129 results on '"Elisabetta Tosti"'
Search Results
52. Regulatory roles of nitric oxide during larval development and metamorphosis in Ciona intestinalis
- Author
-
Elisabetta Tosti, Stefania Comes, Margherita Branno, Marco d'Ischia, Annamaria Locascio, Francesco Silvestre, Anna Palumbo, Gian Luigi Russo, S, Come, A, Locascio, F, Silvestre, D'Ischia, M., Russo, G. L., E, Tosti, M, Branno, A, Palumbo, Comes, S, Locascio, A, Silvestre, F, D'Ischia, Marco, Tosti, E, Branno, M, and Palumbo, A.
- Subjects
DNA, Complementary ,media_common.quotation_subject ,Molecular Sequence Data ,Apoptosis ,Biology ,Nitric Oxide ,Nitric oxide ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Juvenile ,Animals ,Ciona intestinalis ,Amino Acid Sequence ,Metamorphosis ,Cyclic GMP ,Molecular Biology ,In Situ Hybridization ,030304 developmental biology ,media_common ,0303 health sciences ,Larva ,Caspase 3 ,Reverse Transcriptase Polymerase Chain Reaction ,Metamorphosis, Biological ,Gene Expression Regulation, Developmental ,Anatomy ,Cell Biology ,biology.organism_classification ,Tadpole ,Resorption ,Cell biology ,Arginase ,chemistry ,Caspases ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Metamorphosis in the ascidian Ciona intestinalis is a very complex process which converts a swimming tadpole to an adult. The process involves reorganisation of the body plan and a remarkable regression of the tail, which is controlled by caspase-dependent apoptosis. However, the endogenous signals triggering apoptosis and metamorphosis are little explored. Herein, we report evidence that nitric oxide (NO) regulates tail regression in a dose-dependent manner, acting on caspase-dependent apoptosis. An increase or decrease of NO levels resulted in a delay or acceleration of tail resorption, without affecting subsequent juvenile development. A similar hastening effect was induced by suppression of cGMP-dependent NO signalling. Inhibition of NO production resulted in an increase in caspase-3-like activity with respect to untreated larvae. Detection of endogenously activated caspase-3 and NO revealed the existence of a spatial correlation between the diminution of the NO signal and caspase-3 activation during the last phases of tail regression. Real-time PCR during development, from early larva to early juveniles, showed that during all stages examined, NO synthase (NOS) is always more expressed than arginase and it reaches the maximum value at late larva, the stage immediately preceding tail resorption. The spatial expression pattern of NOS is very dynamic, moving rapidly along the body in very few hours, from the anterior part of the trunk to central nervous system (CNS), tail and new forming juvenile digestive organs. NO detection revealed free diffusion from the production sites to other cellular districts. Overall, the results of this study provide a new important link between NO signalling and apoptosis during metamorphosis in C. intestinalis and hint at novel roles for the NO signalling system in other developmental and metamorphosis-related events preceding and following tail resorption.(c) 2007 Elsevier Inc. All rights reserved.
- Published
- 2007
- Full Text
- View/download PDF
53. The Impact of Inflammatory Bowel Disease (IBD) and its Treatment on the Reproductive Process
- Author
-
Anna Maria Anniciello and Elisabetta Tosti
- Subjects
medicine.medical_specialty ,Crohn's disease ,business.industry ,media_common.quotation_subject ,Obstetrics and Gynecology ,medicine.disease ,Inflammatory bowel disease ,Ulcerative colitis ,Gastroenterology ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,Reproductive process ,medicine ,030212 general & internal medicine ,Reproduction ,business ,030217 neurology & neurosurgery ,media_common - Published
- 2007
54. The Ascidian Ciona Intestinalis as Model Organism for Ecotoxicological Bioassays
- Author
-
ra Gallo, Elisabetta Tosti, and Aless
- Subjects
Pollution ,Pollutant ,biology ,Ecology ,ved/biology ,media_common.quotation_subject ,ved/biology.organism_classification_rank.species ,Wildlife ,Biodiversity ,Global change ,biology.organism_classification ,Bioassay ,Ciona intestinalis ,Model organism ,media_common - Abstract
The increase of pollutants in the oceans as well as the global change around the world is having a negative impact on the biodiversity of living organisms. There is a crescent alarm in the scientific community for the different types of pollution (biological, chemical and physical) and for the changes occurring at the level of species and communities. Many parameters endangering marine wildlife contribute to create an unhealthy marine environment that induces reproductive disorders that, in turn, may influence even the survival of marine wildlife.
- Published
- 2015
55. Sperm Vacuoles and Reproductive Outcome
- Author
-
Elisabetta Tosti and Adriana Fortunato
- Subjects
medicine.medical_specialty ,Reproductive medicine ,medicine ,Vacuole ,Biology ,Bioinformatics ,Omics ,Outcome (game theory) ,Sperm - Published
- 2015
56. Cytoskeletal Elements and the Reproductive Success in Animals
- Author
-
Alessandra Gallo and Elisabetta Tosti
- Subjects
0303 health sciences ,030219 obstetrics & reproductive medicine ,Zygote ,media_common.quotation_subject ,Zoology ,Biology ,Oocyte ,Sperm ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,Human fertilization ,Meiosis ,medicine ,Reproduction ,Cytoskeleton ,Gametogenesis ,030304 developmental biology ,media_common - Abstract
Reproduction is a stepwise process that starts at the formation of gametes and ends at the completion of meiosis and the first mitotic divisions of the zygote. The involvement of main cytoskeletal elements in mediating the reproductive processes is now well established. This chapter summarizes recent advances in the field related to cytoskeletal dynamics underpinning the basic events of reproduction such as gametogenesis, gametes reciprocal activation, sperm-oocyte interaction and zygote formation. We have described the distinct patterns of distribution of microfilaments and microtubules and their rearrangements along the entire reproductive process. Evidences are provided that these elements play a dynamic role in the establishment and regulation of basic structure and functions of gametes and zygote during the processes of maturation and fertilization.
- Published
- 2015
57. The Impact of In Vitro Fertilization on the Health of the Mother and the Offspring
- Author
-
Alessandro Settimi, Elisabetta Tosti, and Adriana Fortunato
- Subjects
Andrology ,In vitro fertilisation ,business.industry ,Offspring ,medicine.medical_treatment ,Obstetrics and Gynecology ,Medicine ,business - Published
- 2006
58. Interventions to Improve the Implementation of Evidence Based Practices in Women Care
- Author
-
Alessandro Settimi, Fernando Althabe, María Belizán, Mercedes Colomar, Elisabetta Tosti, and Adriana Fortunato
- Subjects
Intervention (law) ,Evidence-based practice ,Nursing ,business.industry ,Unnecessary Procedure ,Psychological intervention ,Birth attendant ,Obstetrics and Gynecology ,Medicine ,business ,Research findings ,Scientific evidence ,Obstetric care - Abstract
An important component of quality of care is the use of evidence-base d practices. Three studies from three different continents (Africa, Asia and Latin America) have reported the suboptimal use of evidence - based interventions in obstetric care, proving that despite scientific evidence and dissemination efforts, there are harmful and/or unnecessary procedures still used, while others that are beneficial are ignored. This illustrates the existing bridges and barriers between practice guidelines based on research findings and practitioners. A theoretical framework could help explain these barriers and possibly help target interventions to specific barriers. A review of existing literature shows that an intervention aimed to increase the use of evidence-base d practices should have the following components: increase birth attendant concern about the effectiveness of routine clinical practices, stimulate their desire to review the effectiveness of their practice, provide them with skills to perform evidence based clinical guidelines appraisal and development and establish mechanisms through key hospital leaders to implement the guidelines and sustain them over time.
- Published
- 2006
59. Ca2+ and Na+ current patterns during oocyte maturation, fertilization, and early developmental stages ofCiona intestinalis
- Author
-
Rosaria De Santis, Annunziata Cuomo, Francesco Silvestre, and Elisabetta Tosti
- Subjects
Time Factors ,Voltage clamp ,Fertilization in Vitro ,Sodium Channels ,Membrane Potentials ,Oogenesis ,Human fertilization ,Botany ,Genetics ,medicine ,Animals ,Ciona intestinalis ,Metaphase ,Fertilisation ,Zygote ,Germinal vesicle ,biology ,Sodium ,Cell Differentiation ,Cell Biology ,Oocyte ,biology.organism_classification ,Cell biology ,medicine.anatomical_structure ,Fertilization ,Oocytes ,Calcium ,Calcium Channels ,Ion Channel Gating ,Developmental Biology - Abstract
Using the whole-cell voltage clamp technique, the electrical changes in oocyte and embryo plasma membrane were followed during different meiotic and developmental stages in Ciona intestinalis. We show, for the first time, an electrophysiological characterization of the plasma membrane in oocytes at the germinal vesicle (GV) stage with high L-type calcium (Ca2+) current activity that decreased through meiosis. Moreover, the absence of Ca2+ reduced germinal vesicle breakdown (GVBD), which is consistent with a role of Ca2+ currents in the prophase/metaphase transition. In mature oocytes at the metaphase I (MI) stage, Ca2+ currents decreased and then disappeared and sodium (Na+) currents first appeared remaining high up to the zygote stage. Intracellular Ca2+ release was higher in MI than in GV, indicating that Ca2+ currents in GV may contribute to fill the stores which are essential for oocyte contraction at fertilization. The fertilization current generated in Na+ free sea water was significantly lower than the control; furthermore, oocytes fertilized in the absence of Na+ showed high development of anomalous "rosette" embryos. Current amplitudes became negligible in embryos at the 2- and 4-cell stage, suggesting that signaling pathways that mediate first cleavage do not rely on ion current activities. At the 8-cell stage embryo, a resumption of Na+ current activity and conductance occurred, without a correlation with specific blastomeres. Taken together, these results imply: (i) an involvement of L-type Ca2+ currents in meiotic progression from the GV to MI stage; (ii) a role of Na+ currents during electrical events at fertilization and subsequent development; (iii) a major role of plasma membrane permeability and a minor function of specific currents during initial cell line segregation events.
