Crustacean hemocytes are important mediators of immune functions such as coagulation and phagocytosis. We employed an in situ approach to investigate the ultrastructural behavior of hemocytes during coagulation and phagocytosis in the early stages after injury caused by leg amputation, using transmission electron microscopy technique in marbled crayfish Procambarus virginalis. Hemocytes underwent drastic morphological changes during coagulation. The morphology of the cytoplasmic granules changed from electron-dense to electron-lucent forms in an expanding manner. The transformed granules containing amorphous electron-lucent material were observed to merge and discharge their contents into extracellular space for coagulation. We also observed that the contents of the nucleus participate in the process of coagulation. In addition, leg amputation induced extensive muscle degeneration and necrotic tissues were avidly taken up by the phagocytic hemocytes containing distinct phagosomes. Interestingly, we observed for the first time how the digested contents of phagocytized necrotic tissues are incorporated into granules and other cellular components that change the cell morphology by increasing the granularity of the hemocytes. Nevertheless, the degranulation of hemocytes during coagulation can also reduce their granularity. Given that morphological traits are important criteria for hemocyte classification, these morphological changes that occur during coagulation and phagocytosis must be taken into account.
- MeSH
- Arthropods * MeSH
- Phagocytosis MeSH
- Phagosomes MeSH
- Hemocytes * MeSH
- Astacoidea MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Understanding the molecular basis of sexual dimorphism in the cardiovascular system may contribute to the improvement of the outcome in biological, pharmacological, and toxicological studies as well as on the development of sex-based drugs and therapeutic approaches. Label-free protein quantification using high-resolution mass spectrometry was applied to detect sex-based proteome differences in the heart of zebrafish Danio rerio. Out of almost 3000 unique identified proteins in the heart, 79 showed significant abundance differences between male and female fish. The functional differences were mapped using enrichment analyses. Our results suggest that a large amount of materials needed for reproduction (e.g., sugars, lipids, proteins, etc.) may impose extra pressure on blood, vessels, and heart on their way toward the ovaries. In the present study, the female's heart shows a clear sexual dimorphism by changing abundance levels of numerous proteins, which could be a way to safely overcome material-induced elevated pressures. These proteins belong to the immune system, oxidative stress response, drug metabolization, detoxification, energy, metabolism, and so on. In conclusion, we showed that sex can induce dimorphism at the molecular level in nonsexual organs such as heart and must be considered as an important factor in cardiovascular research. Data are available via ProteomeXchange with identifier PXD023506.
- MeSH
- Zebrafish * genetics MeSH
- Sex Characteristics * MeSH
- Zebrafish Proteins * MeSH
- Proteome genetics MeSH
- Proteomics MeSH
- Heart * MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Concerns regarding the potential toxic effects of zinc oxide nanoparticles (ZnO NPs) on aquatic organisms are growing due to the fact that NPs may be released into aquatic ecosystems. This study aimed to investigate the effects of dietary exposure to ZnO NPs on juvenile common carp (Cyprinus carpio). Fish were fed a spiked diets at doses 50 and 500mg of ZnO NPs per kg of feed for 6 weeks followed by a 2-week recovery period. Fish were sampled every 2 weeks for haematology trends, blood biochemistry measures, histology analyses, and determination of the accumulation of zinc in tissues. At the end of the exposure and post-exposure periods, fish were sampled for an assessment of lipid peroxidation levels. Dietborne ZnO NPs had no effects on haematology, blood biochemistry, and lipid peroxidation levels during the exposure period. After the recovery period, aspartate aminotransferase activity significantly (p < 0.05) increased and alanine transferase activity significantly (p < 0.05) decreased in the higher exposure group. The level of lipid peroxidation significantly (p < 0.05) decreased in liver of treated fish after 2 weeks post-exposure period. A histological examination revealed mild histopathological changes in kidneys during exposure. Our results did not show a significant increase of zinc content at the end of experiment in any of tested organs. However, chronic dietary exposure to ZnO NPs might affect kidney and liver function.
