Morphogenesis of the fibrous sheath in the marsupial spermatozoon

The spermatozoon fibrous sheath contains longitudinal columns and circumferential ribs. It surrounds the axoneme of the principal piece of the mammalian sperm tail, and may be important in sperm stability and motility. Here we describe its assembly during spermiogenesis in a marsupial, the brush-tail possum, and compare its structural organization with that of eutherian mammals, birds and reptiles. Transmission electron microscopy showed that possum fibrous sheath assembly is a multistep process extending in a distal-to-proximal direction along the axoneme from steps 4 to 14 of spermiogenesis. For the most part, assembly of the longitudinal columns occurs before that of the circumferential ribs. Immunohistochemical and immunogold labelling showed that fibrous sheath proteins are first present in the spermatid cytoplasm; at least some of the proteins of the sheath precursors differ from those in the mature fibrous sheath. That immunoreactivity develops after initiation of chromatin condensation suggests that fibrous sheath proteins, or their mRNAs, are stored within the spermatid cytoplasmic lobule prior to their assembly along the axoneme. These findings are similar to those in laboratory rats, and thus suggests that the mode of fibrous sheath assembly evolved in a common ancestor over 125 million years ago, prior to the divergence of marsupial and eutherian lineages.     

Spermiogenesis and spermiation in a marsupial, the tammar wallaby (Macropus eugenii)

Fourteen steps of spermatid development in the tammar wallaby (Macropus eugenii), from the newly formed spermatid to the release of the spermatozoon into the lumen of the seminiferous tubules, were recognised at the ultrastructural level using transmission and scanning electron microscopy. This study confirmed that although the main events are generally similar, the process of the differentiation of the spermatid in marsupials is notably different and relatively more complex than that in most studied eutherian mammals and birds. For example, the sperm head rotated twice in the late stage of spermiogenesis: the shape of the spermatid changed from a T-shape at step 10 into a streamlined shape in step 14, and then back to T-shape in the testicular spermatozoa. Some unique figures occurring during the spermiogenesis in other marsupial species, such as the presence of Sertoli cell spurs, the nuclear ring and the subacrosomal space, were also found in the tammar wallaby. However, an important new finding of this study was the development of the postacrosome complex (PAC), a special structure that was first evident as a line of electron dense material on the nuclear membrane of the step 7 spermatid. Subsequently it became a discontinuous line of electron particles, and migrated from the ventral side of the nucleus to the area just behind the posterior end of the acrosome, which was closely located to the sperm–egg fusion site proposed for Monodelphis domestica (Taggart et al. 1993). The PAC and its possible role in both American and Australian marsupials requires detailed examination. Distinct immature features were discovered in the wallaby testicular spermatozoa. A scoop shape of the acrosome was found on the testicular spermatozoa of the tammar wallaby, which was completely different to the compact button shape of acrosome in ejaculated spermatozoa. The fibre network found beneath the cytoplasm membrane of the midpiece of the ejaculated sperm also did not occur in the testicular spermatozoa, although the structure of the principal piece was fully formed and had no obvious morphological difference from that of the epididymal and ejaculated spermatozoa. The time frame of the formation of morphologically mature spermatozoa in the epididymis of the tammar wallaby needs to be determined by further studies.     

Evidence for an early appearance of modern post-switch immunoglobulin isotypes in mammalian evolution (II); cloning of IgE,IgG1 and IgG2 from a monotreme,the duck-billed platypus,Ornithorhynchus anatinus

To trace the emergence of the modern post-switch immunoglobulin (Ig) isotypes in vertebrate evolution we have studied Ig expression in mammals distantly related to eutherians. We here present an analysis of the Ig expression in an egg-laying mammal, a monotreme, the duck-billed platypus (Ornithorhynchus anatinus). Fragments of platypus IgG and IgE cDNA were obtained by a PCR-based screening using degenerate primers. The fragments obtained were used as probes to isolate full-length cDNA clones of three platypus post-switch isotypes, IgG1, IgG2, and IgE. Comparative amino acid sequence analysis against IgY, IgE and IgG from various animal species revealed that platypus IgE and IgG form branches that are clearly separated from those of their eutherian (placental) counterparts. However, the platypus IgE and IgG still conform to the general structure displayed by the respective Ig isotypes of eutherian and marsupial mammals. According to our findings, all of the major evolutionary changes in the expression array and basic Ig structure that have occurred since the evolutionary separation of mammals from the early reptile lineages, occurred prior to the separation of monotremes from marsupial and placental mammals. Hence, our results indicate that the modern post-switch isotypes appeared very early in the mammalian lineage, possibly already 310-330 million years ago.     

