Histochemical, enzyme histochemical and ultrastructural investigation on the sperm-duct glands of Padogobius martensi (Pisces, Gobiidae)

The functions of the sperm-duct glands of Padogobius martensi were investigated by histochemical, enzyme histochemical and fine structural methods during the prereproductive and reproductive periods, i.e. during their maximum activity. The epithelium lining the spermduct gland tubules produces strongly acidic sulphomucins, probably conjugated with proteins and the variety of sulphomucins increases during the reproductive period when the exocrine epithelium is also involved in the phagocytosis of degenerating sperm cells. A contractile network of myoid cells surround the tubules. The gland interstitium contains cells similar in enzyme histochemical and ultrastructural features to the testicular interstitial cells (Leydig cell homologues). Unlike the latter, however, the interstitial cells of the sperm-duct gland are able to produce steroid glucuronides, which act possibly as pheromones.     

The digestive gland of Viviparus ater (Mollusca, Gastropoda, Prosobranchia): an ultrastructural and histochemical study

The digestive gland of Viviparus ater was studied using histochemical and ultrastructural methods. Only one cell type was observed in the tubule epithelium of the gland. The cells are involved in an endocytotic process mediated by clathrin-coated vesicles and in the intracellular digestion of food materials (thus they can be regarded as digestive cells). The different stages of digestion and exocytotic extrusion of residual bodies into the tubule lumen were shown by electron microscopy. Very few, small mucocytes are scattered among the digestive cells. Calcium concretions, glycogen-containing cells and endocrine cells are scattered in the area of connective tissue present among the digestive tubules.     

Histological, fine-structural and histochemical differences in the testicular glands of gobiid and blenniid fishes

The testicular gland (t.g.) is a glandular tissue situated adjacent to the testis of blenniid and several gobiid species. In the present study the t.g. of Blennius pavo Risso and Gobius niger L. were compared by histological and histochemical methods. In B. pavo the spermatozoa have to cross the t.g. to reach the vas deferens and thus they come into contact with the gland cells, whereas in G. niger the vas deferens is situated between the testis and the t.g. The fine structure and histo-chemistry of the t.g. cells reveal that in B.pavo the cells of the t.g. have exocrine as well as endocrine functions. The t.g. cells of B. pavo contain large amounts of lipids, form a secretion containing acid mucopolysaccharides, show positive reaction for acid phosphatase, and some cells stain for 3β-HSD and G6PD. The function of the t.g. of G. niger is exclusively endocrine. Characteristics of the gland cells of this species are well developed smooth ER and tubulovesicular or paracristal-line mitochondria. The stainings for 3β-HSD, G6PD and UDPGD give strong positive results in the whole t.g., indicating the presence of steroids and steroid glucuronides.     

Absence of cross-linking via trans-glutaminase in barnacle cement and redefinition of the cement

Balanomorphan barnacles attach their calcareous bases to a variety of substrata, including others of the same species, through secretion of an underwater adhesive, commonly referred to as cement. In this multi-functional process of underwater attachment, curing of the adhesive is crucial for the formation of a secure attachment. To date, there has been no direct evidence presented to suggest the involvement of cross-linking or polymerization in the cement curing process, despite the emergence of this hypothesis in the recent literature. A recently proposed mechanism for cement curing involves glutamyl-lysine cross-linking via the action of trans-glutaminase. However, in the opinion of the author, inadequate attention may have been paid to sample collection during the study and the conditions used in the analysis may not be adequate to support the conclusions of the paper. Indeed, further investigation, the results of which are presented here, did not provide any evidence to support adhesive curing via glutamyl-lysine cross-linking. Therefore, the hypothesis that the process of cement curing is similar to the clotting system of barnacle hemolymph is not compatible with the data reported so far. In order to allay any potential confusion, a new definition of the barnacle cement is proposed.     

Ultrastructural and histochemical comparison in two haplotaxids

Pelodrilus leruthi Hrabe, 1958 and Haplotaxis gordioides Hartman, 1821 have been investigated from a histochemical point of view. A different distribution and number of fast and slow fibres has been detected in the two species. Pelodrilus leruthi specimens living in different caves, show some differences in the body wall muscle thickness and in the structure of the ventral nerve cord. A correlation with their different life styles is proposed.     

