasm. Most of the spermatozoa (the intact ones) accumulated carboxyfluorescein in all compartments; however, a few cells (those with damaged plasma membranes) accumulated the stain only in the acrosome and/or the mitochondria, while others (all of whose membranes were damaged) remained entirely unstained. The impermeant propidium iodide did not stain any of the (intact) spermatozoa that accumulated carboxyfluorescein throughout their length, but stained all the others (the heads fluoresced red). The technique appeared to provide more reliable estimations of the percentage of functional cells than did motility estimations or assessments of acrosomal integrity (presence of normal apical ridge). The technique also demonstrated the sensitivity of the sperm plasma membrane to cold shock: virtually all cells rapidly became permeable to the stains after such stress. Assessments of boar sperm samples during preparative incubation for in-vitro fertilization indicated a considerable increase in the percentage of cells with damaged plasma membranes as incubation proceeded, in advance of the increase in the percentage of cells with discharged acrosomes.     

The origin of nematode sperm: Progenesis at the cellular level

Analysis of the development and structure of aberrant sperm of nematodes and other metazoans with internal insemination showed that these spermatozoa have several unusual, but shared features: (1) the absence of a flagellum and an axoneme, an unusual arrangement of centrioles; (2) an amoeboid shape and amoeboid motility due to cytoskeleton components; (3) the poor condensation of nuclear chromatin, which may be diffuse, thread-like, and discrete; (4) the absence of a nuclear envelope; (5) multiple unmodified mitochondria; (6) the absence of an acrosome; (7) unique membranous components derived from the Golgi complex; and (8) the large size of spermatozoa due to prominent cytoplasm filled with a great number of components. These shared features of aberrant spermatozoa may be explained by the conservation of a number of features that are characteristic of the primitive undifferentiated cell (the predecessor of all specialized gametes). The primitive cell features of numerous versions of aberrant sperm reflect the arrest of cytoplasm specialization of male gametes at an early stage of development. This pattern of gamete evolution is quite consistent with the conception of progenesis (retention of juvenile characters by precocious, sexually mature morphologically juvenile stage). Thus, the origin of the aberrant sperm of nematodes and many other metazoans may be interpreted as progenesis at the cellular level.     

Cryopreservation of epididymal sperm in tree shrews (Tupaia belangeri)

Cryopreservation of sperm from tree shrews, which are considered primitive primates, would enhance genetic management and breeding programs. Epididymal sperm were surgically harvested from male tree shrews, cryopreserved in two Tes-Tris-based cryodiluents, and used in four experiments. In Experiment 1, there were no significant differences in motility and acrosome integrity among five concentrations of egg yolk in TTE after cooling to 4 °C. However, sperm frozen in TTE containing 20% egg yolk at −172 °C/min had better (P < 0.05) post-thaw motility and acrosome integrity. In Experiment 2, sperm held for 10 min prior to storage in liquid nitrogen had greater motility than those held for 5 or 15 min (P < 0.05), but acrosome integrity was not different (P > 0.05) among treatments. In Experiment 3, sperm frozen in TTE diluent had higher (P < 0.05) motility and acrosome integrity than those in TEST diluent. In Experiment 4, there were no differences (P > 0.05) in the fertilization rate of oocytes and the proportion of tree shrews yielding fertilized oocytes, following AI with fresh versus frozen sperm. In conclusion, tree shrew epididymal sperm were successfully cryopreserved, as assessed by post-thaw motility, acrosome integrity, and fertilizing ability.