Those other mammals: The immunoglobulins and T cell receptors of marsupials and monotremes

This review summarizes analyses of marsupial and monotreme immunoglobulin and T cell receptor genetics and expression published over the past decade. Analyses of recently completed whole genome sequences from the opossum and the platypus have yielded insight into the evolution of the common antigen receptor systems, as well as discovery of novel receptors that appear to have been lost in eutherian mammals. These species are also useful for investigation of the development of the immune system in organisms notable for giving birth to highly altricial young, as well as the evolution of maternal immunity through comparison of oviparous and viviparous mammals.     

Development of membrane differentiations in the guinea pig spermatid during spermiogenesis

We have studied the development of membrane differentiations in guinea pig spermatids during spermiogenesis, using electron microscopy of thin sections, freeze-fracturing, and filipin labeling as an indicator of membrane cholesterol distribution. The development of the distinctive membrane specializations closely correlated with the developmental steps of spermatid differentiation. The annulus, the zipper, and the circumferential strands of particles overlying the mitochondrial gyres of the midpiece appeared in the plasma membrane over the tail during the last two steps of spermiogenesis. The outer acrosomal membrane showed trnsient crenations during the acrosome phase and serrations over the equatorial segment of the acrosome during the maturation phase. On the inner nuclear membrane, a depression appeared opposite the inner zone of the acrosome during the Golgi phase and persisted throughout spermiogenesis. The implantation fossa also appeared during this phase, as a bulging on the inner nuclear envelope covered with small membrane particles. By the cap phase, small and large 15–20-nm particles were present at the implantation-fossa site, and clusters of 9-nm particles were seen over the postacrosomal segment of the nuclear envelope. The migration of the nuclear pores began when close contacts were established between adjacent nuclear membranes. It started simultaneously at the anterior pole and the implantation fossa and later progressed over the segment of membrane between the caudal margin of the acrosome and the posterior ring. By the maturation phase, the nuclear pores had migrated to the redundant nuclear envelope. After filipin labeling, a gradient was visible in the distribution of the cholesterol-filipin complexes, decreasing from the plasma membrane to the nuclear membrane. Within the same membrane, cholesterol distribution varied from one pole of the cell to the other. On the differentiations of the plasma membrane over the tail, cholesterol-filipin complexes were absent, but they were profuse in the membrane over the head. In the acrosomal membrane, the complexes were gathered in clumps associated with the crenations. They became fewer in the rostral segment but remained moderate in the caudal equatorial segment as spermiogenesis proceeded. The nuclear membrane contained cholesterol-free regions opposite the inner acrosome and at the site of the implantation fossa. Otherwise, the acrosomal segment of the nuclear envelope demonstrated a homogeneous distribution of cholesterol-filipin complexes, while the postacrosomal segment showed small clumps of complexes adjacent to nuclear pores. We propose that each of the spermatid cell membranes is not an autonomous system, but is dependent upon its exchanges with the immediate external environment under the one side of the membrane and its close coupling to the internal environment (i.e., membrane-associated structures: annulus, zipper, etc.) under the otehr side of the membrane. We suggest that the cholesterol in spermatid cell membranes contributes to the establishment of their mosaicism and regulates the modeling of the spermatid by modulating the internal fluidity of individual membrane segments during spermiogenesis.     