- MeSH
- Time Factors MeSH
- Water Pollutants, Chemical metabolism toxicity MeSH
- Diet MeSH
- Liver drug effects metabolism MeSH
- Carps metabolism physiology MeSH
- Metal Nanoparticles toxicity MeSH
- Kidney drug effects metabolism MeSH
- Environmental Monitoring methods MeSH
- Zinc Oxide metabolism toxicity MeSH
- Oxidative Stress drug effects MeSH
- Lipid Peroxidation drug effects MeSH
- Tissue Distribution MeSH
- Dose-Response Relationship, Drug MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
Detection of patterns of subcellular calcium distribution in the cardiovascular system can contribute to understanding its role in cardiac and blood function. The present study localized calcium in heart atrium, ventricle, and bulbus arteriosus as well as in erythrocytes of zebrafish Danio rerio using an oxalate-pyroantimonate technique combined with transmission electron microscopy. Intracellular calcium stores were detected in caveolae, mitochondria, and the nuclei of several zebrafish cardiac cell types. Melanin pigmentation containing calcium stores was detected in the pericardial cavity. Melanin might be an extracellular source of calcium for heart beating and/or a lubricant to prevent friction during beating process. Calcium deposits were also detected in the plasma membrane, cytoplasm and nucleus of erythrocytes as well as in blood plasma. Possible exchange of calcium between erythrocytes and blood plasma was observed. Interactions of such calcium stores and possible contribution of extracellular calcium stores such as melanin pigmentation to supply calcium for vital functions of heart cells should be addressed in future studies.
- MeSH
- Cell Membrane metabolism MeSH
- Cell Nucleus metabolism MeSH
- Zebrafish physiology MeSH
- Erythrocytes metabolism MeSH
- Myocytes, Cardiac metabolism MeSH
- Caveolae metabolism MeSH
- Melanins metabolism MeSH
- Mitochondria metabolism MeSH
- Heart Ventricles cytology MeSH
- Heart Atria cytology MeSH
- Microscopy, Electron, Transmission MeSH
- Calcium analysis MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Eggs of sterlet are discharged outside into ambient aquatic environment where egg activation and fertilization occur. Effects of different activation media including freshwater and clay suspension on protein abundances of egg were quantified in sterlet Acipenser ruthenus. In-gel digestion and high resolution mass spectrometry were used for label-free protein quantification in the eggs of five females. No significant (p > 0.05) difference was found between protein abundances in eggs activated with different media. However, results showed significant (p < 0.05, fold change ≥2) reduction in the abundances of nine proteins including six glycoproteins, enolase and heat shock protein in activated groups compared to freshly ovulated eggs as control. The fact that abundance of proteasome subunit alpha significantly reduced only in eggs which were activated by clay suspension suggests that activation medium can somehow intervene with protein regulation during fertilization. In conclusion, external fertilization in sturgeon egg is accompanied by huge release of proteins into the external environment that may participate in the construction of a transient microenvironment around egg for attraction and protection of spermatozoa to ensure ensuing fertilization. Data are available via ProteomeXchange with identifier PXD006232.
