The development of the oviducal gland in the Rajid thornback ray, <Emphasis Type="Italic">Raja clavata</Emphasis>

The reproductive processes of chondrichthyans are complex. Knowledge of the development and maturation of the oviducal gland is vital for understanding the reproductive biology of a species. This study represents the first contribution of this subject for skates. In the oviparous thornback ray, Raja clavata, oviducal gland development begins early in the developing stage with the formation of gland tubules and the distinct lamellae of each zone: club, papillary, baffle and terminal. Oviducal development is complete by the end of the developing stage when the storage and secretion of products is evident within the gland tubules of each zone. Periodic acid-Schiff and alcian blue histological staining showed that the secretory mucous cells of the club and papillary zones produce neutral and sulfated acid mucins. The last row of gland tubules of the papillary zone stains intensely for sulfated acid mucins. The baffle zone, which is responsible for the production of the egg capsule, represented 60–80% of the glandular zone of the oviducal gland. Sperm bundles were observed in the deeper recesses of the baffle zone during the maturation process, and during capsule extrusion, sperm were detected near the lumen. The terminal zone was composed of two types of gland tubules: serous (producing protein fibres) and mucous glands (producing sulfated acid mucins).     

Uterine epithelial-sperm interaction, endometrial cycle and sperm storage in the terminal zone of the oviducal gland in the placental smoothhound, Mustelus canis.

The fate of spermatozoa deposited within the female reproductive tract has been described in the smoothhound, Mustelus canis. Evidence of uterine epithelial-sperm interaction is presented, as well as documentation of sperm storage specifically in the terminal zone of the oviducal gland. Sperm fate is correlated with morphology of the endometrial cycle and specificity of storage in the oviducal gland. The endometrium of M. canis undergoes dramatic tissue remodeling associated with gestation. In females harboring fertilized ova or preimplantation yolk-reliant embryos, the uterine epithelium is simple cuboidal with mucous droplets for lubrication. The presence of the embryo elicits a response from the uterus, which becomes modified for nutrient and respiratory exchange into vascular uterine attachment sites that abut the distal aspect of the yolk sac. Areas of the uterus adjacent to the uterine attachment sites are termed paraplacental sites. Uterine attachment sites are simple squamous while the paraplacental epithelium is simple columnar. Paraplacental cells have basal metachromatic vesicles and a dense array of apical cytoplasmic filaments. Immediately postpartum the uterine attachment sites, now termed uterine or placental scars, begin to remodel to a mucous epithelium for the next gestational cycle. Paraplacental cells slough off the apical filamentous portion, and sperm become embedded in the epithelium. Bundled sperm occur throughout gestation in the terminal zone of the oviducal gland. Sperm are not embedded in the terminal zone epithelium as in the uterus. Following sperm release from the uterus, the paraplacental epithelium reverts to a mucous epithelium for the next reproductive cycle. Fertilization is presumed to occur in the anterior oviduct above the oviducal gland. The physiological mechanisms that mediate sperm-uterus attachment, release, and storage in the terminal zone of the oviducal gland are currently under investigation.     

Oviducal gland microstructure of Raja miraletus and Dipturus oxyrinchus (Elasmobranchii,Rajidae)