Posttesticular development of spermatozoa of the tammar wallaby (Macropus eugenii)

Tammar wallaby spermatozoa undergo maturation during transit through the epididymis. This maturation differs from that seen in eutherian mammals because in addition to biochemical and functional maturation there are also major changes in morphology, in particular formation of the condensed acrosome and reorientation of the sperm head and tail. Of spermatozoa released from the testes, 83% had a large immature acrosome. By the time spermatozoa reached the proximal cauda epididymis 100% of sperm had condensed acrosomes. Similarly 86% of testicular spermatozoa had immature thumb tack or T shape head-tail orientation while only 2% retained this immature morphology in the corpus epididymis. This maturation is very similar to that reported for the common brush tail possum, Trichosurus vulpecula. However, morphological maturation occurred earlier in epididymal transit in the tammar wallaby. By the time spermatozoa had reached the proximal cauda epididymis no spermatozoa had an immature acrosome and thumbtack orientation. Associated with acrosomal maturation was an increase in acrosomal thiols and the formation of disulphides which presumably account for the unusual stability of the wallaby sperm acrosome. The development of motility and progressive motility of tammar wallaby spermatozoa is similar to that of other marsupials and eutherian mammals. Spermatozoa are immotile in the testes and the percentage of motile spermatozoa and the strength of their motility increases during epididymal transit. During passage through the caput and corpus epididymis, spermatozoa first became weakly motile in the proximal caput and then increasingly progressively motile through the corpus epididymis. Tammar wallaby spermatozoa collected from the proximal cauda epididymis had motility not different from ejaculated spermatozoa. Ultrastructural studies indicated that acrosomal condensation involved a complex infolding of the immature acrosome. At spermiation the acrosome of tammar wallaby spermatozoa was a relatively large flat or concave disc which projected laterally and anteriorly beyond the limits of the nucleus. During transit of the epididymal caput and proximal corpus the lateral projections folded inwards to form a cup like structure the sides of which eventually met and fused. The cavity produced by this fusion was lost as the acrosome condensed to its mature form as a small button-like structure contained within the depression on the anterior end of the nucleus. During this process the dorsal surface of the immature acrosome and its outer acrosomal membrane and overlying plasma membrane were engulfed into the acrosomal matrix. This means that the dorsal surface of the acrosomal region of the testicular tammar wallaby sperm head is a transient structure. The dorsal acrosomal surface of the mature spermatozoon appears ultrastructurally to be the relocated ventral surface of the acrosomal projections which previously extended out beyond the acrosomal depression on the dorsal surface of the nucleus of the immature spermatozoon.     

Evolution and Development of the Atrial Septum

The complete division of the atrial cavity by a septum, resulting in a left and right atrium, is found in many amphibians and all amniotes (reptiles, birds, and mammals). Surprisingly, it is only in eutherian, or placental, mammals that full atrial septation necessitates addition from a second septum. The high incidence of incomplete closure of the atrial septum in human, so-called probe patency, suggests this manner of closure is inefficient. We review the evolution and development of the atrial septum to understand the peculiar means of forming the atrial septum in eutherian mammals. The most primitive atrial septum is found in lungfishes and comprises a myocardial component with a mesenchymal cap on its leading edge, reminiscent to the primary atrial septum of embryonic mammals before closure of the primary foramen. In reptiles, birds, and mammals, the primary foramen is closed by the mesenchymal tissues of the atrioventricular cushions, the dorsal mesenchymal protrusion, and the mesenchymal cap. These tissues are also found in lungfishes. The closure of the primary foramen is preceded by the development of secondary perforations in the septal myocardium. In all amniotes, with the exception of eutherian mammals, the secondary perforations do not coalesce to a secondary foramen. Instead, the secondary perforations persist and are sealed by myocardial and endocardial growth after birth or hatching. We suggest that the error-prone secondary foramen allows large volumes of oxygen-rich blood to reach the cardiac left side, needed to sustain the growth of the extraordinary large offspring that characterizes eutherian mammals. Anat Rec, 302:32–48, 2019. © 2018 The Authors. The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.     