- Published
- 2006
60. Calcium currents correlate with oocyte maturation during the reproductive cycle inOctopus vulgaris
- Author
-
Elisabetta Tosti, Carlo Di Cristo, Anna Di Cosmo, Marina Paolucci, and Annunziata Cuomo
- Subjects
medicine.medical_specialty ,Patch-Clamp Techniques ,Voltage clamp ,Octopodiformes ,Biology ,Prophase ,Meiosis ,Internal medicine ,medicine ,Animals ,Metaphase ,Progesterone ,Germinal vesicle ,Spermatozoon ,Vitellogenesis ,Oocyte ,Cell biology ,Endocrinology ,medicine.anatomical_structure ,Oocytes ,Calcium ,Female ,Animal Science and Zoology ,Calcium Channels - Abstract
Using the whole-cell voltage clamp technique, we have studied the Ca2+ currents and the steady-state conductance during different oocyte growth stages and during the reproductive cycle of the female of Octopus vulgaris. Evidence is presented that L-type Ca2+ currents are high in small pre-vitellogenic oocytes (80-150 microm diameter) and significantly lower in early vitellogenic oocytes (180-300 microm diameter). Similarly, a significant decrease of the steady-state conductance occurred from the pre to early- vitellogenic oocytes. Octopus oocytes showed larger Ca2+ currents in the reproductive rather than non-reproductive periods. These data indicates that ion and L-type Ca2+ currents play a role in oocyte growth and cytoplasmic maturation, and possibly in preparing the plasma membrane to the interaction with the spermatozoon. By using fluorescent microscopy, we show that oocytes from 80 to 400 microm diameter have the large germinal vesicle characteristic of the immature oocytes. In subsequent stages of growth (up to 1000 microm diameter) the nucleus is no more visible and the metaphase spindle appears. These data demonstrate that Octopus vulgaris oocytes are arrested in the first meiotic prophase up to the early-vitellogenic stage and resume meiosis at this stage up to a second block presumably in metaphase I. We discuss a possible role for progesterone as the hormonal stimulus for the first prophase-metaphase meiotic transition.
- Published
- 2005
61. Biochemical and Functional Characterization of Protein Kinase CK2 in Ascidian Ciona intestinalis Oocytes at Fertilization
- Author
-
Elisabetta Tosti, Annalisa Mupo, Mariarosaria Tosto, Immacolata Castellano, Gian Luigi Russo, and Annunziata Cuomo
- Subjects
0303 health sciences ,Cyclin-dependent kinase 1 ,animal structures ,Protein subunit ,fungi ,030302 biochemistry & molecular biology ,Oocyte activation ,Cell Biology ,Biology ,Oocyte ,biology.organism_classification ,Biochemistry ,Molecular biology ,Cell biology ,03 medical and health sciences ,medicine.anatomical_structure ,embryonic structures ,medicine ,Ciona intestinalis ,Casein kinase 2 ,Protein kinase A ,Molecular Biology ,Metaphase ,030304 developmental biology - Abstract
The ubiquitous and pleiotropic dual specificity protein kinase CK2 has been studied and characterized in many organisms, from yeast to mammals. Generally, the enzyme is composed of two catalytic (α and/or α′) and two regulatory (β) subunits, forming a differently assembled tetramer. Although prone to controversial interpretation, the function of CK2 has been associated with fundamental biological processes such as signal transduction, cell cycle progression, cell growth, apoptosis, and transcription. Less known is the role of CK2 during meiosis and the early phase of embryogenesis. In this work, we studied CK2 activity during oocyte activation, a process occurring at the end of oocyte maturation and triggered by fertilization. In ascidian Ciona intestinalis, an organism whose complete genome has been published recently, CK2 was constitutively active in unfertilized and fertilized oocytes. The enzymatic activity oscillated through meiosis showing three major peaks: soon after fertilization (metaphase I exit), before metaphase II, and at the exit from metaphase II. Biochemical analysis of CK2 subunit composition in activated oocytes indicated that CK2-α was catalytically active as a monomer, independently from its regulatory subunit β; however, CK2-β was only detectable in unfertilized oocytes where it was associated with a bona fide identified ascidian mitogen-activated protein kinase. After fertilization, CK2-β was undetectable, suggesting its rapid degradation. Protein sequence analysis of CK2-α and -β cDNA indicated a high identity compared with vertebrate homologs. In addition, the absence of putative phosphorylation sites for Cdc2 kinase on both α and β subunits suggested an important role for CK2 in regulating meiotic cell cycle in C. intestinalis oocytes.
- Published
- 2004
62. Electrical events during gamete maturation and fertilization in animals and humans
- Author
-
Raffaele Boni and Elisabetta Tosti
- Subjects
Male ,Spermatozoon ,Obstetrics and Gynecology ,Biology ,Oocyte ,Spermatozoa ,Resting potential ,Ion Channels ,Calcium in biology ,Cell biology ,Electrophysiology ,Human fertilization ,medicine.anatomical_structure ,Reproductive Medicine ,Fertilization ,Second messenger system ,Botany ,Oocytes ,medicine ,Animals ,Humans ,Gamete ,Calcium ,Female ,Ion channel - Abstract
Gamete cells are electrogenic, i.e. capable of responding to electrical stimuli and modifying their electrical properties during the crucial periods of maturation and fertilization. Ion channels have been widely demonstrated on the plasma membrane of the oocyte and spermatozoon in all animals studied, and electrical modifications in gametes are due to ion currents that are modulated via these ion channels. The modification of intracellular calcium levels in gametes has been extensively studied, and these modifications are recognized to be a second messenger system for gamete maturation and fertilization. Other ions also move through the plasma membrane, either in association with or independent of calcium, and these generate typical features such as fertilization currents and oscillation of resting potential. These modifications were first studied in marine invertebrates, and the observations subsequently compared with mammalian systems, including human. The precise role played by these currents in the processes of maturation and fertilization is still poorly understood; however, recent research opens new frontiers for their clinical and technological application.
- Published
- 2004
63. Bioactive aldehydes from diatoms block the fertilization current in ascidian oocytes
- Author
-
Adrianna Ianora, Antonio Miralto, Elisabetta Tosti, Isabella Buttino, Giovanna Romano, and Annunziata Cuomo
- Subjects
Patch-Clamp Techniques ,Time Factors ,Gating ,Biology ,Human fertilization ,Botany ,Genetics ,medicine ,Animals ,Ciona intestinalis ,Channel blocker ,Actin ,Diatoms ,Aldehydes ,Microscopy, Confocal ,Embryo ,Cell Biology ,Blastomere ,biology.organism_classification ,Cell biology ,Mechanism of action ,Fertilization ,Larva ,Oocytes ,Female ,medicine.symptom ,Developmental Biology - Abstract
The effects of bioactive aldehydes from diatoms, unicellular algae at the base of the marine food web, were studied on fertilization and early development processes of the ascidian Ciona intestinalis. Using whole-cell voltage clamp techniques, we show that 2-trans-4-trans-decadienal (DD) and 2-trans-4-cis-7-cis-decatrienal (DT) inhibited the fertilization current which is generated in oocytes upon interaction with the spermatozoon. This inhibition was dose-dependent and was accompanied by inhibition of the voltage-gated calcium current activity of the plasma membrane. DD and DT did not inhibit the subsequent contraction of the cortex. Moreover, DD specifically acted as a fertilization channel inhibitor since it did not affect the steady state conductance of the plasma membrane or gap junctional (GJ) communication within blastomeres of the embryo. On the other hand, DD did affect actin reorganization even though the mechanism of action on actin filaments differed from that of other actin blockers. Possibly this effect on actin reorganization was responsible for the subsequent teratogenic action on larval development. The effect of DD was reversible if oocytes were washed soon after fertilization indicating that DD may specifically target certain fertilization mechanisms. Thus, diatom reactive aldehydes such as DD may have a dual effect on reproductive processes, influencing primary fertilization events such as gating of fertilization channels and secondary processes such as actin reorganization which is responsible for the segregation of cell lineages. These findings add to a growing body of evidence on the antiproliferative effects of diatom-derived aldehydes. Our results also report, for the first time, on the action of a fertilization channel blocker in marine invertebrates.
- Published
- 2003
64. Antifouling Compounds Endangering Marine Invertebrates Reproduction
- Author
-
Aless, ra Gallo, and Elisabetta Tosti
- Subjects
Biofouling ,chemistry.chemical_compound ,chemistry ,Ecology ,Reproduction (economics) ,Marine invertebrates ,Biology ,Xenobiotic - Abstract
Reproduction and embryo development of marine animals occur in the sea water and are processes very sensitive to substances present in the environment that come from industrial and agricultural activities. These compounds are collectively known as xenobiotics due to their extraenvironmental origin. Among xenobiotics present in the sea water, a relevant role is played by the anti-fouling compounds. These are used to prevent settlement and growth of organisms on submerged structures of vessels and ships that otherwise should create serious problems of speed slow down.
- Published
- 2014
65. Progesterone induces activation inOctopus vulgaris spermatozoa
- Author
-
Elisabetta Tosti, Anna Di Cosmo, Giovanni Gragnaniello, Carlo Di Cristo, and Annunziata Cuomo
- Subjects
Male ,endocrine system ,medicine.medical_specialty ,Octopodiformes ,Acrosome reaction ,Ionophore ,chemistry.chemical_element ,Stimulation ,Calcium ,Biology ,Andrology ,Lectins ,Internal medicine ,Genetics ,medicine ,Animals ,Acrosome ,Calcimycin ,Progesterone ,reproductive and urinary physiology ,Fluorescent Dyes ,Ionophores ,urogenital system ,Acrosome Reaction ,Lectin ,Cell Biology ,Spermatozoa ,Membrane progesterone receptor ,Endocrinology ,chemistry ,Spermatophore ,biology.protein ,Female ,Fluorescein-5-isothiocyanate ,Developmental Biology - Abstract
The purpose of the present study was to determine whether Octopus vulgaris spermatozoa are activated by progesterone stimulation. Spermatozoa were collected from the spermatophores in the Needham's sac of the male lMSr and from the spermathecae of oviducal glands of the female lFSr. We used transmission lTEMr and scanning lSEMr electron microscopy to study the morphology of untreated, Ca2p ionophore A23187 and progesteronehtreated MS spermatozoa, and untreated FS spermatozoa. We showed that ionophore and progesterone stimulation of MS spermatozoa induce breakdown of the membranes overlapping the acrosomal region, exposing the spiralized acrosome. These modifications resemble the acrosome reaction observed in other species. FS stored in the spermathecae did not show the membranes covering the acrosomal region present in the MS spermatozoa. When ionophore and progesterone treatments were performed in Ca2phfree artificial sea water, no changes were observed, suggesting the role of external calcium in modifying membrane morphology. Lectin studies showed a different fluorescence distribution and membrane arrangement of FShuntreated spermatozoa with respect to the MS, suggesting that spermatozoa transferred in the female genital tract after mating, are stored in a prehactivated state. The plasma membrane of the untreated MS and FS spermatozoa was labelled with ProgesteronehBSAhFITC, indicating the presence of plasma membrane progesterone receptor. Taken together these data suggest that progesterone induces an acrosomeh like reaction in MS spermatozoa similar to that induced by calcium elevation. In addition progesterone may play a role in the prehactivation of spermatozoa stored in the female tract, further supporting the hypothesized parallelism between cephalopods and vertebrates. Mol. Reprod. Dev. 59:97–105, 2001. © 2001 WileyhLiss, Inc.