We studied the morphology and histology of the oviducal gland (OG) in the brown ray (Raja miraletus) and the long‐nosed skate (Dipturus oxyrinchus) to understand its functional role in the reproductive strategy of these species. The external morphology of the gland was similar in both species, with lateral extensions like those found in other members of the Rajidae. Microscopic analysis showed a similar internal organization in both species. Immature and developing glands did not react to histochemical techniques. On reaching maturity, the OG had the largest width due to an increase in the production of secretory materials. In both species, the club zone of the gland showed a strong reaction to Periodic acid‐Schiff (PAS) and alcian blue (AB) stains, indicating production of neutral and sulfated acid mucins. The secretory material produced by the papillary zone varied greatly between the two species. Both displayed tubular glands similar to those observed in the club zone, but in D. oxyrinchus the region near the lumen was intensely PAS+, whereas the last row of tubules of the brown ray stained intensely for a mixture of neutral and sulfated mucins. The baffle zone was the most conspicuous and extensive segment of all OGs, and it did not react to PAS/AB. The terminal zone, which is responsible for production of hair filaments, differed between the two species in terms of composition and organization of serous and mucous glands. This difference probably is related to the different substrates in which they release the egg capsules. Individual sperm detected in the brown ray baffle lamellae could be the result of a recent mating, whereas their presence in the deep recesses of the baffle and in the terminal zone of the long‐nosed skate might indicate sperm storage. J. Morphol. 276:1392–1403, 2015. © 2015 Wiley Periodicals, Inc.     

Evidence for sperm storage in common smoothhound shark Mustelus mustelus and paternity assessment in a single litter from South Africa

The present histological study of adult female common smoothhound shark Mustelus mustelus demonstrates that the species can store sperm in their oviducal glands. In addition, single paternity of a litter of 22 pups is reported, contributing to further knowledge regarding the genetic and behavioural mating strategy of M. mustelus.     

Sperm transfer and storage in the lizard,Anolis carolinensis

The relationship of the hemipenis to the cloaca in copula and sperm storage and transport in the female oviduct were studied in Anolis carolinensis using light and scanning electron microscopy. During copulation, the hemipenis does not penetrate beyond the cloaca, but the two apical openings of the bifurcate sulcus spermaticus appose the openings of the oviducts from the cloaca. Sperm enter the sperm storage tubules between 2 and 6 hr after insemination and small amounts of sperm reach the infundibulum 6 to 24 hr following mating. Sperm storage tubules are embedded in the wall of the utero-vaginal transition, and are formed by the folding and fusion of the oviducal epithelium. The importance of the hemipenile-cloacal relationship and the role of sperm storage in the life history of A. carolinensis are discussed.     

Ultrastructure of the reproductive system of the black swamp snake (Seminatrix pygaea): Part I. Evidence for oviducal sperm storage

Oviducal sperm storage in the viviparous (lecithotrophic) colubrid snake Seminatrix pygaea was studied by light and electron microscopy. Out of 17 adult snakes examined from May–October, sperm were found in the oviducts of only two specimens. In a preovulatory female sacrificed 14 May, sperm were found in the oviducal lumen and sperm storage tubules (SSTs) of the posterior infundibulum. In a nonvitellogenic female sacrificed 9 June, sperm were found in the lumen and glands of the posterior uterus and anterior vagina, indicating a recent mating. The glands in the posterior infundibulum and vagina were simple or compound tubular, whereas glands in the uterus always were simple tubular. The epithelium of the sperm storage glands was not modified from that lining the rest of the oviduct. The cuboidal or columnar epithelium consisted of alternating ciliated and secretory areas. The secretory product released into the lumen by a merocrine process contained mucoprotein. Lipid droplets also were numerous in the epithelium. Portions of sperm sometimes were embedded in the apical cytoplasm or in secretory material. A carrier matrix containing a mucoid substance, desquamated epithelium, lipids, membranous structures, and possibly phagocytes was found around sperm in the posterior uterus. J. Morphol. 241:1–18, 1999. © 1999 Wiley‐Liss, Inc.     

Seasonal changes of sperm storage and correlative structures in male and female soft-shelled turtles, Trionyx sinensis