Ultrastructure of sperm and spermiogenesis ofPterastericola astropectinis (Platyhelminthes,Rhabdocoela, Pterastericolidae)

During development of the spermatid, two free flagella are transported distally from the main spermatid mass by elongation of the shaft. They rotate back towards the shaft and lie parallel with it prior to fusion in a distal-proximal direction. This process conforms to that found in other free-living platyhelminths that have fused or adjoined axonemes in their sperm (Acoela, Kalyptorhynchia, Polycladida). It is basically different from the process of fusion in the majority of neodermatan species (major groups of parasitic platyhelminths), where the attachment of axonemes remains near the main spermatid body, a median process grows out between them, the nucleus and mitochondria migrate into this process and axonemes then fuse with it in a proximal-distal manner.Pterastericola astropectinis also differs from the Neodermata in the presence of dense bodies in the sperm. The ultrastructure of the sperm, spermiogenesis and protonephridia does not support the view of a close affinity between Pterastericolidae and Neodermata.     

Cloning of the MHC class II DRB cDNA from the brushtail possum (Trichosurus vulpecula)

The cell-surface glycoproteins encoded by the major histocompatibility complex (MHC) bind to processed foreign antigens and present them to T lymphocytes. Two classes of MHC molecules and their corresponding gene sequences have been extensively studied in eutherian mammals and birds, but data on the marsupial MHC are limited. Marsupials split from eutherian mammals about 125 million years ago and represent a distinct branch in mammalian evolution. Here the cDNA cloning of MHC class II genes of the brushtail possums (Trichosurus vulpecula) is reported. The sequences obtained were found to be relatively conserved when compared to the red-necked wallaby (Macropus rufogriseus) and an South American marsupial, Monodelphis domestica. The T. vulpecula sequence shared an average overall sequence identity of 75.4% at the deduced amino acid level with M. rufogriseus and M. domestica, respectively.     

Localization of boar sperm proacrosin during spermatogenesis and during sperm maturation in the epididymis

The localization of proacrosin was determined by using colloidal gold labeling and electron microscopy of boar germ cells during spermiogenesis to post-ejaculation. Proacrosin was first localized in round spermatids during the Golgi phase of spermiogenesis; it was associated with the electron-dense granule, or acrosomal granule that was conspicuous within the acrosome. It remained within the acrosomal granule during the cap and acrosome phases of spermiogenesis. At these stages, there was no apparent association of the proacrosin molecule with the acrosomal membranes. During the maturation phase of spermiogenesis, proacrosin was seen to become dispersed into all regions of the acrosome except the equatorial segment. When sperm from different segments of the epididymis and ejaculated sperm were examined, localization was observed throughout the acrosome except for the equatorial segment. Here proacrosin appeared to be localized on both the inner and outer acrosomal membranes as well as with the acrosomal matrix, although further studies are required to verify the membrane localization. No labeling was seen on the plasma membrane. These data suggest that the synthesis and movement of proacrosin to sites in the acrosome are controlled by an as yet unknown process. The absence of proacrosin on the plasma membrane of mature ejaculated sperm makes it unlikely that this enzyme plays a role in sperm-zona adhesion prior to capacitation.     

Sleep in the platypus.

We have conducted the first study of sleep in the platypus Ornithorhynchus anatinus. Periods of quiet sleep, characterized by raised arousal thresholds, elevated electroencephalogram amplitude and motor and autonomic quiescence, occupied 6-8 h/day. The platypus also had rapid eye movement sleep as defined by atonia with rapid eye movements, twitching and the electrocardiogram pattern of rapid eye movement. However, this state occurred while the electroencephalogram was moderate or high in voltage, as in non-rapid eye movement sleep in adult and marsupial mammals. This suggests that the low-voltage electroencephalogram is a more recently evolved feature of mammalian rapid eye movement sleep. Rapid eye movement sleep occupied 5.8-8 h/day in the platypus, more than in any other animal. Our findings indicate that rapid eye movement sleep may have been present in large amounts in the first mammals and suggest that it may have evolved in pre-mammalian reptiles.     