- MeSH
- Fertilization * MeSH
- Phosphopyruvate Hydratase metabolism MeSH
- Glycoproteins metabolism MeSH
- Ovum metabolism MeSH
- Heat-Shock Proteins metabolism MeSH
- Proteome metabolism MeSH
- Fishes metabolism MeSH
- Animals MeSH
- Check Tag
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Calcium plays a variety of vital regulatory functions in many physiological and biochemical events in the cell. The aim of this study was to describe the ultrastructural distribution of calcium during different developmental stages of spermatogenesis in a model organism, the zebrafish (Danio rerio), using a combined oxalate-pyroantimonate technique. Samples were treated by potassium oxalate and potassium pyroantimonate during two fixation stages and examined using transmission electron microscopy to detect electron dense intracellular calcium. The subcellular distribution of intracellular calcium was characterized in spermatogonium, spermatocyte, spermatid, and spermatozoon stages. The area which is covered by intracellular calcium in different stages was quantified and compared using software. Isolated calcium deposits were mainly detectable in the cytoplasm and the nucleus of the spermatogonium and spermatocyte. In the spermatid, calcium was partially localized in the cytoplasm as isolated deposits. However, most calcium was transformed from isolated deposits into an unbound pool (free calcium) within the nucleus of the spermatid and the spermatozoon. Interestingly, in the spermatozoon, calcium was mainly localized in a form of an unbound pool which was detectable as an electron-dense mass within the nucleus. Also, sporadic calcium deposits were scattered in the midpiece and flagellum. The proportional area which was covered by intracellular calcium increased significantly from early to late stages of spermatogenesis. The extent of the area which was covered by intracellular calcium in the spermatozoon was the highest compared to earlier stages. Calcium deposits were also observed in the somatic cells (Sertoli, myoid, Leydig) of zebrafish testis. The notable changes in the distribution of intracellular calcium of germ cells during different developmental stages of zebrafish spermatogenesis suggest its different homeostasis and physiological functions during the process of male gamete development.
- MeSH
- Cell Nucleus ultrastructure MeSH
- Zebrafish metabolism MeSH
- Spermatids cytology ultrastructure MeSH
- Spermatogenesis * MeSH
- Subcellular Fractions metabolism ultrastructure MeSH
- Testis cytology ultrastructure MeSH
- Calcium metabolism MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Calcium regulates many intracellular events such as growth and differentiation during different stages of gamete development. The aim of this study was to localize and quantify the intracellular distribution of calcium during different developmental stages of spermatogenesis in sterlet, Acipenser ruthenus, using a combined oxalate-pyroantimonate technique. The distribution of calcium was described in spermatogonium, spermatocyte, spermatid, and spermatozoon stages. In the spermatogonium and spermatocyte, calcium deposits were mainly localized in the nucleus and cytoplasm. The spermatid had calcium in the nucleus, developing acrosomal vesicle, and cytoplasm. Intracellular calcium transformed from scattered deposits in spermatogonia and spermatocyte stages into an unbound form in spermatid and the spermatozoon. The proportion of area covered by calcium increased significantly (p<0.05) from early to late stages of spermatogenesis. The largest proportion of area covered by calcium was observed in the nucleus of the spermatozoon. In conclusion, although most of the intracellular calcium is deposited in limited areas of the spermatogonium and spermatocyte, it is present an unbound form in the larger area of spermatids and spermatozoa which probably reflects changes in its physiological function and homeostasis during the process of male gamete production in spermatogenesis.
- MeSH
- Fishes anatomy & histology metabolism physiology MeSH
- Spermatids ultrastructure MeSH
- Spermatogenesis physiology MeSH
- Spermatogonia ultrastructure MeSH
- Spermatozoa ultrastructure MeSH
- Calcium metabolism MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Calcium plays prominent roles in regulating a broad range of physiological events in reproduction. The aim of this study was to describe the subcellular distribution of calcium deposits during stages of oogenesis in zebrafish using a combined oxalate-pyroantimonate technique. The oocyte development of zebrafish was categorized into four stages: primary growth, cortical-alveolus, vitellogenic, and maturation, based on morphological criteria. Calcium deposits in the primary growth stage were detected in the cytoplasm, mitochondria, nucleus, and follicular cells. At the cortical-alveolus stage, calcium particles were transported from follicular cells and deposited in the cortical alveoli. In the vitellogenic stage, some cortical alveoli were compacted and transformed from flocculent electron-lucent to electron-dense objects with the progression of the stage. Calcium deposits were transformed from larger to smaller particles, coinciding with compaction of cortical alveoli. In the maturation stage, calcium deposits in all oocyte compartments decreased, with the exception of those in mitochondria. The proportion of area covered by calcium deposits in the mitochondria and cortical alveoli of oocytes at different stages of development was significantly different (p<0.05). The extent of calcium deposits in the cortical alveoli of mature oocytes was substantially lower than in earlier stages. Basic information about calcium distribution during zebrafish oogenesis may contribute to better understanding of its role in oogenesis.