Reproductive ducts of male and female soft-shelled turtles, Trionyx sinensis were examined throughout the year (March, May, September, December) using brightfield and electron microscopes (TEM and SEM), to determine the location and histomorphological characteristics of sperm storage structures as well as their changes at different phases of the seasonal reproductive cycle. Sperm stored in the epididymis were also examined. In the male, spermatogenesis is initiated in spring (May), and then the mature sperm are released in autumn as an episodic event. Spermatogenesis is inactive in winter. However, in this species, the epididymis contains sperm throughout the entire year. Sperm observed in the epididymis are intact and some structures are uniquely different from other reptiles, and is characterized by 35–40 concentric mitochondria with a dense core in the centre. Many glycogen granules are observed in the cytoplasm of the midpiece. However, the epithelial cell type of epididymal duct change in different seasons. The cells are fully developed with a highly secretory activity in September. The materials secreted from the epithelium might have the function as nourishment for the stored sperm. Sperm storage structures in the form of tubules are observed in the wall of the isthmus of the oviduct in hibernating females but are absent in the groups of May and September. These tubules develop either by folding or fusion of the oviductal mucosal folds and are lined by both ciliated and secretory cells. These tubules might provide a microenvironment for the sperm to enable its long-term storage. After being separated 4 months (December–March) from the male, sperm are observed in the tubules of the isthmus of the oviduct. The unique character of the sperm combined with the special sperm storage structures enable the sperm to maintain fertility and activity during their storage.     

Multiple paternity in the starry smooth‐hound shark Mustelus asterias (Carcharhiniformes: Triakidae)

Multiple paternity was investigated in the starry smooth‐hound shark, Mustelus asterias Cloquet, 1821. Analysis of 12 pregnant females and their embryos, at four microsatellite loci, showed that at least 58% carried multiply‐sired litters. Paternal skew was observed, with one male siring most of the embryos within a litter, although no patterns of association were detected between the identity of the father and the size or the uterine position of the embryos. Sperm storage was observed in the oviducal glands of all twelve females. Results shed light on the complexity of reproductive strategies and selective processes in M. asterias. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 119–125.     

Microanatomy of the male and female reproductive tracts in the long‐tailed butterfly ray Gymnura poecilura,an elasmobranch with unusual characteristics

The anatomy of the male and female reproductive systems was investigated in the long‐tailed butterfly ray Gymnura poecilura using gross observation and light microscopy. The testes are highly asymmetrical, to the extent that only the left testis is functional and the right testis is completely absent. Both of the male genital ducts are present and symmetrical, although spermatozoa only occur in the left duct. The genital ducts are straight and unconvoluted, with regular incomplete internal partitions throughout. Females do not possess a right ovary, nor do the oviducal glands exhibit distinct club and papillary zones, and the baffle zone lacks baffle plates. In all sections of the gland, the tubules display different secretory activities depending on the proximity to the gland lumen. The gland produces a thin egg membrane that encases each egg individually, while the endometrium is formed into trophonemata.     

Seasonal variation in the oviduct of female Agkistrodon piscivorus (Reptilia:Squamata): An ultrastructural investigation

The annual oviductal cycle of the Cottonmouth, Agkistrodon piscivorus, is described using electron microscopy. This is only the second such study on a snake and the first on a viperid species. Specimens were collected in reproductive and nonreproductive condition throughout the year and five ultrastructurally unique regions were recognized: the anterior infundibulum, posterior infundibulum, glandular uterus, nonglandular uterus, and vagina. Except for the anterior infundibulum and vagina, which exhibit no seasonal variation in ultrastructure, the oviduct becomes highly secretory at the start of vitellogenesis. This includes the entire luminal border of the uterus, the tubular glands of the glandular uterus, and the luminal border and sperm storage tubules of the posterior infundibulum. The secretory materials produced in the oviduct vary among regions of the oviduct, and also can vary among time periods in the same region of the oviduct. Variation is especially evident in the sperm storage tubules. Secretory activity in the sperm storage tubules ceases after ovulation, but the tubular glands of the glandular uterus remain secretory until parturition, at which time secretory activity in the varying sections of the oviduct decreases dramatically. After parturition, the oviduct remains in a dormant state until the next reproductive season. The seasonal variation in oviducal morphology mirrors the temperate primitive reproductive cycle known for some pitvipers. Uterine glands of A. piscivorous are more similar in secretory activity to those of an oviparous lizard than a viviparous colubrid snake, suggesting variation in uterine gland morphology between snakes of different families. J. Morphol., 2008. © 2008 Wiley‐Liss, Inc.     