Evidence for an early appearance of modern post-switch isotypes in mammalian evolution; cloning of IgE,IgG and IgA from the marsupial Monodelphis domestica

In birds, reptiles and amphibians the IgY isotype exhibits the functional characteristics of both of IgG and IgE. Hence, the gene for IgY most likely duplicated some time during early mammalian evolution and formed the ancestor of present day IgG and IgE. To address the question of when IgY duplicated and formed two functionally distinct isotypes, and to study when IgG and IgA lost their second constant domains, we have examined the Ig expression in a non-placental mammal, the marsupial Monodelphis domestica (grey short-tailed opossum). Screening of an opossum spleen cDNA library revealed the presence of all three isotypes in marsupials. cDNA clones encoding the entire constant regions of opossum IgE (ϵ chain), IgG (γ chain) and IgA (α chain) were isolated, and their nucleotide sequences were determined. A comparative analysis of the amino acid sequences for IgY, IgA, IgE and IgG from various animal species showed that opossum IgE, IgG and IgA on the phylogenetic tree form branches clearly separated from their eutherian counterparts. However, they still conform to the general structure found in eutherian IgE, IgG and IgA. Our findings indicate that all the major evolutionary changes in the Ig isotype repertoire, and in basic Ig structure that have occurred since the evolutionary separation of mammals from the early reptile lineages, occurred prior to the evolutionary separation of marsupials and placental mammals.     

The region homologous to the X-chromosome inactivation centre has been disrupted in marsupial and monotreme mammals

Summary Marsupial, as well as eutherian, mammals are subject to X chromosome inactivation in the somatic cells of females, although the phenotype and the molecular mechanism differ in important respects. Monotreme mammals appear to subscribe at least to a form of dosage compensation of X-borne genes. An important question is whether inactivation in these non-eutherian mammals involves co-ordination by a control locus homologous to the XIST gene and neighbouring genes, which play a key regulatory role in human and mouse X inactivation. We mapped BACs containing several orthologues of protein-coding genes that flank human and mouse XIST and genes that lie in the homologous region in chicken and frog. We found that these genes map to two distant locations on the opossum X, and also to different locations on a platypus autosome. We failed to find any trace of an XIST orthologue in any marsupial or monotreme or on any flanking BAC, confirming the conclusion from recent work that non-eutherian mammals lack XIST. We propose the region homologous to the human and mouse X-inactivation centre expanded in early mammals, and this unstable region was disrupted independently in marsupial and monotreme lineages. In the eutherian lineage, inserted and existing sequences provided the starting material for the non-translated RNAs of the X-inactivation centre, including XIST. Electronic supplementary material The online version of this article (doi: ) contains supplementary material, which is available to authorized users.     

Uterine remodelling during pregnancy and pseudopregnancy in the brushtail possum (Trichosurus vulpecula; Phalangeridae)

The formation of a placenta is critical for successful mammalian pregnancy and requires remodelling of the uterine epithelium. In eutherian mammals, remodelling involves specific morphological changes that often correlate with the mode of embryonic attachment. Given the differences between marsupial and eutherian placentae, formation of a marsupial placenta may involve patterns of uterine remodelling that are different from those in eutherians. Here we present a detailed morphological study of the uterus of the brushtail possum (Trichosurus vulpecula; Phalangeridae) throughout pregnancy, using both scanning and transmission electron microscopy, to identify whether uterine changes in marsupials correlate with mode of embryonic attachment as they do in eutherian mammals. The uterine remodelling of T. vulpecula is similar to that of eutherian mammals with the same mode of embryonic attachment (non‐invasive, epitheliochorial placentation). The morphological similarities include development of large apical projections, and a decrease in the diffusion distance for haemotrophes around the period of embryonic attachment. Importantly, remodelling of the uterus in T. vulpecula during pregnancy differs from that of a marsupial species with non‐invasive attachment (Macropus eugenii; Macropodidae) but is similar to that of a marsupial with invasive attachment (Monodelphis domestica; Didelphidae). We conclude that modes of embryonic attachment may not be typified by a particular suite of uterine changes in marsupials, as is the case for eutherian mammals, and that uterine remodelling may instead reflect phylogenetic relationships between marsupial lineages.     