The endocrine cells in the gastroenteropancreatic system of the bowfin, Amia calva L.: an immunohistochemical, ultrastructural, and immunocytochemical analysis.

The gastroenteropancreatic (GEP) endocrine system of bowfin (Amia calva) was described using light and electron microscopy and immunological methods. The islet organ (endocrine pancreas) consists of diffusely scattered, mostly small islets and isolated patches of cells among and within the exocrine acini. The islets are composed of abundant, centrally located B cells immunoreactive to bovine and lamprey insulin antisera and D cells showing a widespread distribution and specificity to somatostatin antibodies. A and F cells are present at the very periphery of the islets and are immunoreactive with antisera against glucagon (and glucagon-like peptide) and several peptides of the pancreatic polypeptide (PP)-family, respectively. The peptides of the two families usually collocates within the same peripheral islet cells and are the most common immunoreactive peptides present in the extra-islet tissue. Immunocytochemistry and fine structural observations characterised the granule morphology for B and D cells and identified two cell types with granules immunoreactive to glucagon antisera. These two putative A cells had similar granules, which were distinct from either B or D cells, but one of the cells had rod-shaped cytoplasmic inclusions within cisternae of what appeared to be rough endoplasmic reticulum. The inclusions were not immunoreactive to either insulin or glucagon antisera. Only small numbers of cells in the stomach and intestine immunoreacted to antisera against somatostatin, glucagon, and PP-family peptides. The paucity of these cells was reflected in the low concentrations of these peptides in intestinal extracts. The GEP system of bowfin is not unlike that of other actinopterygian fishes, but there are some marked differences that may reflect the antiquity of this system and/or may be a consequence of the ontogeny of this system in this species.     

Histological,histochemical, and ultrastructural investigation of the male copulatory apparatus of Haminoea navicula (Gastropoda,Cephalaspidea)

Due to its biological and systematic importance, the morphology and function of the male copulatory apparatus of Haminoea navicula, a Cephalaspidea gastropod mollusk, was investigated by light and electron microscopy. These systems are poorly understood in haminoids, but are often used in the taxonomy of the genus. In H. navicula, the male copulatory apparatus comprises the penis within a penial sheath, a seminal duct and the prostate with two lobes. The penis is a muscular structure with a tip covered by spikes formed by muscular cells. The penial sheath consists of muscular tissue folds lined by an epithelium. Below this epithelium, polysaccharide‐secreting cells and pigment cells were observed. A large number of vacuolar cells were found below the ciliated epithelium of the seminal duct. The proximal lobe of the prostate was formed by tubules that could be divided in basal, middle and apical zones, containing cells that secrete polysaccharides and proteins. The tubules of the prostate distal lobe contained a single type of secretory cells with vesicles that were stained by histochemical techniques for detection of polysaccharides and proteins. Ciliated cells were present along the tubules in both lobes of the prostate. This study revealed a complex prostate with five types of secretory cells, which secrete substances that should be involved in the maintenance and eventually also in the maturation of spermatozoa. A comparison with previous publications, shows that the male copulatory apparatus can differ substantially among cephalaspideans, even between H. navicula and non‐European species attributed to this genus.     

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.     

Structural and histochemical changes in the salivary glands of Rhipicephalus appendiculatus during feeding

The salivary glands of the brown ear tick of cattle, R. appendiculatus, from both sexes and at all stages of feeding, were examined as whole glands and as sections for ultrastructural and histochemical changes. The type 1 acinus consists of a basal labyrinth formed by the interdigitations of a central cell and four peripheral cells. These cells form a specialized border with a central constrictor cell which surrounds the acinar duct. The plasma membrane of the central cell is exposed to the duct. The type 1 acini do not appear to secrete active saliva components involved in feeding. The type 2 acini undergo a great increase in synthetic and secretory activity during feeding in both sexes and secrete a lipoprotein probably to form part of the attachment cone and also glycoproteins and esterases of unknown functions. The type 3 acini of both sexes also secrete a lipoprotein probably to form part of the attachment cone. The f cells of these acini in the females transiently secrete a glycoprotein of unknown function and then transform to become part of a water excreting unit. In the males the secretory activity of the granular cells of the type 2 and 3 acini is maintained for further attachments. The type 4 acini of the males accumulate masses of proteinaceous granules. The system of interstitial cells and intercellular spaces in types 2, 3 and 4 acini is large and increasingly active during feeding.     