Oviductal sperm storage structure and their changes during the seasonal (dissociated) reproductive cycle in the soft-shelled turtle Lissemys punctata punctata

The oviduct of the Indian fresh water soft-shelled turtle Lissemys punctata punctata was examined throughout the year under light and scanning electron microscopes to determine the location, histomorphological characteristics, and function of sperm storage structure, as well as their changes at different phases of the seasonal reproductive cycle. Sperm storage structures in the form of tubules were observed in the wall of isthmus throughout the year. These tubules developed either by folding or fusion of the oviductal mucosal folds and were lined by both ciliated and nonciliated epithelial cells. The height and secretory activities of the epithelia were markedly high during the breeding phase (August to September) but low in the nonbreeding phase (October to June). A few short tubules lined by cuboidal epithelium appear in the wall of infundibulum only during the breeding phase. Following mating (May), inseminated sperm were stored within the tubules of isthmus up to the pre-ovulatory stage (August). Thereafter, sperm associated with PAS-positive materials secreted from the epithelium (referred to as a carrier matrix) moved forward to the infundibulum and were stored within the storage tubules of the infundibulum for a short time. Subsequently, sperm evacuated the storage tubules and entered the oviductal lumen to fertilize the subsequently ovulated eggs during or prior to ovulation. The isthmus-tubules become shorter and narrower in the regressive phase (October to November) and remained so until the early preparatory phase (April). Sperm release might have been stimulated by estrogen secreted from the ovarian follicles of pre-ovulatory turtles. Stored sperm not utilized for fertilization remained viable not less than six months in the present turtle species.     

Sperm aggregations in the spermatheca of the red back salamander (Plethodon cinereus)

The spermatheca of Plethodon cinereus is a compound tubular gland that stores sperm from mating in early spring (March–April) to oviposition in summer (June–July). The seasonal variation of sperm storage in this species has previously been studied by light and transmission electron microscopy. In this paper, sperm aggregations, interaction of sperm with the spermathecal epithelium, and spermathecal secretions are studied using scanning electron microscopy. Within spermathecal tubules, relatively small groups of sperm are aligned along their entire lengths in parallel arrays. This pattern is similar to other plethdontids with complex spermathecae. Lumina of spermathecal tubules are filled with secretory material in April prior to the arrival of sperm, and after sperm appear, a coating of secretory material persists on the apices of the spermathecal epithelium. Sperm peripheral to the central luminal mass can become embedded in the secretory matrix or pushed deeper into the spermathecal epithelium. The spermathecal secretions may serve to attract and prolong the viability of sperm, but sperm that become enmeshed in the secretions or epithelium are phagocytized. Sperm and spermathecal secretions are largely absent after ovulation and in summer months, and new secretory vacuoles are formed in fall, although mating does not occur until spring.     

Numbers and size of sperm storage tubules and the duration of sperm storage in birds: a comparative study

Estimates of the numbers of sperm storage tubules (SSTs) in the utero-vaginal junction of 11 bird species are presented. Numbers of SSTs varied by a factor of 40 between species, and ranged from 500 to 20000. Body mass accounted for over 50% of the variation in SST mumbers. SST length was positively correlated with the length of spermatozoa across species. The duration of sperm storage was not correlated with the number of SSTs or the volume of sperm storage tissue. However, the number of 'active' SSTs appears to vary between species and it was not possible to make allowance for this. Sperm storage duration was weakly, positively correlated wth clutch size, but showed a significant positive relationship with the number of days over which laying occurred. The number of SSTs was also positively correlated with the number of sperm per ejaculate. The best predictor of sperm storage duration was a multiple regression equation using the spread of laying and the length of sperm storage tubules. The duration of sperm storage in birds which remain together during the pre-laying period is such that a single insemination immediately before the start of laying could fertilize the entire clutch.     