- Published
- 2001
66. Functional Gap Junctions in the Early Sea Urchin Embryo Are Localized to the Vegetal Pole
- Author
-
Elisabetta Tosti, Brian Dale, and Ikuko Yazaki
- Subjects
Polarity in embryogenesis ,blastomeres ,Biology ,Cleavage (embryo) ,Morula ,sea urchin ,chemistry.chemical_compound ,Animals ,Molecular Biology ,gap junctions ,Body Patterning ,Lucifer yellow ,Gap junction ,Electric Conductivity ,Blastomere ,Anatomy ,Gastrula ,Cell Biology ,1-Octanol ,Embryonic Induction ,Coupling (electronics) ,chemistry ,Sea Urchins ,Biophysics ,intercellular communication ,Archenteron ,Developmental Biology - Abstract
Using the whole-cell voltage-clamp technique we have studied electrical coupling and dye coupling between pairs of blastomeres in 16- to 128-cell-stage sea urchin embryos. Electrical coupling was established between macromeres and micromeres at the 16-cell stage with a junctional conductance ( G j ) of 26 nS that decreased to 12 nS before the next cleavage division. G j between descendants of macromeres and micromeres was 12 nS falling to 8 nS in the latter half of the cell cycle. Intercellular current intensity was independent of transjunctional voltage, nondirectional, and sensitive to 1-octanol and therefore appears to be gated through gap junction channels. There was no significant coupling between other pairs of blastomeres. Lucifer yellow did not spread between these electrically coupled cell pairs and in fact significant dye coupling between nonsister cells was observed only at the 128-cell stage. Since 1-octanol inhibited electrical communication between blastomeres at the 16- to 64-cell stage and also induced defects in formation of the archenteron, it is possible that gap junctions play a role in embryonic induction.
- Published
- 1999
- Full Text
- View/download PDF
67. Nitric Oxide Gates Fertilization Channels in Ascidian Oocytes through Nicotinamide Nucleotide Metabolism
- Author
-
Elisabetta Tosti, Brian Dale, Lucia Grumetto, Antony Galione, Martin Wilding, M. L. De Simone, Grumetto, Lucia, Wilding, M., De Simone, M. L., Tosti, E., Galione, A., and Dale, B.
- Subjects
Intracellular Fluid ,Male ,Niacinamide ,Nitroprusside ,Biophysics ,chemistry.chemical_element ,Calcium ,Nitric Oxide ,Biochemistry ,Cyclic ADP-ribose ,Ion Channels ,Calcium in biology ,Nitric oxide ,chemistry.chemical_compound ,medicine ,Animals ,Ciona intestinalis ,Molecular Biology ,ascidia oocytes ,Adenosine Diphosphate Ribose ,Cyclic ADP-Ribose ,biology ,Nicotinamide ,Cell Biology ,biology.organism_classification ,Spermatozoa ,chemistry ,nicotinamide metabolism ,Fertilization ,Second messenger system ,Oocytes ,Sodium nitroprusside ,Ion Channel Gating ,medicine.drug - Abstract
In this paper we use the nitric oxide (NO) donor sodium nitroprusside to examine the response of the unfertilised oocyte of the ascidian Ciona intestinalis to nitric oxide. We show that the release of NO triggers an inward current that displays similar properties to the ascidian fertilisation current. Furthermore, the production of NO causes the release of intracellular calcium through a ruthenium-red sensitive mechanism. Our data suggest that these effects are due to the stimulation of nicotinamide nucleotide metabolism, but the active second messenger is not cyclic adenosine diphosphate ribose (cADPr). Finally, we show that NO production increases at fertilisation. The results suggest that ascidian sperm trigger the release of NO and this second messenger causes the breakdown of nicotinamide nucleotides leading to the production of a second messenger which induces the fertilisation current and may assist in the production of the increase in calcium.
- Published
- 1997
68. Polarized distribution of L-type calcium channels in early sea urchin embryos
- Author
-
Elisabetta Tosti, I. Yazaki, and Brian Dale
- Subjects
Blastomeres ,Embryo, Nonmammalian ,Potassium Channels ,Calcium Channels, L-Type ,Nifedipine ,Physiology ,chemistry.chemical_element ,Calcium ,Paracentrotus lividus ,biology.animal ,Animals ,L-type calcium channel ,Sea urchin ,Ion transporter ,Calcium metabolism ,Voltage-dependent calcium channel ,biology ,Chemistry ,Cell Polarity ,Gastrula ,Cell Biology ,Anatomy ,Blastomere ,Calcium Channel Blockers ,biology.organism_classification ,Sea Urchins ,Microscopy, Electron, Scanning ,Biophysics ,Calcium Channels ,Cell Division - Abstract
Using the whole cell clamp technique, we have measured calcium-dependent currents and steady-state conductance in early sea urchin blastomeres. The calcium currents in M phase decreased from 8.5 microA/cm2 at the four-cell stage to 5.4 microA/cm2 at the eight-cell stage. In 16-cell stage embryos, calcium currents were 7.4 microA/cm2 in the mesomeres, 2.3 microA/cm2 in the macromeres, and were not detected in the micromeres. In contrast, the micromeres had a two- to threefold higher steady-state conductance than the mesomeres or macromeres, which may be due to potassium ion conductivity. Nifedipine, an L-type channel antagonist, delays cleavage division at a concentration of 0.05-0.1 mM and causes developmental defects, such as poor skeletal differentiation in later sea urchin embryos.
- Published
- 1997
69. Distribution pattern and activity of mitochondria during oocyte growth and maturation in the ascidian Styela plicata
- Author
-
Francesco Silvestre, Alessandra Gallo, Saïda Tekaya, Elisabetta Tosti, and Amina Bezzaouia
- Subjects
Cytoplasm ,DiOC6 ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Human fertilization ,Microscopy, Electron, Transmission ,medicine ,Animals ,Urochordata ,030304 developmental biology ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Germinal vesicle ,biology ,Oocyte activation ,Cell Biology ,biology.organism_classification ,Oocyte ,Cell biology ,Mitochondria ,Styela plicata ,medicine.anatomical_structure ,chemistry ,Oocytes ,Female ,Vitellogenesis ,Developmental Biology - Abstract
SummaryThe process of oocyte maturation is underlined by a redistribution of cellular organelles, among which mitochondria play a functional role for the acquisition of fertilization and developmental competence. In this paper, we applied electron and confocal microscopy by using DIOC6and JC-1 stain to evaluate mitochondria distribution pattern and activity during different stages of oocyte growth in the ascidianStyela plicata. Three categories of oocytes at the germinal vesicle stage underlying the vitellogenic process were characterized on the basis of size, pigmentation and accessory cells. Mitochondria were spread throughout the cytoplasm at the smallest oocyte stage and gradually migrated to the periphery of the subcortical cytoplasm at the intermediate stage. At the fully grown oocyte stage, mitochondria were aggregated in the subcortical cytoplasm. This pattern of polarized mitochondria distribution correlates significantly with an increase in mitochondria potential and activity. In this paper we discuss the relationship of mitochondria to the acquisition of oocyte developmental competence.
- Published
- 2013
70. Sperm DNA Fragmentation Assays Correlate with Sperm Abnormal Morphology and the Pregnancy Outcome
- Author
-
Sofia Casale, Francesca Liguori, Giuseppina Nacchia, Elisabetta Tosti, Adriana Fortunato, and Rita Leo
- Subjects
endocrine system ,In vitro fertilisation ,urogenital system ,medicine.medical_treatment ,Biology ,Sperm ,Chromatin ,Andrology ,chemistry.chemical_compound ,Prophase ,chemistry ,Sperm chromatin decondensation ,medicine ,DNA fragmentation ,Fragmentation (cell biology) ,reproductive and urinary physiology ,DNA - Abstract
Purpose: A retrospective study of 89 hypo-fertile male patients attending for in vitro fertilization was undertaken in order to evaluate possible correlations among sperm DNA damage, sperm analysis parameters and pregnancies. Methods: Sperm parameters (concentration, normal morphology and multiple anomalies) were evaluated according to the World Health Organization guidelines. DNA damages were simultaneously evaluated on each sperm sample by i) sperm chromatin decondensation test; ii) sperm DNA fragmentation evaluated by Sperm Chromatin Dispersion and Halo sperm Kit; iii) sperm DNA fragmentation evaluated by the Terminal Uridine Nick-End Labelling procedure. Results: Statistical analysis was performed by using analysis of variance and least squares regression. The sperm chromatin dispersion and fragmentation assays showed a statistically significant positive correlation (P=0.0017) in all the samples confirming the good efficacy of either of the two tests in detecting sperm DNA damage. Both the two test negatively correlated with normal sperm morphology (P=0.008), however only by using Halo sperm test we obtained a significant correlation with multiple sperm pathological morphologies (P=0.029) and an inverse correlation with pregnancies outcome (P=0.013). No correlation was detected among DNA damages, sperm concentration and chromatin decondensation. Conclusions: These data suggest a similar efficacy of sperm chromatin dispersion and Terminal Uridine Nick-End Labelling in detecting sperm DNA integrity. Due to the higher sensitivity of Halo test, its prognostic role in the diagnosis of fertilizing ability of a semen sample and possible pregnancy rate is discussed
- Published
- 2013
71. T-type Ca2+ current activity during oocyte growth and maturation in the ascidian Styela plicata
- Author
-
Gian Luigi Russo, Alessandra Gallo, and Elisabetta Tosti
- Subjects
Embryology ,Anatomy and Physiology ,lcsh:Medicine ,Calcium Channels, T-Type ,Sperm-Egg Interactions ,0302 clinical medicine ,Molecular Cell Biology ,lcsh:Science ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Multidisciplinary ,Zygote ,biology ,Voltage-dependent calcium channel ,Hyperpolarization (biology) ,Cell biology ,Electrophysiology ,medicine.anatomical_structure ,Research Article ,Signal Transduction ,medicine.medical_specialty ,Animal Types ,Embryonic Development ,chemistry.chemical_element ,Calcium ,Signaling Pathways ,03 medical and health sciences ,Internal medicine ,Calcium-Mediated Signal Transduction ,medicine ,Animals ,Urochordata ,Biology ,030304 developmental biology ,Germinal vesicle ,Cell Membrane ,Embryogenesis ,lcsh:R ,Oocyte ,biology.organism_classification ,Styela plicata ,Endocrinology ,chemistry ,Fertilization ,Oocytes ,Veterinary Science ,lcsh:Q ,Organism Development ,Developmental Biology ,Aquatic Animals - Abstract
Voltage-dependent calcium currents play a fundamental role during oocyte maturation, mostly L-type calcium currents, whereas T-type calcium currents are involved in sperm physiology and cell growth. In this paper, using an electrophysiological and pharmacological approach, we demonstrated, for the first time in oocytes, that T-type calcium currents are present with functional consequences on the plasma membrane of growing immature oocytes of the ascidian Styela plicata. We classified three subtypes of immature oocytes at the germinal vesicle stage on the basis of their size, morphology and accessory cellular structures. These stages were clearly associated with an increased activity of T-type calcium currents and hyperpolarization of the plasma membrane. We also observed that T-type calcium currents oscillate in the post-fertilization embryonic stages, with minimal amplitude of the currents in the zygote and maximal at 8-cell stage. In addition, chemical inhibition of T-type calcium currents, obtained by applying specific antagonists, induced a significant reduction in the rate of cleavage and absence of larval formation. We suggest that calcium entry via T-type calcium channels may act as a potential pacemaker in regulating cytosolic calcium involved in fertilization and early developmental events.