An ultrastructural investigation of the testes and spermiogenesis in Myobia murismusculi (Schrank) (Acari,Actinedida: Myobiidae)

Summary

The stages of spermiogenesis in Myobia murismusculi were investigated on the basis of ultrastructural analysis of both the testes and the female organs: receptaculum seminis and seminal duct. The walls of the testes consist of a thin epithelial layer. Germ and secretory cells lie free in the lumen of the testes. In the early stages of differentiation, both cell types represent clusters of sister cells joined by intercellular bridges. Each secretory cell contains prominent RER and Golgi complex, which produce single dense granule. Growing gradually the granule fills the whole volume of the cell's cytoplasm. Mature secretory cells disintegrate and the secretory product discharges into the testicular lumen. The germ cells are represented by the early, the intermediate and the late spermatids as well as the immature sperm (prospermia). Neither spermatogonia nor meiotic figures were observed in adult males. As spermiogenesis starts, numerous narrow invaginations of the outer membrane (peripheral channels) develop on the cell surface. They form a wide circumferential network connected to pinocytotic vesicles. Owing to the secretory activity of the Golgi complex, a large acrosomal granule is formed in the early spermatids. A long acrosomal filament runs along the intranuclear canal. Nuclear material condenses and forms two spherical bodies of different electron density. The lighter one can be observed until the stage of the late spermatids, when the nuclear envelope almost completely disappears. The electron-dense nuclear body transforms into a definite chromatin body, which is observed in the mature sperm as a cup-shaped structure. The late spermatids are characterized by the presence of a large electronlucent vacuole, which seems to be unique for the process of spermiogenesis in Actinedida. After the spermia enter the female genital tract, the peripheral channels disappear as well as the vacuole. The cells form long amoeboid arms with a special microtubular layer underneath the plasma membrane. The chromatin body is encircled by a large acrosomal granule of complex shape provided by long extensions running deep into the cytoplasm. The cytoplasm contains no organelles except for a group of unmodified mitochondria in the post-nuclear region. The main characteristics of the Myobia spermiogenesis are discussed with regard to other actinedid mites.     

Ultrastructural and histochemical study of the gallbladder epithelia of rainbow trout and tench

The simple, high columnar gallbladder epithclia of the rainbow trout (Salmo gairdneri, Rich.) and the tench (Tinca linca, L.) are composed of light and dark cells. Within the dark cells two types must be distinguished: less hydrated, but normal epithelial cells and degenerating cells. The latter seem to be eliminated from the epithelium through phagocytosis by monocytes and macrophages, which possibly correspond with the ‘basal regenerative cells’ of earlier investigations. Since they are mainly found near the basement membrane, an hypothesis is put forth that they do not enter the gallbladder lumen, but migrate back into the subepithelial connective tissue after phagocytosis. Light and less hydrated dark cells are specialized for absorption of water and dissolved bile salts and for secretion of mucus, which mainly consists of acid glycoproteins. Interspecific differences are seen with regard to both functions. So, in the tench only single microvilli arc developed apically, whereas a true brush border is seen in the rainbow trout. Thus, a higher absorptive rate can be assumed in the rainbow trout. Or, with respect to mucus secretion, in the rainbow trout the granular endoplasmic reticulum predominates, whereas in the tench the smooth endoplasmic reticulum, often associated with glycogen and even forming ‘glycogen bodies’, is better developed. In the rainbow trout fibrillar granules of highly condensed acid glycoproteins are seen to be secreted. In the tench only vesicles of flocculent content occur. Perhaps, these morphological and subsequent metabolic differences may be related to the various feeding habits of the predatory rainbow trout and the omnivorous tench. Seasonal variations as expected especially in the tench after winter rest could not be established.     