Optimization of handling and refrigerated storage of guppy Poecilia reticulata sperm

Sperm collection methods and the effect of osmolality, ions, sugar, temperature, pH and dilution ratio on sperm motility were investigated in guppies Poecilia reticulata. The present study revealed that the sperm was motile in a wide range of osmolalities (200–470 mOsm kg^?1) either in Hanks balanced‐salt solution (HBSS) or in non‐electrolyte solutions such as glucose or sucrose. Sperm collected from crushing testes yielded lower motility and shorter motility duration than samples collected without crushing but gentle disruption. Dilution ratios within the range of 1:50 to 1:500 of sperm to HBSS had minimal effect on sperm motility during extended refrigerated storage. Examination of storage temperature showed that refrigerated storage at 4° C was superior to room temperature (25° C). Sperm was found to tolerate a wide range of pH from 5·6 to 7·8, but motility was affected negatively by pH values >7·8.     

Sperm aggregations in female Agkistrodon piscivorus (Reptilia:Squamata): a histological and ultrastructural investigation

Upon copulation in female Agkistrodon piscivorus, sperm migrate up the oviduct to sperm storage tubules (SSTs) in the posterior infundibulum. The epithelium of the SSTs is composed of ciliated and secretory cells and differs ultrastructurally from that of the epithelium lining the lumen of the posterior infundibulum. Sperm pass through an area composed primarily of ciliated cells at the opening of each gland before aligning themselves in parallel arrays with their nuclei facing an area composed primarily of secretory cells at the base of the tubules. Sperm are also found embedded inter- and intracellularly in the SSTs. The secretory vacuoles in the SSTs become highly electron dense after the start of the fall mating season along with the synthesis of lipid droplets. Histochemical analysis reveals that the alteration in secretory material density is caused by the production of neutral carbohydrates. Some sperm remain in aggregates in the nonglandular section of the posterior uterus until the time of ovulation. However, ultrastructural evidence indicates these sperm degrade before ovulation. Therefore, sperm in posterior aggregates have no role in fertilization of ovulated ova. The data presented here support the hypothesis that infundibular sperm storage is the mode that snakes utilize to sequester viable sperm until ovulation.     

Sperm storage within the oviduct of turtles

Tubules containing sperm were identified by light microscopy in the oviducts from 11 species of turtles representing six different families. Sperm storage tubules were found in a small region of the posterior portion of the egg albumin-secreting section of the oviduct located between the infundibulum and the uterus. This location of storage tubules, midway between the ovary and vagina, is unique among vertebrates. Ducts, restricted to the posterior albumin region, connect the tubules to the oviduct lumen, allow entrance of sperm to the tubules. Sperm were identified in tubules of female turtles isolated from males for as long as 423 days.     

Sperm aggregations in the spermatheca of female desmognathine salamanders (Amphibia: Urodela: Plethodontidae)

The alignment of sperm in a cloacal sperm storage gland, the spermatheca, was studied in female desmognathine salamanders by scanning and transmission electron microscopy. Females representing nine species and collected in spring, late summer, and fall in the southern Appalachian Mountains contained abundant sperm in their spermathecae. The spermatheca is a compound tubuloalveolar gland connected by a single common tube to the middorsal wall of the cloaca. Sperm enter the common tube in small groups aligned in parallel along their axes, and continue in a straight course until encountering divisions of the common tube (neck tubules) or luminal borders of distal bulbs, which can act as barriers. Sperm may form tangles, in which small clusters retain their mutual alignment, at the branches of the neck tubules from the common tube, or in the lumen of the distal bulbs, where subsequent waves of sperm collide with sperm already present. The nuclei of some sperm from the initial group to encounter the walls of the distal bulbs appear to become embedded in secretory material on the luminal border or in the apical cytoplasm of the spermathecal epithelial cells. We propose that these sperm become trapped in the spermatheca and are ultimately degraded. J. Morphol. 238:143–155, 1998. © 1998 Wiley-Liss, Inc.