- Published
- 2013
72. Maturation Promoting Factor in ascidian oocytes is regulated by different intracellular signals at meiosis I and II
- Author
-
Elisabetta Tosti, Gian Luigi Russo, Keiichiro Kyozuka, Brian Dale, and Livio Antonazzo
- Subjects
Maturation-Promoting Factor ,Molecular Sequence Data ,Maturation promoting factor ,chemistry.chemical_element ,Protein Serine-Threonine Kinases ,Biology ,Calcium ,Calcium in biology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,BAPTA ,Cyclins ,medicine ,Animals ,Amino Acid Sequence ,Telophase ,Kinase activity ,Egtazic Acid ,Molecular Biology ,Chelating Agents ,030304 developmental biology ,0303 health sciences ,Heparin ,Cell Membrane ,Oocyte ,Ciona intestinalis ,Enzyme Activation ,Meiosis ,medicine.anatomical_structure ,chemistry ,Biochemistry ,Oocytes ,Biophysics ,biology.protein ,030217 neurology & neurosurgery ,Signal Transduction ,Developmental Biology ,MPF complex - Abstract
Using the fluorescent dye Calcium Green-dextran, we measured intracellular Ca2+ in oocytes of the ascidian Ciona intestinalis at fertilization and during progression through meiosis. The relative fluorescence intensity increased shortly after insemination in a single transient, the activation peak, and this was followed by several smaller oscillations that lasted for approximately 5 minutes (phase 1). The first polar body was extruded after the completion of the phase 1 transients, about 9 minutes after insemination, and then the intracellular calcium level remained at baseline for a period of 5 minutes (phase 2). At 14 minutes postinsemination a second series of oscillations was initiated that lasted 11 minutes (phase 3) and terminated at the time of second polar body extrusion. Phases 1 and 3 were inhibited by preloading oocytes with 5 mM heparin. Simultaneous measurements of membrane currents, in the whole-cell clamp configuration, showed that the 1–2 nA inward fertilization current correlated temporally with the activation peak, while a series of smaller oscillations of 0.1–0.3 nA amplitude were generated at the time of the phase 3 oscillations. Biochemical characterization of Maturation Promoting Factor (MPF) in ascidian oocytes led to the identification of a Cdc2-like kinase activity. Using p13suc1-sepharose as a reagent to precipitate the MPF complex, a 67 kDa (67×103Mr) protein was identified as cyclin B. Histone H1 kinase activity was high at metaphase I and decreased within 5 minutes of insemination reaching a minimum level during phase 2, corresponding to telophase I. During phase 3, H1 kinase activity increased and then decayed again during telophase II. Oocytes preloaded with BAPTA and subsequently inseminated did not generate any calcium transients, nonetheless H1 kinase activity decreased 5 minutes after insemination, as in the controls, and remained low for at least 30 minutes. Injection of BAPTA during phase 2 suppressed the phase 3 calcium transients, and inhibited both the increase in H1 kinase activity normally encountered at metaphase II and second polar body extrusion.
- Published
- 1996
73. Reducing the time of sperm-oocyte interaction in human in-vitro fertilization improves the implantation rate
- Author
-
Brian Dale, Sergio Panzella, M. Cristina Magli, Elisabetta Tosti, Luca Gianaroli, Anna Pia Ferraretti, and A. Fiorentino
- Subjects
Adult ,Male ,endocrine system ,Time Factors ,medicine.medical_treatment ,Fertilization in Vitro ,Biology ,Andrology ,03 medical and health sciences ,0302 clinical medicine ,Human fertilization ,medicine ,Humans ,Embryo Implantation ,Prospective Studies ,030304 developmental biology ,Sperm-Ovum Interactions ,0303 health sciences ,030219 obstetrics & reproductive medicine ,In vitro fertilisation ,urogenital system ,Rehabilitation ,Obstetrics and Gynecology ,Embryo ,Oocyte ,Polyspermy ,Sperm ,Pregnancy rate ,medicine.anatomical_structure ,Reproductive Medicine ,Infertility ,Female - Abstract
Human oocyte development was evaluated after a reduced time exposure to spermatozoa in vitro. A total of 119 patients were assigned to two study groups in a randomized prospective study in which each patient's oocytes were exposed to spermatozoa for either 1 h (group 1 - 58 patients) or the standard 16 h incubation period (group 2 - 61 patients). The fertilization rate obtained in group 1 was higher than in group 2 (285/393, 73%, and 272/410, 66% respectively), suggesting that the spermatozoa-oocyte interaction occurs within 1 h. This was confirmed in a study in vitro using fluorescently labelled spermatozoa and normal oocyte-cumulus complexes. Spermatozoa enter the cumulus complex within 15 min, traverse the cumulus layer within 3 h, and first appear in the oocyte cortex at 4 h post-insemination. The incidence of polyspermy was higher in oocytes exposed to spermatozoa for 16 h (3%) than for 1 h (1%). There was no difference in the cleavage rate or morphological characteristics of embryos from both study groups. However, when evaluating the timing of embryo development, group 1 generated a significantly higher percentage of four to five cell embryos when compared to group 2 (55 versus 39%; P < 0.001), documented at 40 h post-insemination. The implantation and pregnancy rates for group 1 were 11 and 28%, while the corresponding rates for group 2 were 8 and 15%. This suggests that a reduced exposure of oocyte to spermatozoa favours embryo viability, possibly due to a decrease in potential damage from sperm metabolic waste products.
- Published
- 1996
74. Oocyte Maturation and Fertilization: A Long History for a Short Event
- Author
-
Elisabetta, Tosti, Raffaele, Boni, Elisabetta, Tosti, and Raffaele, Boni
- Subjects
- Fertilization (Biology), Ovum, Reproduction, Conception
- Abstract
'This book is an integrated approach to the study of the basic events involved reproduction, and contains recent achievements described by most of the outstanding scientists of this field. General and basic patterns of oocyte maturation and fertilization are described in a modern context of integrated morphological and biochemical methods, up to the practical application of this knowledge. Because of their typical external fertilization, simple marine invertebrate models (sea urchin, ascidians, etc.) initially provided relevant and unique sources of information on reproductive biology. A major impetus for transferring and comparing this information in mammals coincided with the development of protocols that also allowed external fertilization in mammals, first in laboratory animals, and then in livestock and human. In Vitro Fertilization (IVF) technology represents a revolution in the general knowledge of reproductive biology, opening new doors that lead to biomedical applications.'--Page ii.
- Published
- 2011
75. Is sperm chromatin packaging relevant for IVF success?
- Author
-
Elisabetta Tosti and Adriana Fortunato
- Subjects
Spermatid ,Spermatozoon ,urogenital system ,Biology ,Oocyte ,medicine.disease ,Protamine ,Sperm ,Chromatin ,Male infertility ,Andrology ,Semen quality ,medicine.anatomical_structure ,medicine ,biology.protein - Abstract
Spermiogenesis is the last phase of the process that give rise to a mature and competent spermatozoon. It occurs from a dramatic morphological and structural change of the spermatid, in particular replacement of DNA-linked histones by protamines leading to a highly compact chromatin structure consisting of DNA and heterogeneous nucleoproteins. The target of a fertilizing spermatozoon is to deliver into the oocyte the paternal genome and regulatory factors that are required for proper embryonic development [1]. To do this the sperm must be capable of undergoing decondensation at a peculiar moment of the fertilization process. Increasing evidence on the strong paternal effect on preimplantation embryo development [2] focus on the importance to identify a reliable sperm quality parameter. Although many different causes may give rise to male infertility [3], traditionally in IVF centres, routine laboratory investigations evaluate seminal parameters such as concentration, motility and morphology in order to assess semen quality prior to undergo assisted reproduction. Recent acquisitions correlate poor chromatin condensation to a failure in fertilization, embryo development and repeated miscarriages indicating even that sperm DNA damage over 30% impedes natural pregnancy.
- Published
- 2012
76. IMSI, Useful, Useless or Harmful?
- Author
-
Y Menezo and Elisabetta Tosti
- Subjects
Gamete donation ,Reproduction (economics) ,Wish ,Business ,Human body ,Principle of legality ,Database transaction ,Object (philosophy) ,Human being ,health care economics and organizations ,humanities ,Law and economics - Abstract
Treatment with gametes donated by subjects other than those who wish to be parents is offered as part of assisted reproduction techniques in many countries. Ethical and legal concerns concerning gamete donation are discussed. Gamete donation constitutes a legal transaction, whose form is basically that of a contract, and in fact we are dealing here with a strictly regulated contractual object. The debate lies in considerations of legality and proprietorship. However, gametes cannot be compared with other parts of the human body. Their ability to originate another human being gives them a special status, which entails moral concerns. Nevertheless, and despite the fact that ethical evaluations may be a critical aspect, they cannot determine the legality of the act.
- Published
- 2012
77. Regulation of the fertilization current in ascidian oocytes by intracellular second messengers
- Author
-
Elisabetta Tosti and Brian Dale
- Subjects
medicine.medical_specialty ,Receptors, Cytoplasmic and Nuclear ,Biology ,Phosphatidylinositols ,Second Messenger Systems ,Calcium in biology ,Membrane Potentials ,03 medical and health sciences ,chemistry.chemical_compound ,Human fertilization ,Internal medicine ,Sulfhydryl reagent ,Genetics ,medicine ,Animals ,Inositol 1,4,5-Trisphosphate Receptors ,030304 developmental biology ,0303 health sciences ,Heparin ,Hydrolysis ,030302 biochemistry & molecular biology ,Cell Polarity ,Neomycin ,Cell Biology ,Oocyte ,Cell Compartmentation ,Ciona intestinalis ,Cell biology ,EGTA ,Endocrinology ,medicine.anatomical_structure ,chemistry ,Fertilization ,Second messenger system ,Oocytes ,Membrane channel ,Calcium ,Calcium Channels ,Intracellular ,Developmental Biology - Abstract
Neomycin, injected into ascidian oocytes to a final concentration of 10–50 mM, inhibits both the fertilization current and the surface contraction, showing that phosphoinositide hydrolysis is required for these early activation events. Sperm-activated fertilization currents are not inhibited in the presence of 100 μg/ml intracellular heparin, suggesting that these currents are not directly gated by InsP3. The sulfhydryl reagent thimerosal at 100 μM, in contrast, significantly increases the fertilization current presumably by sensitizing the channel receptor. Since heparin inhibits the surface contraction, InsP3 receptors are shown to play a role in the propagation of the activation response in ascidian oocyte. Depleting intracellular calcium stores by microinjecting 50 mM EGTA into oocytes does not activate fertilization channels; however, subsequent fertilization of these EGTA loaded oocytes leads to a significantly larger and faster fertilization current. Thus in contrast to somatic cells studied to date, second messenger operated plasma membrane channels in ascidian oocytes are not gated by calcium released from intracellular stores. © 1994 Wiley-Liss, Inc.