Spermiogenesis in Soft‐Shelled Turtle,Pelodiscus sinensis

Spermiogenesis in the soft‐shelled turtle, Pelodiscus sinensis, was examined by transmission electron microscopy. The process includes nuclear elongation, chromatin condensation, acrosomal and flagellar development, and elimination of excess cytoplasm. In stage I, the proacrosomal vesicle occurs next to a shallow fossa of the nucleus, and a dense acrosomal granule forms beneath it. A smaller subacrosomal granule in the middle of the fibrous layer is related to the development of intranuclear tubules. The nucleus begins to move eccentrically. In stage II, the round proacrosomal vesicle is flattened by protrusion of the nuclear fossa, and the dense acrosomal granule diffuses into the vesicle, as the fibrous layer forms the subacrosomal cone. Circular manchettes develop around the nucleus, and the chromatin coagulates into small granules. The movement of the nucleus causes rearrangement of the cytoplasm. In stage III, the front of the elongating nucleus protrudes out of the spermatid and is covered by the flat acrosome; coarse granules replace the small ones within the nucleus. At the posterior pole of the head, mitochondria move backward. Numerous microtubules begin to assemble the axoneme of flagellum. In stage IV, the chromatin concentrates to dense homogeneous phase. The circular manchette is reorganized longitudinally. The Sertoli process covers the acrosome and the residues of the cytoplasmic lobes are eliminated. In stage V, the sperm head matures. After dissolution of the longitudinal manchette, the mitochondria arrange themselves around the proximal and distal centrioles. Caudal to the mitochondrial mass, a fibrous sheath surrounds the proximal portion of the flagellum. Anat Rec, 290:1213‐1222, 2007. © 2007 Wiley‐Liss, Inc.     

TCR gamma chain diversity in the spleen of the duckbill platypus (Ornithorhynchus anatinus)

TCR gamma (TRG) chain diversity in splenic gammadelta T cells was determined for an egg-laying mammal (or monotreme), the duckbill platypus. Three distinct V subgroups were found in the expressed TRG chains and these three subgroups are members of a clade not found so far in eutherian mammals or birds. Each subgroup contains approximately five V gene segments, and their overall divergence is much less than is found in eutherians and birds, consistent with their recent evolution from an ancestral V gene segment. The platypus TRG locus also contains three C region genes and many of the residues involved in TCR function, such as interactions with CD3, were conserved in the monotreme C regions. All non-eutherian mammals (monotremes and marsupials) lacked the second cysteine residue necessary to form the intradomain disulfide bond in the C region, a loss apparently due to independent mutations in marsupials and monotremes. Monotreme TRGC regions also had among the most variation in the length of the connecting peptide region described for any species due to repeated motifs.     

The origin and loss of the ubiquitin activating enzyme gene on the mammalian Y chromosome

Mammalian sex chromosomes are thought to be descended from a homologous pair of autosomes: a testis-determining allele which defined the Y chromosome arose, recombination between the nascent X and Y chromosomes became restricted and the Y chromosome gradually lost its non-essential genetic functions. This model was originally inferred from the occurrence of few Y-linked genetic traits, pairing of the X and Y chromosomes during male meiosis and, more recently, the existence of X- Y homologous genes. The comparative analysis of such genes is a means by which the validity of this model can be evaluated. One well-studied example of an X-Y homologous gene is the ubiquitin activating enzyme gene ( UBE1 ), which is X-linked with a distinct Y-linked gene in many eutherian ('placental') and metatherian (marsupial) mammals. Nonetheless, no UBE1 homologue has yet been detected on the human Y chromosome. Here we describe a more extensive study of UBE1 homologues in primates and a prototherian mammal, the platypus. Our findings indicate that UBE1 lies within the X-Y pairing segment of the platypus but is absent from the human Y chromosome, having been lost from the Y chromosome during evolution of the primate lineage. Thus UBE1 illustrates the key steps of 'autosomal to X-specific' evolution of genes on the sex chromosomes.      