Variation among families for characteristics of the adhesive plaque in the barnacle Balanus amphitrite

A quantitative genetics approach was used to examine variation in the characteristics of the adhesive plaque of the barnacle Balanus amphitrite Darwin attached to two silicone substrata. Barnacles settled on silicone polymer films occasionally form thick, soft adhesive plaques, in contrast to the thin, hard plaques characteristic of attachment to other surfaces. The proportion of barnacles producing a thick adhesive plaque was 0.31 for Veridian, a commercially available silicone fouling-release coating, and 0.18 for Silastic T-2, a silicone rubber used for mold-making. For both materials, significant variation among maternal families in the proportion of barnacles producing a thick adhesive plaque was observed, which suggests the presence of genetic variation, or maternal environmental effects, for this plaque characteristic. For the Veridian coating, barnacles expressing the thick adhesive plaque also exhibited significantly reduced tenacity. This represents the first reported case for potential genetic control of intraspecific phenotypic variation in the physical characteristics and tenacity of the adhesive of a fouling invertebrate.     

An ultrastructural investigation of the early stages of oocyte differentiation in Actinia fragacea (Cnidaria; Anthozoa)

Individuals from a population of the intertidal sea anemone Actinia fragacea (Tugwell) were collected at approximately monthly intervals over an 18 month period. Samples of gonad were removed from each anemone and examined by light and electron microscopy. During late spring and early summer, large numbers of small cells were seen in the endoderm of the female gonads, lying close to the mesoglea. For convenience, these cells were classified into three types. Type I cells are 6–9 μm in diameter, with relatively very large nuclei, which may contain synaptinemal complexes, and scant cytoplasm containing few organelles. Type II cells are larger, reaching 15 μ m in diameter, with more abundant cytoplasm containing more organelles and inclusions. The nucleus is more dense, but may also contain synaptinemal complexes. Type III cells are less common. They are similar in size to Type II cells, but their nuclei contain irregular dense chromatin masses, and the nuclear envelope is incomplete or absent. The possible significance of the various cell types is discussed. It is suggested that Type I cells are oocytes at a very early stage of differentiation and that Type II cells are rather later oocytes. The status of the Type III cells is uncertain.     

Morphological and histochemical features of the digestive tract of Leiarius marmoratus (Gill, 1870)

Leiarius marmoratus, a freshwater catfish from Pimelodidae family, shows great biological and commercial relevance because of its geographic distribution and adaptation to fish-farm. The knowledge of the morphological characteristics of the digestive tract is fundamental to the understanding of fish physiology and nutrition, which helps in the planning of diets to provide better management and success in fish farming. Thus, this work described the morphology and histochemistry of the digestive tract of L. marmoratus adults. After euthanasia, the animals were dissected for analysis of the digestive tract. The oesophagus is a short and distensive organ with longitudinal folds that allow the passage of large food, e.g., other fishes. Oesophageal mucosa layer shows a stratified epithelium with goblet cells and club cells. The secretion of goblet cells is composed of neutral and acidic mucins that are anchored in the epithelium luminal face by epithelial cells fingerprint-like microridges, lubricating the surface to facilitate the food sliding. Club cells have protein secretion that can be involved in alarm signals when epithelium is damaged and in immunological defence. The saccular stomach is highly distensible to store large food. Gastric mucosa layer is composed of epithelial cells with intense secretion of neutral mucin to protect against self-digestion of gastric juice. Cardiac and fundic regions of stomach show well-developed gastric glands composed of oxynticopeptic cells. These cells have numerous mitochondria, highlighting their intense activity in the synthesis of acid and enzymes. The intestine is divided into three regions: anterior, middle and posterior. Although it is a short tube, intestine shows longitudinal folds and microvilli of enterocytes to increase the contact surface. These folds are higher in the anterior region of the intestine, highlighting their function in digestion and absorption. Intestinal goblet cells have acidic and neutral mucins that lubricate the epithelium and aid in digestive processes. These cells increase in number towards aboral, and they are related to the protection and lubrication to expulsion of faecal bolus.     