- Published
- 1994
78. Voltage clamp of the nuclear envelope
- Author
-
Brian Dale, Luigia Santella, Elisabetta Tosti, Louis J. DeFelice, and Keiichiro Kyozuka
- Subjects
Germinal vesicle ,General Immunology and Microbiology ,Chemistry ,Voltage clamp ,Analytical chemistry ,General Medicine ,Resting potential ,General Biochemistry, Genetics and Molecular Biology ,Membrane ,medicine.anatomical_structure ,Cytoplasm ,medicine ,Biophysics ,Patch clamp ,General Agricultural and Biological Sciences ,Nucleus ,Ion channel ,General Environmental Science - Abstract
By using conventional intracellular microelectrodes, we show that the starfish germinal vesicle in situ has a resting potential of +4 mV to +30 mV with respect to the cytoplasm. Patch-clamp electrodes reveal that the surface of the isolated nucleus contains ion channels of 120 pS single-channel conductance. Under whole-cell voltage-clamp conditions, the starfish germinal vesicle generates asymmetric time-variant currents that depend on the ionic environment. Fluorescent dyes in the whole-cell clamp electrode fill the nucleus, showing that the time-variant and the steady-state conductances are properties of the outer and inner nuclear membranes or something that spans them. Because scanning electron microscopy shows a density of nuclear pores in the range 60—110 per square micrometre, and the steady-state resistance in the range is 30—100 MΩ, it seems probable that the pores are not freely permeable to ions. Furthermore, fluorescent dyes of molecular weight up to 500 Da micro-injected either into the cytoplasm or the nucleus of intact oocytes accumulate in the nucleus. Our results show that the nucleus acts as an electrically isolated compartment that separates charged low molecular mass molecules including inorganic ions.
- Published
- 1994
79. Sperm nucleus decondensation, hyaluronic acid (HA) binding and oocyte activation capacity: different markers of sperm immaturity? Case reports
- Author
-
Moncef Benkhalifa, Elisabetta Tosti, Marc Cohen, Didier De la fontaine, Yves Menezo, Am Junca, and Beatriz Gonzalez Marti
- Subjects
Male ,Fertilization in Vitro ,Gamete Biology ,Genetics ,medicine ,Humans ,Epigenetics ,Sperm Injections, Intracytoplasmic ,Hyaluronic Acid ,Genetics (clinical) ,Cell Nucleus ,biology ,Obstetrics and Gynecology ,Oocyte activation ,General Medicine ,Oocyte ,Sperm ,Spermatozoa ,Cell biology ,Chromatin ,Histone ,medicine.anatomical_structure ,Reproductive Medicine ,biology.protein ,Oocytes ,DNA fragmentation ,Female ,Spermatogenesis ,Developmental Biology - Abstract
During the early time of IVF, sperm competence was defined as the ability to fertilize an oocyte. However, with the advent of ICSI, despite the capacity to reach 2–4cell stage, it is impossible for numerous patients, to establish a pregnancy. Instead the consensus now is that male fertility has to be defined as the capacity to produce sperm cells able to establish a full term pregnancy [1–3] Basic parameters such as concentration, motility and morphology are of limited value in determining sperm capacity to allow full embryonic development to term. Determination of DNA changes like fragmentation and condensation are independent parameters [2] and obviously of major importance. DNA fragmentation, related or not to reactive oxygen species (ROS) insult is only one piece of the problem. Indeed, sperm chromatin tertiary structure seems to be critical for correct epigenetic regulation and maintenance, and further on for male fertility [4, 5]. During very early embryogenesis, an adequate chromatin structure is also necessary for the very first cleavages [6–9]. Methylation is one of the most important regulators of the tertiary structure and imprinting; it affects sperm DNA and histones packaging it; in this way a correct recycling of homocysteine is mandatory during spermatogenesis and embryogenesis. The sperm is not just a carrier of paternal genome: it has a relevant epigenetic contribution. A defective wrapping of DNA is often linked to immaturity. Hyaluronic acid (HA) binding ability has been proposed as a tool to select the more mature spermatozoa having reached their final nuclear and cytoplasmic maturation [10, 11], even if controversies exist [12], and it is used as well in veterinary medicine [13]. In this case report we have tested the relation between HA binding and methylation effector supplementation, by comparison to what we have observed with such a supplementation for patients with poor sperm condensation.
- Published
- 2011
80. Ion currents involved in oocyte maturation, fertilization and early developmental stages of the ascidian Ciona intestinalis
- Author
-
Elisabetta, Tosti, Alessandra, Gallo, and Francesco, Silvestre
- Subjects
Embryo, Nonmammalian ,Oogenesis ,Time Factors ,Fertilization ,Oocytes ,Animals ,Embryonic Development ,Urochordata ,Ion Channels ,Ciona intestinalis ,Electrophysiological Phenomena ,Membrane Potentials - Abstract
Electrophysiological techniques were used to study the role of ion currents in the ascidian Ciona intestinalis oocyte plasma membrane during different stages of growth, meiosis, fertilization and early development. Three stages of immature oocytes were discriminated in the ovary, with the germinal vesicle showing specific different features of growth and maturation. Stage-A (pre-vitellogenic) oocytes exhibited the highest L-type calcium current activity and were incompetent for meiosis resumption. Stage-B (vitellogenic) oocytes showed a progressive disappearance of calcium currents and the first appearance of sodium currents that remained high during the maturation process, up to the post-vitellogenic stage-C oocytes. The latter had acquired meiotic competence, undergoing spontaneous in vitro maturation and interacting with the spermatozoon. However, fertilized oocytes did not produce normal larvae, suggesting that cytoplasmic maturation may affect embryo development. In mature oocytes at the metaphase I stage, sodium currents were present and remained high up to the zygote stage. Oocytes fertilized in the absence of sodium showed significant reduction of the fertilization current amplitude and high development of anomalous "rosette" embryos. Current amplitudes became negligible in embryos at the 2- and 4-cell stage, whereas resumption of all the current activities occurred at the 8-cell embryo. Taken together, these results suggest: (i) an involvement of L-type calcium currents in initial oocyte meiotic progression and growth; (ii) a role of sodium currents at fertilization; (iii) a role of the fertilization current in ensuring normal embryo development.
- Published
- 2011
81. Index
- Author
-
Elisabetta Tosti and Raffaele Boni
- Published
- 2011
82. Oocyte Maturation and Fertilization: A Long History for a Short Event
- Author
-
Mahbubur M. Rahman, Georgia Pennarossa, Raffaele Boni, Christopher Malcuit, A. Vanelli, Kay Elder, Yves Menezo, Gian Luigi Russo, Grazyna Ptak, Nava Dekel, Alex Tsafriri, Fulvio Gandolfi, Pasqualino Loi, Marc-André Sirard, Rafael A. Fissore, Brian Dale, Peter Sutovsky, Young-Joo Yi, Francesco Silvestre, Tiziana Al Brevini, Martin Wilding, Stefania Bilotto, Poul Hyttel, and Elisabetta Tosti
- Subjects
Andrology ,Human fertilization ,medicine.anatomical_structure ,Event (relativity) ,medicine ,Biology ,Oocyte - Abstract
Events of reproduction occurring from meiotic resumption of the immature oocyte up to its exit from the second meiotic block following activation will be revealed. Morphological modifications of the oocyte during maturation will be related to signal transduction mechanisms involving primary or secondary messengers. A detailed view will be addressed to genetic and epigenetic control of oocyte maturation. At fertilization, reciprocal gamete activation and novel knowledge related to sperm factor and cascade mechanisms occurring in the oocyte will be focused. A detailed description of ion currents occurring during fertilization will depict another point of view of oocyte activation mechanisms, further supporting for their complexity. Finally, all basic information related to this short time lapse will be considered in relation to clinical application of assisted reproductive technologies. New frontiers, such as stem cells and cloning technologies, will be analyzed and future applications and improvements will be hypothesised.
- Published
- 2011
83. Inositol tri-phosphate in human and ascidian spermatozoa
- Author
-
Brian Dale, Elisabetta Tosti, and Anna Palumbo
- Subjects
Male ,Acrosome reaction ,Ionophore ,chemistry.chemical_element ,Inositol 1,4,5-Trisphosphate ,Calcium ,chemistry.chemical_compound ,Species Specificity ,Genetics ,medicine ,Animals ,Humans ,Ciona intestinalis ,Inositol ,Phosphatidylinositol ,Inositol phosphate ,Calcimycin ,chemistry.chemical_classification ,biology ,Spermatozoon ,urogenital system ,Cell Biology ,biology.organism_classification ,Spermatozoa ,Microscopy, Electron ,medicine.anatomical_structure ,Biochemistry ,chemistry ,embryonic structures ,Acrosome ,Developmental Biology - Abstract
Using a specific protein binding assay we have shown that a spermatozoon of the ascidian Ciona intestinalis contains 1.58 +/- 0.74 x 10(-19) moles of inositol 1,4,5-tri-phosphate (InsP3), while a human spermatozoon contains 6.4 +/- 0.14 x 10(-19) moles. Induction of the acrosome reaction (AR) in both species, by exposure to the calcium ionophore A23187, does not significantly alter levels of InsP3, suggesting that phosphatidylinositol (PI) turnover is not necessary for the calcium ionophore induced AR. Furthermore, PI turnover in ascidian spermatozoa appears to be insensitive to lithium and phorbol ester. The high intracellular concentration of InsP3 in spermatozoa, corresponding to 50-200 microM, suggests it may play a role in egg activation.
- Published
- 1993
84. Role of cyclic AMP in the maturation of Ciona intestinalis oocytes
- Author
-
Francesco Silvestre, Tiziana Covino, Annunziata Cuomo, Alessandra Gallo, and Elisabetta Tosti
- Subjects
Adenylate kinase ,Oogenesis ,03 medical and health sciences ,Follicle ,0302 clinical medicine ,medicine ,Cyclic AMP ,Animals ,Ciona intestinalis ,030304 developmental biology ,Cell Nucleus ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Germinal vesicle ,biology ,Chemistry ,Phosphodiesterase ,Embryo ,Cell Biology ,Oocyte ,biology.organism_classification ,Cell biology ,medicine.anatomical_structure ,Oocytes ,Developmental Biology ,Adenylyl Cyclases ,Signal Transduction - Abstract
SummaryImmature oocytes are arrested at prophase I of the meiotic process and maturation onset is indicated by oocyte nuclear disassembly (germinal vesicle breakdown or GVBD). Signaling pathways that elevate intracellular cyclic AMP (cAMP) may either prevent or induce oocyte maturation depending on the species. In some marine invertebrates and, in particular, in ascidian oocytes, cAMP triggers GVBD rather than blocking it. In this paper, we tested different cAMP elevators in fully grown oocytes at the germinal vesicle stage (GV) of the ascidianCiona intestinalis. We demonstrated that through the activation of adenylate cyclase or the inhibition and phosphodiesterases the oocyte remained at the GV stage. This effect was reversible as the GV-arrested oocytes, rinsed and incubated in sea water, are able to undergo spontaneous maturation and extrusion of follicle cells. In addition, oocytes acquire the ability to be fertilized and start early development. However, morphology of follicle cells, embryos and larvae fromin vitromatured oocytes showed different morphology from those derived fromin vivomature oocytes. The role and the transduction mechanism of cAMP in the regulation of oocyte maturation were discussed. Finally, we indicated a variation of biological mechanisms present in the ascidian species; moreover, we sustain evidence proving that tunicates share some biological mechanisms with vertebrates. This information provided new hints on the importance of ascidians in the evolution of chordates.