Structural changes of the head components of the rat spermatid during late spermiogenesis

The transformation of the nucleus, acrosomic system, and perinuclear theca (perforatorium and post-acrosomal dense lamina) was analyzed during the maturation phase, i.e., steps 14 to 19 of spermiogenesis. Following partial condensation of chromatin from steps 11–14, the nucleus continues to condense during the following steps until the end of spermiogenesis. The redundant nuclear envelope which forms along the apical and ventral aspects of the nucleus and around the implantation fossa regresses during steps 17–19. The acrosomic system splits into two portions early in step 15 to give rise to: (a) the main portion with its crest-like acrosome running along the dorsal aspect of the nucleus and head cap extending over the lateral surfaces of the nucleus; and (b) a smaller head-cap segment which is seen in steps 15 and 16 along one side of the nucleus at its apical extremity. This separated head-cap segment reaches the apical-ventral aspect of the head during step 17 and condenses in synchrony with the rest of the acrosomic system in step 19 of spermiogensis. The large crescentic acrosome, which in step 15 forms a large fin at the caudal extremity of the acrosomic apparatus, moves anteriorly during steps 16 and 17, while the whole acrosomic system extends farther apically beyond the tip of the nucleus. The perforatorium and post-acrosomal dense lamina form a rigid capsule (perinuclear theca) that covers tightly the sickle-shaped nucleus and binds the inner acrosomal membrane and the post-acrosomal membranes. The post-acrosomal dense lamina, which includes the ventral spur, appears early in step 15 as a dense cytoplasmic layer applied to the nucler envelope at the caudal extremity of the nucleus except over the perifossal zone. The perforatorium forms during step 19 of spermiogenesis as a result of the condensation of a wispy cytoplasmic material which has accumulated in the subacrosomal space during steps 14–18. Thus the spermatid's head is deeply modified during the maturation phase and takes its definitive shape only at the last step of spermiogenesis.     

大鼠和小鼠精子发生过程中线粒体结构增殖和分布的研究

本研究采用超薄切片和冷冻蚀刻复型膜技术,制备大鼠和小鼠曲细精管样品,在透射电镜下观察各种生精细胞内线粒体的形态、数量、分布和膜结构。线粒体的增殖、发育和分布,与精子发生过程密切配合,表现为:1.线粒体的数量以休止期精原细胞最少;初级精母细胞线粒体自身分裂,迅速增殖,数量增加;精子细胞中数量比较恒定。2.线粒体的分布在精原细胞中随机分布,精母细胞内集中成群,高尔基期精子细胞线粒体分散在高尔基复合体附近或质膜下,头帽期、顶体期移至尾侧,成熟精子线粒体规则地排列在尾部中段密纤维外。3.线粒体的形态和发一育,在精原细胞线粒体为圆形或椭圆形,嵴平行,精母细胞线粒体延伸成杆状,并分裂形成新线粒体。新形成的线粒体缺少嵴,线粒体基质电子密度很低,成熟精子的线粒体嵴不规则。线粒体膜为三夹层结构,蛋白颗粒主要分布在外膜,内嵴蛋白颗粒很少,或缺少蛋白颗粒。     

An ultrastructural study of developing spermatids and associated Sertoli cells during spermiogenesis in the kowari (crest-tailed marsupial rat), Dasyuroides byrnei

Ultrastructural changes of developing spermatids and associated Sertoli cells during spermiogenesis in the kowari, Dasyuroides byrnei, were observed by transmission electron microscopy. The early round spermatid in the kowari showed an acrosomal vacuole containing sparsely distributed material. The acrosomal vacuole grew to some degree and then collapsed upon itself and decreased in size. After the diminution of the vacuole, flattening and condensation of the nucleus began. At this period, the manchette, nuclear ring and caudal plaque appeared and the ectoplasmic specialization of the Sertoli cell developed in association with the acrosomal region of the spermatid head. Microtubules of the manchette were arranged obliquely or perpendicular to the long axis of the spermatid. As the spermatid developed further, the nucleus displayed a horseshoe shape in cross section and was flattened in longitudinal section. The ectoplasmic specialization which was the most developed at this period appeared like horns protruding from the spermatid nucleus. Immediately before spermiation, the flattened aspect of the spermatid head contacted an apical process of the Sertoli cell. The Sertoli cell apical process extended the spermatid head into the lumen. Long tubulobulbar complexes appeared in the Sertoli cell, after the ectoplasmic specialization had dissociated.