Changes in the levels and distribution of glycogen during regeneration of cutaneous wounds in Heteropneustes fossilis: a histochemical investigation

Incisional wounds of the same length and depth were made in the skin of catfish Heteropneustes fossilis , and the response in terms of levels of glycogen in cells during healing was studied histochemically. After injury, glycogen disappeared completely from the cells in the migrating epidermis. This disappearance is correlated with the migration of the epidermal cells toward the wound gap for epithelialization and the increased production of mucous cells at the initial stages of healing. The gradual accumulation of glycogen in the epithelial cells and the basal cells after epithelialization of the wound during the early stages of healing suggests that in these cells gluconeogenesis predominates. The gradual disappearance of glycogen from the epithelial cells 72 h after injury and from the basal cells 36 h after injury indicates the gradual resumption of their metabolic activities. The increased gradients of glycogen in club cells that are undergoing vacuolization and disintegration are discussed. No significant levels of glycogen were observed during the formation of granulation tissue and biosynthesis of the sub-cutis. The depletion of glycogen in the degenerating muscle bundles is correlated with the synthesis of lactic acid which contributes to local acidity and favours the autolysis of muscles.     

An ultrastructural and cytochemical study of the digestive system in Oxyposthia praedator (Turbellaria,Acoela)

The central parenchyma in Oxyposthia praedator consists of multifunctional cells. These cells digest food material intracellularly, can effect extracellular digestion through release of digestive enzymes by cell lysis and clasmatosis, and synthesize reserve nutritional substances.     

Ultrastructural and histochemical observations on the tegument of Gastrodiscoides hominis (Paramphistoma: Digenea).

The tegument of the paramphistome, Gastrodiscoides hominis, is basically similar to that of other digeneans. It is folded into concentrically arranged furrows and ridges bearing numerous tightly packed tubercules, and extends into the oral cavity. An area of specialized tegument is present on the ventral surface, anterior to the disc region. Mitochondria are absent from the tegumental syncytium and underlying tegumental cells, suggesting that the tegument may serve principally as a protective layer rather than in active uptake phenomena. However, extensions of the lymph and parenchyma systems are closely associated with the base of the tegumental syncytium and may provide ATP for active processes. Ciliated and non-ciliated sensory papillae are present, particularly around the oral opening. Numerous lymph channels are present in the sub-tegument and may be involved in osmoregulation.     

Morphological and histochemical studies on the prostate gland of developing and adult Paramphistomum cervi (Digenea: Paramphistomatidae)

Morphological and histochemical studies have been made on the development of the prostate gland of the digenetic trematode, Paramphistomum cervi during the course of its infection in sheep; histochemical characterization of prostate gland secretion in adult worms and its functional relationship with the transport of spermatozoa have also been studied. In 6-wk-old worms, the terminal portion of the male genital duct is formed of what appears to be a syncytial epithelium containing a relatively large number of nuclei compared to the rest of the male duct. Cellular organization of the prostate gland becomes conspicuous in 8-wk-old worms and the prostate gland is fully developed by 16 wk. Two types of prostate gland cells, characteristic of the adult, become distinct in 16-wk-old worms which contain spermatozoa in the lumen of their vas deferens and pars prostatica. Adult worms show two types of prostate gland cells: type I containing mainly glycoprotein and type II mainly phospholipid granules. Weak-to-strong activities of acid phosphatase, alkaline phosphatase, esterase, lipase, adenosine triphosphatase, tetrazolium reductase, NAD-diaphorase, isocitrate dehydrogenase, glyceraldehyde dehydrogenase, α-glycerophosphate dehydrogenase and glucose-6-phosphate dehydrogenase have been observed in the prostate gland. The release of prostate gland secretion and the passage of spermatozoa through the lumen of pars prostatica appear to be synchronized events.