- Published
- 2010
85. Dynamic roles of ion currents in early development
- Author
-
Elisabetta Tosti
- Subjects
Embryo, Nonmammalian ,Embryonic Development ,Biology ,Ion Channels ,Ion ,Membrane Potentials ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Genetics ,Animals ,Humans ,Urochordata ,Ion channel ,030304 developmental biology ,0303 health sciences ,Zygote ,Cell Membrane ,Fishes ,Embryo ,Cell Biology ,Anatomy ,Blastomere ,Blastula ,Embryo, Mammalian ,Electrophysiology ,Sea Urchins ,Biophysics ,Cattle ,Signal transduction ,Anura ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Excitable cells have the capacity to modify their electrical properties in response to different stimuli. This specific feature is due to a flux of ion currents that flow via ion channels in the plasma membrane. In all species so far studied, ion channels are proteins expressed in the zygote and in the blastomeres of the developing embryo, and their activity is subject to dynamic changes throughout the early cleavage stages. Although these complex patterns imply that ion currents play a role in signal transduction and the control of embryogenesis, a specific developmental function for the appearance, loss, and alterations of the channels remains to be elucidated. This review reports several aspects surrounding the involvement of ion currents in early embryo development, from invertebrates to human. It focuses on the occurrence, modulation, and dynamic role of ion fluxes through external, intra- and inter-cellular ion channels from the zygote up to the blastula and pre-implantation stages. The implications for a role of ion currents in development, and their possible clinical and technological applications are discussed.
- Published
- 2010
86. 5-HT-receptive structures are localized in the interblastomere cleft of Paracentrotus lividus early embryos
- Author
-
Elisabetta Tosti, Francesco Silvestre, and Yu. B. Shmukler
- Subjects
Agonist ,Male ,Blastomeres ,Serotonin ,Embryo, Nonmammalian ,medicine.drug_class ,Cleavage (embryo) ,Ligands ,Paracentrotus lividus ,Membrane Potentials ,biology.animal ,medicine ,Animals ,Receptor ,Sea urchin ,5-HT receptor ,biology ,Quipazine ,Embryogenesis ,Cell Membrane ,Cell Biology ,Anatomy ,biology.organism_classification ,Spermatozoa ,Cell biology ,Electrophysiology ,Receptors, Serotonin ,Paracentrotus ,Female ,Developmental Biology ,medicine.drug - Abstract
SummaryLocal application of the agonists of serotonin receptors of third type 5-HTQ, SR57277A and quipazine into the interblastomere cleft of the sea urchinParacentrotus lividusembryo during first cleavage division, evokes specific membrane currents, whereas application of these drugs out of contact area show currents of lower amplitude and longer latent period. At the same time 5-HT3-receptor agonist quipazine imitates interblastomere signal in half embryos, but corresponding antagonists prevent it. Present data develop the hypothesis of protosynapse, demonstrating that the distribution of membrane serotonin receptors is limited to the period of cleavage division and localized in the interblastomere contact area. A possible role of spatial–temporal restriction of receptors at the interblastomere contact area is discussed in relation to the subsequent embryo development.
- Published
- 2008
87. Cytotoxic and apoptogenic activity of a methanolic extract from the marine invertebrate Ciona intestinalis on malignant cell lines
- Author
-
Elisabetta Tosti, Gian Luigi Russo, Rosa Anna Siciliano, Gaetano Ciarcia, Emilio Presidente, Russo, Gl, Ciarcia, Gaetano, Presidente, E, Siciliano, Ra, and Tosti, E.
- Subjects
Ciona intestinali ,ascidian ,Antineoplastic Agents ,tunicate ,010501 environmental sciences ,Ecteinascidia turbinata ,01 natural sciences ,Aplidium ,03 medical and health sciences ,In vivo ,Cell Line, Tumor ,Drug Discovery ,Botany ,Animals ,Humans ,Ciona intestinalis ,Invertebrate ,030304 developmental biology ,0105 earth and related environmental sciences ,Cell Proliferation ,0303 health sciences ,biology ,Cell growth ,Caspase 3 ,apoptosis ,Marine invertebrates ,biology.organism_classification ,apoptosi ,In vitro ,Tunicate ,Nucleosomes ,Biochemistry ,cytotoxicity - Abstract
Marine invertebrates provide a series of natural products with different biological activities. Several of these compounds and their derivatives showed a potent anticancer effect. Tunicates represent an important source of bioactive agents, leading to the isolation of ecteinascidin-743 (ET-743), a compound isolated from the Caribbean sea squirt Ecteinascidia turbinata with a potent cytotoxic activity against a variety of tumours in vitro and in vivo. Current phase II clinical trials against soft tissue sarcomas in Europe and the United States indicate that ET-743 represents a highly promising anticancer agent. Another example is aplidine from the Mediterranean tunicate Aplidium albicans, with a broad spectrum activity against various types of cancers, such as colorectal, lymphoma, thyroid and renal cancers. In the present work, we reported, for the first time, that a partially purified methanolic extract prepared from the ascidian Ciona intestinalis inhibited cell proliferation in human cell lines of different origin, including Caco2, HPB-ALL, U-937 and HL-60 and induced early apoptotic events, such as caspase-3 activation and internucleosomal DNA degradation. We suggest the presence in the Ciona intestinalis extract of bioactive compounds possessing anticancer activity.
- Published
- 2008
88. Calcium and other ion dynamics during gamete maturation and fertilization
- Author
-
Elisabetta Tosti, Raffaele Boni, Roberto Gualtieri, Riccardo Talevi, Boni, R, Gualtieri, Roberto, Talevi, Riccardo, and Tosti, E.
- Subjects
Male ,medicine.medical_specialty ,chemistry.chemical_element ,Calcium ,Biology ,Models, Biological ,03 medical and health sciences ,0302 clinical medicine ,Human fertilization ,Food Animals ,Capacitation ,Internal medicine ,medicine ,Animals ,Humans ,Small Animals ,030304 developmental biology ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Ion Transport ,Equine ,Oocyte activation ,Oocyte ,Sperm ,In vitro maturation ,Cell biology ,Sperm Maturation ,medicine.anatomical_structure ,Endocrinology ,Germ Cells ,chemistry ,Fertilization ,Oocytes ,Gamete ,Animal Science and Zoology ,Female ,Sperm Capacitation - Abstract
Ion currents and cytosolic free calcium ([Ca(2+)](i)) elevations are crucial events in triggering the complex machinery involved in both gamete maturation and fertilization. Oocyte maturation is triggered by hormone signaling which causes ion currents and [Ca(2+)](i) increase. Extracellular calcium seems to be required for meiosis progression since: (i) calcium depletion in the maturation medium severely affects oocyte developmental competence; (ii) the activity of plasma membrane L-type Ca(2+) currents decreases during maturation; (iii) the exposure to verapamil, a specific Ca(2+) channel blocker, decreases in vitro maturation efficiency. In spermatozoa, maturation initiates inside the epididymis and ends in the female genital tract. During their journey through the female reproductive tract, sperm undergo a dramatic selection and capacitation achieving fertilization competence. Adhesion to the tubal epithelium extends sperm life through depression of [Ca(2+)](i) until capacitation signals trigger an [Ca(2+)](i) elevation followed by sperm release. At fertilization, egg-sperm interaction evokes well-described transient and almost simultaneous events: i.e., fertilization current, a change in resting potential, and an increase in free [Ca(2+)](i) concentration. These events, termed oocyte activation, are the direct consequence of sperm interaction via either activation of a receptor or entry of a sperm factor. The latter hypothesis has been recently supported by the discovery of PCLzeta, a sperm-specific isozyme triggering a dramatic [Ca(2+)](i) increase via inositol 1,4,5-trisphosphate (IP(3)) production. The course of ion currents and [Ca(2+)](i) transients during maturation and fertilization plays a pivotal role in correct embryo development.
- Published
- 2007
89. APEX/Ref-1 (Apurinic/Apyrimidic Endonuclease DNA-Repair Gene) Expression in Human and Ascidian (Ciona intestinalis) Gametes and Embryos
- Author
-
Gian Luigi Russo, Stefania Bilotto, Said El-Mouatassim, Elisabetta Tosti, and Yves Menezo
- Subjects
Male ,Embryology ,Embryo, Nonmammalian ,DNA Repair ,DNA damage ,DNA repair ,Molecular Sequence Data ,Gene Expression ,Evolution, Molecular ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,DNA-(Apurinic or Apyrimidinic Site) Lyase ,Genetics ,Animals ,Humans ,Ciona intestinalis ,AP site ,Amino Acid Sequence ,RNA, Messenger ,Molecular Biology ,Phylogeny ,030304 developmental biology ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Base Sequence ,biology ,Obstetrics and Gynecology ,Cell Biology ,biology.organism_classification ,Spermatozoa ,Apex (geometry) ,Cell biology ,Ciona ,Blastocyst ,Reproductive Medicine ,chemistry ,Oocytes ,DNA fragmentation ,Female ,DNA ,Developmental Biology - Abstract
In recent years, the impact of sperm DNA damage on fertility has become an important issue. The different technologies developed to check sperm DNA fragmentation lead to the same conclusion: DNA damage negatively impacts upon reproductive processes. Oocyte DNA repair capacity is one of the cues to understanding embryo developmental arrest. APEX/Ref-1 (apurinic/apyrimidic endonuclease) is an enzyme involved in the DNA base excision repair pathway removing the abasic sites, the most common DNA decays. In humans, APEX has a multifunctional role, including the control of the redox status of transcription factors. RT-PCR allowed us to detect human APEX transcripts in oocytes, spermatozoa and preimplantation blocked embryos. In parallel, a comparative study on sea squirt Ciona intestinalis (ascidian) indicated that APEX transcripts are clearly detectable in oocytes and embryos until the larva stage, but not in spermatozoa, suggesting the appearance of the paternal contribution to DNA repair during development having arisen only late in Vertebrate evolution. Of additional phylogenetic significance is the observation that sea squirt APEX appears to lack redox transcriptional activity.