- MeSH
- Zebrafish physiology MeSH
- Microscopy methods MeSH
- Oocytes chemistry MeSH
- Oogenesis * MeSH
- Image Processing, Computer-Assisted methods MeSH
- Calcium analysis MeSH
- Animals MeSH
- Check Tag
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
After mating, spermatophores of signal crayfish are stored on the body of the female for a period before fertilization. This study compared the post-mating protein profile and pattern of protein tyrosine phosphorylation of the signal crayfish spermatophore to that of the freshly ejaculated spermatophore and found substantial differences. Two major bands of tyrosine-phosphorylated proteins of molecular weights 10 and 50kDa were observed in the freshly ejaculated spermatophore of the signal crayfish. While the tyrosine-phosphorylated protein band with molecular weight 10kDa was formed by protein(s) of similar pH, the band with molecular weight of 50kDa consisted of proteins of varying pH. In the post-mating spermatophore, the band with molecular weight of 50kDa was not detected, and an increase in the level of protein tyrosine phosphorylation was observed in the 10kDa band. The microtubular radial arms of the spermatozoon showed a positive reaction to an anti-tyrosine antibody conjugated with gold particles in both the freshly ejaculated and post-mating spermatophores. In conclusion, the male gamete of the signal crayfish undergoes molecular modification during post-mating storage on the body of the female including changes in the level of protein expression and protein tyrosine phosphorylation. Structural similarity of the radial arms in the crayfish immotile spermatozoon with flagellum, which is the main site of protein tyrosine phosphorylation in the mammalian motile spermatozoa, raises questions regarding evolution and function of such organelles across the animal kingdom that must be addressed in the future studies.
- MeSH
- Phosphorylation physiology MeSH
- Copulation MeSH
- Sperm Motility physiology MeSH
- Proteins genetics metabolism MeSH
- Astacoidea physiology MeSH
- Spermatogonia physiology MeSH
- Spermatozoa physiology MeSH
- Tyrosine metabolism MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
The freshly ejaculated spermatophore of crayfish undergoes a hardening process during post-mating storage on the body surface of female. The ultrastructural distribution of calcium deposits were studied and compared in freshly ejaculated and post-mating noble crayfish spermatophores, using the oxalate-pyroantimonate technique, to determine possible roles of calcium in post-mating spermatophore hardening and spermatozoon maturation. Small particles of sparsely distributed calcium deposits were visible in the wall of freshly ejaculated spermatophore. Also, large amount of calcium deposits were visible in the membranes of the freshly ejaculated spermatozoon. Five minutes post-ejaculation, granules in the spermatophore wall appeared as porous formations with numerous electron lucent spaces. Calcium deposits were visible within the spaces and scattered in the spermatophore wall matrix, where smaller calcium deposits combined to form globular calcium deposits. Large numbers of the globular calcium deposits were visible in the wall of the post-mating spermatophore. Smaller calcium deposits were detected in the central area of post-mating spermatophore, which contains the sperm mass, and in the extracellular matrix and capsule. While the density of calcium deposits decreased in the post-mating spermatozoon membranes, numerous small calcium deposits appeared in the subacrosomal zone and nucleus. Substantial changes in calcium deposit distribution in the crayfish spermatophore during post-mating storage on the body of female may be involved in the processes of the spermatophore hardening and spermatozoon maturation.
- MeSH
- Cell Nucleus ultrastructure MeSH
- Calcification, Physiologic physiology MeSH
- Reproduction MeSH
- Astacoidea cytology metabolism MeSH
- Spermatogonia cytology metabolism MeSH
- Calcium metabolism MeSH
- Animals MeSH
- Check Tag
- Male MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