- Published
- 2007
90. Calcium ion currents mediating oocyte maturation events
- Author
-
Elisabetta Tosti
- Subjects
Calcium Channels, L-Type ,lcsh:QH471-489 ,chemistry.chemical_element ,Review ,Biology ,Calcium ,Oogenesis ,lcsh:Gynecology and obstetrics ,Endocrinology ,Prophase ,Meiosis ,medicine ,Animals ,Humans ,lcsh:Reproduction ,Ion channel ,lcsh:RG1-991 ,Germinal vesicle ,Voltage-dependent calcium channel ,Cell Membrane ,Obstetrics and Gynecology ,Oocyte ,Cell biology ,medicine.anatomical_structure ,Reproductive Medicine ,chemistry ,Oocytes ,Calcium Channels ,Developmental Biology - Abstract
During maturation, the last phase of oogenesis, the oocyte undergoes several changes which prepare it to be ovulated and fertilized. Immature oocytes are arrested in the first meiotic process prophase, that is morphologically identified by a germinal vesicle. The removal of the first meiotic block marks the initiation of maturation. Although a large number of molecules are involved in complex sequences of events, there is evidence that a calcium increase plays a pivotal role in meiosis re-initiation. It is well established that, during this process, calcium is released from the intracellular stores, whereas less is known on the role of external calcium entering the cell through the plasma membrane ion channels. This review is focused on the functional role of calcium currents during oocyte maturation in all the species, from invertebrates to mammals. The emerging role of specific L-type calcium channels will be discussed.
- Published
- 2006
91. Intracellular calcium and protein tyrosine phosphorylation during the release of bovine sperm adhering to the fallopian tube epithelium in vitro
- Author
-
Elisabetta Tosti, Raffaele Boni, Maria Zagami, Riccardo Talevi, Roberto Gualtieri, Gualtieri, Roberto, Boni, R., Tosti, E., Zagami, Maria, and Talevi, Riccardo
- Subjects
Intracellular Fluid ,Male ,Embryology ,medicine.medical_specialty ,Thapsigargin ,media_common.quotation_subject ,Motility ,Biology ,Epithelium ,chemistry.chemical_compound ,Endocrinology ,Capacitation ,Internal medicine ,Culture Techniques ,medicine ,Cell Adhesion ,Animals ,Phosphorylation ,Ovulation ,Fallopian Tubes ,media_common ,Hyperactivation ,urogenital system ,Heparin ,Obstetrics and Gynecology ,Tyrosine phosphorylation ,Cell Biology ,Protein-Tyrosine Kinases ,Sperm ,Spermatozoa ,Cell biology ,Reproductive Medicine ,chemistry ,Oviduct ,Calcium ,Cattle ,Female - Abstract
In mammals, sperm adhesion to the epithelial cells lining the oviductal isthmus plays a key role in the maintenance of motility and in the selection of superior quality subpopulations. In the bovine species, heparin and other sulfated glycoconjugates powerfully induce the synchronous release of sperm adhering to tubal epitheliumin vitroand may represent the signal which triggers release at ovulationin vivo. Sperm detachment may be due either to surface remodeling or to hyperactivation brought about by capacitation. In this paper, the dynamics of intracellular free Ca2+concentration ([Ca2+]i) and protein tyrosine phosphorylation in sperm during and after heparin-induced release fromin vitrocultured oviductal monolayers were assessed to determine whether this event is due to capacitation. Moreover, Ca2+-ionophore A23187, thapsigargin, thimerosal and caffeine were used to determine whether [Ca2+]iincrease and/or hyperactivation can induce sperm release. Results showed that: 1. heparin-released sperm have significantly higher [Ca2+]ithan adhering sperm; 2. heparin induces a [Ca2+]ielevation in the sperm head followed by detachment from the monolayers; 3. external Ca2+is not required for heparin-induced release; 4. [Ca2+]iincrease and/or hyperactivation are unable to release sperm; and 5. heparin-released sperm have an increased level of tyrosine phosphorylated proteins compared with adhering sperm. In conclusion, although heparin is considered a long-lasting capacitation agent, it quickly modulates the capacitation of bovine sperm adhering to the fallopian epithelium, probably leading to surface remodeling and therefore to the release of sperm selected and stored within the oviduct through adhesion.
- Published
- 2004
92. Biochemical and functional characterization of protein kinase CK2 in ascidian Ciona intestinalis oocytes at fertilization. Cloning and sequence analysis of cDNA for alpha and beta subunits
- Author
-
Gian Luigi, Russo, Mariarosaria, Tosto, Annalisa, Mupo, Immacolata, Castellano, Annunziata, Cuomo, and Elisabetta, Tosti
- Subjects
DNA, Complementary ,Time Factors ,MAP Kinase Signaling System ,Protein Conformation ,Immunoblotting ,Molecular Sequence Data ,Protein Serine-Threonine Kinases ,Catalysis ,Catalytic Domain ,Serine ,Animals ,Humans ,Amino Acid Sequence ,RNA, Messenger ,Cloning, Molecular ,Phosphorylation ,Casein Kinase II ,Metaphase ,DNA Primers ,Binding Sites ,Base Sequence ,Dose-Response Relationship, Drug ,Sequence Homology, Amino Acid ,Sequence Analysis, DNA ,Precipitin Tests ,Ciona intestinalis ,Protein Structure, Tertiary ,Meiosis ,Fertilization ,Chromatography, Gel ,Oocytes ,Electrophoresis, Polyacrylamide Gel - Abstract
The ubiquitous and pleiotropic dual specificity protein kinase CK2 has been studied and characterized in many organisms, from yeast to mammals. Generally, the enzyme is composed of two catalytic (alpha and/or alpha') and two regulatory (beta) subunits, forming a differently assembled tetramer. Although prone to controversial interpretation, the function of CK2 has been associated with fundamental biological processes such as signal transduction, cell cycle progression, cell growth, apoptosis, and transcription. Less known is the role of CK2 during meiosis and the early phase of embryogenesis. In this work, we studied CK2 activity during oocyte activation, a process occurring at the end of oocyte maturation and triggered by fertilization. In ascidian Ciona intestinalis, an organism whose complete genome has been published recently, CK2 was constitutively active in unfertilized and fertilized oocytes. The enzymatic activity oscillated through meiosis showing three major peaks: soon after fertilization (metaphase I exit), before metaphase II, and at the exit from metaphase II. Biochemical analysis of CK2 subunit composition in activated oocytes indicated that CK2-alpha was catalytically active as a monomer, independently from its regulatory subunit beta; however, CK2-beta was only detectable in unfertilized oocytes where it was associated with a bona fide identified ascidian mitogen-activated protein kinase. After fertilization, CK2-beta was undetectable, suggesting its rapid degradation. Protein sequence analysis of CK2-alpha and -beta cDNA indicated a high identity compared with vertebrate homologs. In addition, the absence of putative phosphorylation sites for Cdc2 kinase on both alpha and beta subunits suggested an important role for CK2 in regulating meiotic cell cycle in C. intestinalis oocytes.
- Published
- 2004
93. Antifouling Compounds Endangering Marine Invertebrates Reproduction
- Author
-
Elisabetta Tosti, Alessandra Gallo, primary
- Published
- 2014
- Full Text
- View/download PDF
94. Fertilization and activation currents in bovine oocytes
- Author
-
Raffaele Boni, Elisabetta Tosti, and Annunziata Cuomo
- Subjects
Embryology ,medicine.medical_specialty ,Thapsigargin ,Patch-Clamp Techniques ,Potassium Channels ,Voltage clamp ,Parthenogenesis ,chemistry.chemical_element ,Fertilization in Vitro ,Inositol 1,4,5-Trisphosphate ,Calcium ,03 medical and health sciences ,chemistry.chemical_compound ,Potassium Channels, Calcium-Activated ,0302 clinical medicine ,Endocrinology ,Oogenesis ,Internal medicine ,Caffeine ,medicine ,Animals ,Egtazic Acid ,Ion channel ,030304 developmental biology ,Chelating Agents ,Sperm-Ovum Interactions ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Ethanol ,Ionophores ,Ryanodine receptor ,Cell Membrane ,Obstetrics and Gynecology ,Cell Biology ,Hyperpolarization (biology) ,Oocyte ,Stimulation, Chemical ,EGTA ,medicine.anatomical_structure ,Reproductive Medicine ,chemistry ,Biophysics ,Oocytes ,Cattle ,Female - Abstract
One of the first events that occurs at fertilization is a transient modification of the electrical properties of the oocyte plasma membrane. The whole-cell voltage clamp technique was used to demonstrate an outward ion current and a hyperpolarization of the plasma membrane after fertilization in bovine oocytes. These electrical events, together with measurement of internal calcium concentrations, were also recorded after injection with sperm factor and exposure to parthenogenetic activators, such as Ca(2+) ionophore, ethanol and thapsigargin. Experiments were carried out simultaneously in immature and in vitro matured oocytes. Significant differences were recorded in the activation current and hyperpolarization among oocyte activators and between immature and matured oocytes. However, outward ion current and Ca(2+) release showed similar dynamics. The injection of the calcium chelator EGTA completely abolished both ion current and hyperpolarization, indicating that these electrical events are calcium dependent. Addition of specific calcium releasers, such as 1,4,5-inositol trisphosphate (IP(3)) and caffeine, triggered ion activation current and hyperpolarization indicating that IP(3) and ryanodine receptors are active in both immature and matured oocytes. Different ion channel inhibitors were used to characterize the channels underlying outward currents. Only addition of rIberiotoxin caused a complete inhibition of the current, indicating the involvement of high conductance Ca(2+)-activated K(+) channels in generating activation current. In conclusion, these findings provide evidence that bovine oocyte activation is associated with Ca(2+)-dependent electrical events. Oocytes have the potential to react to different activators even when immature; however, oocyte maturation seems to increase sensitivity to physiological activators, such as spermatozoa and sperm factor, and chemicals, such as ethanol.
- Published
- 2003
95. Ca(2+) current activity decreases during meiotic progression in bovine oocytes
- Author
-
Elisabetta Tosti, Raffaele Boni, and Annunziata Cuomo
- Subjects
medicine.medical_specialty ,Patch-Clamp Techniques ,Membrane permeability ,Calcium Channels, L-Type ,Nifedipine ,Physiology ,Biology ,In Vitro Techniques ,Membrane Potentials ,Cell membrane ,03 medical and health sciences ,0302 clinical medicine ,Meiosis ,Internal medicine ,Follicular phase ,medicine ,Animals ,Ovarian follicle ,Metaphase ,Calcimycin ,030304 developmental biology ,0303 health sciences ,030219 obstetrics & reproductive medicine ,Germinal vesicle ,Ionophores ,Cell Membrane ,Cell Biology ,Oocyte ,Calcium Channel Blockers ,Cell biology ,medicine.anatomical_structure ,Endocrinology ,Verapamil ,Oocytes ,Calcium ,Cattle - Abstract
By using the whole cell voltage-clamp technique, we studied changes in plasma membrane permeability at different meiotic stages of bovine oocytes. Follicular oocytes were matured in vitro and activated by Ca2+ionophore. Oocytes at germinal vesicle (GV), germinal vesicle breakdown (GVBD), metaphase I (MI), metaphase II (MII), and meiosis exit were used for electrophysiological recording. By clamping the oocytes at −30 mV, we found that the L-type voltage-dependent Ca2+channels were active at the GV stage and that their activity decreased after the GVBD stage. Furthermore, the resting potential decreased from the GV to the MI stage and increased again at MII. A significant decrease of the steady-state conductance occurred from the GV to the MI stage, followed by a sharp increase at the MII stage. With the addition of organic L-type Ca2+channel blockers (nifedipine and verapamil), we inhibited the Ca2+currents. However, only in the case of verapamil was there a decrease of in vitro maturation efficiency. Our results suggest that, in addition to the cumulus-oocyte junctions, the plasma membrane channels provide another mode of Ca2+entry into bovine oocytes during meiosis.
- Published
- 2000
96. Intercellular communication in in vivo- and in vitro-produced bovine embryos
- Author
-
Raffaele Boni, Elisabetta Tosti, S. Roviello, and Brian Dale
- Subjects
Blastomeres ,animal structures ,Cell Communication ,Fertilization in Vitro ,Biology ,Cell junction ,Andrology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,In vivo ,medicine ,Inner cell mass ,Animals ,Blastocyst ,reproductive and urinary physiology ,030304 developmental biology ,Fluorescent Dyes ,0303 health sciences ,Lucifer yellow ,030219 obstetrics & reproductive medicine ,Embryo ,Cell Biology ,General Medicine ,Blastomere ,Isoquinolines ,In vitro ,Microscopy, Electron ,medicine.anatomical_structure ,Reproductive Medicine ,chemistry ,embryonic structures ,Immunology ,Cattle - Abstract
In vivo bovine embryos were obtained by nonsurgical flushing of uterine horns of cows submitted to superovulatory treatment, while in vitro embryos were generated from oocytes collected from slaughtered donors. Lucifer Yellow injected into single blastomeres did not diffuse into neighboring cells until the morula stage in in vivo embryos and the blastocyst stage in in vitro embryos. In both cases diffusion was limited to a few cells. In contrast, diffusion was extensive in microsurgically isolated inner cell mass (ICM) but absent in the trophectoderm (TE). At the blastocyst stage, diffusion was always more extensive in in vivo than in in vitro embryos. Ultrastructural analyses confirmed these functional observations, and gap junction-like structures were observed at the blastocyst stage. These structures were diffuse in the ICM of in vivo embryos, scarce in the ICM of in vitro embryos and in the TE of in vivo embryos, and not observed in the TE of in vitro embryos. Blastomeres at all stages of development from the 2-cell stage to the blastocyst stage in in vitro embryos and at the morula and blastocyst stage in in vivo embryos were electrically coupled, and the junctional conductance (Gj) decreased in in vitro embryos from 4.18 +/- 1.70 nS (2-cell stage) to 0.37 +/- 0.12 nS (blastocyst stage). At each developmental stage, in vivo embryos showed a significantly (P < 0. 05) higher Gj than in vitro-produced embryos. Moreover, a significantly (P < 0.01) higher Gj was found in isolated ICM than in the respective blastocyst in both in vivo- and in vitro-produced embryos (3.5 +/- 1.4 vs. 0.7 +/- 0.3 and 2.6 +/- 1.6 vs. 0.37 +/- 0. 12 nS, respectively). The electrical coupling in absence of dye coupling in the early bovine embryo agrees with observations for embryos from other phyla. The late and reduced expression of intercellular communicative devices in in vitro-produced embryos may be one of the factors explaining their developmental low efficiency.
- Published
- 1999
97. Activation of Ciona Intestinalis at Fertilisation is Controlled by Nicotinamide Nucleotide Metabolism
- Author
-
M. L. De Simone, Marcella Marino, Martin Wilding, Brian Dale, Elisabetta Tosti, Gian Luigi Russo, and L. Grumetto
- Subjects
0303 health sciences ,030219 obstetrics & reproductive medicine ,biology ,Chemistry ,Meiosis II ,chemistry.chemical_element ,Calcium ,Oocyte ,biology.organism_classification ,Calcium in biology ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,medicine.anatomical_structure ,Biochemistry ,Second messenger system ,medicine ,Ciona intestinalis ,Intracellular ,Fertilisation ,030304 developmental biology - Abstract
Fertilisation is characterised by an ion current across the oocyte plasma membrane (DeFelice et al., 1986), a transient increase in intracellular calcium (see Jaffe, 1985), and the release of the cell cycle block controlled at the level of MPF (Newport and Kirschner, 1984). The fertilisation calcium signal appears to trigger the resumption of the cell cycle at fertilisation in oocytes arrested at meiosis II or after the completion of meiosis (reviewed in Whitaker and Patel, 1990). However, the role of intracellular second messengers, including calcium, in the control of progression through the early meiotic cell cycle, is less well defined (see Whitaker and Patel, 1990; Whitaker, 1996).
- Published
- 1998
98. Meiotic Cell Cycle Control by Mos in Ascidian Oocytes
- Author
-
Keiichiro Kyozuka, Elisabetta Tosti, Marcella Marino, Brian Dale, Maria Laura de Simone, Martin Wilding, and Gian Luigi Russo
- Subjects
0303 health sciences ,Cell division ,DNA replication ,Biology ,Cell cycle ,Cell biology ,Chromosome segregation ,03 medical and health sciences ,0302 clinical medicine ,Meiosis ,Signal transduction ,Mitosis ,030217 neurology & neurosurgery ,030304 developmental biology ,Anaphase - Abstract
Mitotic cell division cycle is regulated by several control mechanisms generally known as checkpoints, whose main function is to ensure that critical events in the cell division such as DNA replication and chromosome segregation occur with high fidelity and in the correct order and time. A recent view of cell cycle regulation, indicate that checkpoints work as signal transduction pathways with their initiating signals, sensors, transducers, and effectors (Elledge, 1996). Similarly, meiotic cell cycle regulation have specific checkpoints and two of them have been recently characterized at molecular level. The first ensures the completion of recombination before the formation of meiosis I spindle; the second blocks anaphase I until all the paired chromosomes are correctly attached to the spindle (Page and Orr-Weaver, 1997). However, the most intriguing difference between mitosis and meiosis respect to cell cycle regulation, is the ability of animal oocytes to arrest at specific stage during maturation, and maintain this block for extremely long time: from years (frog) to decades (human) (Sagata, 1996a,b). It is universally accepted that the meiotic block is due to the activity of a cytostatic factor (CSF) primarily identified by Masui (Masui and Markert, 1971).
- Published
- 1998
99. A soluble sperm factor gates Ca(2+)-activated K+ channels in human oocytes
- Author
-
Brian Dale, Elisabetta Tosti, Adriana Fortunato, and Vincenzo Monfrecola
- Subjects
Cell Extracts ,Male ,medicine.medical_specialty ,Patch-Clamp Techniques ,Potassium Channels ,Ionophore ,chemistry.chemical_element ,Semen ,Gating ,Calcium ,Andrology ,Human fertilization ,Internal medicine ,Genetics ,medicine ,Humans ,Genetics (clinical) ,Calcimycin ,Sperm-Ovum Interactions ,Ionophores ,urogenital system ,Obstetrics and Gynecology ,General Medicine ,Iberiotoxin ,Oocyte ,Sperm ,Spermatozoa ,Endocrinology ,medicine.anatomical_structure ,Reproductive Medicine ,chemistry ,Solubility ,Oocytes ,Female ,Peptides ,Ion Channel Gating ,Developmental Biology - Abstract
Purpose: Our goal was to study the activation current in physiologically competent metaphase II human oocytes, i.e., not previously exposed to spermatozoa or aged in vitro, and, in particular, to determine whether a soluble sperm factor triggers a fertilization current comparable to that observed with intact spermatozoa and to characterize the current involved. Methods: The whole-cell voltage-clamp technique was used on spare metaphase II oocytes, obtained with patient consent from IVF programs. In this configuration a soluble fraction from human spermatozoa was microinjected, and the current recorded. Results: Metaphase II human oocytes generate bell-shaped outward currents of 400–1000 pA (X=706±322;n=10), following injection of a cytosolic extract from human spermatozoa. The amount of sperm extract injected was less than 10% of the total oocyte volume and was equivalent to 1–10 spermatozoa. A similar current was generated following exposure to 20 µM of the calcium ionophore A23187 (n=10). The steady-state conductance of the oocyte increased from 10 to 19.8 nS (n=10) following injection of the sperm factor and from 5.3 to 27.7 nS following ionophore exposure. Both sperm factor- and ionophore-induced currents were reduced in amplitude when the unfertilized oocyte was preexposed to 25–75 µM iberiotoxin (n=8) and eliminated at a concentration of 100 µM iberiotoxin. Conclusions: The data support the hypothesis of a soluble sperm factor involved in the activation of human oocytes and shows that the initial activation response in the human oocyte is the gating of Ca2+-activated K+ channels.
- Published
- 1996
100. Is the plasma membrane of the human oocyte reorganised following fertilisation and early cleavage?
- Author
-
Brian Dale, M. Iaccarino, and Elisabetta Tosti
- Subjects
Blastomeres ,Zygote ,chemistry.chemical_compound ,Lectins ,medicine ,Concanavalin A ,Humans ,Fertilisation ,Fluorescent Dyes ,biology ,Microvilli ,Chemistry ,Cell Membrane ,Lectin ,Embryo ,Cell Biology ,Blastomere ,Anatomy ,Plants ,Oocyte ,Embryo, Mammalian ,Cell biology ,Sialic acid ,medicine.anatomical_structure ,Fertilization ,biology.protein ,Microscopy, Electron, Scanning ,Oocytes ,Female ,Plant Lectins ,Developmental Biology - Abstract
SummaryThe purpose of the present study was to determine whether the plasma membrance of the human oocyte is reorganised following fertlisation and during early cleavage. In order to characterise and localise the major sugar moieties on surface glycoporteins, oocytes and embroys were labelled with a range of flourescent lectins. Regional organisation of plasma membrane microvilli in oocytes and embryos was also studied using scanning electron microscopy (SEM). The plasma membrance of human oocytes, zygotes and early blastomeres stained strongly and homogeneously with concanavalin A andTriticum vulgarislectin (WGA), indicationg the presence of plasma membrance glycoconjugates with α-D-mannosyl residues, sialic acid and β-NAc-glucosaminyl groups. We did not observe regional domains in oocytes and zygotes, suggesting that the plasma membrane is not topographically reorganised following fertilisation. SEM shows the surface of the human zygote to be organised into short microvilli 0.2–3.0 μm in length and at a density of 5–20/μm2. In early cleavage stages the microvilli are shorter and less frequent (0.2–1.0 μm; 1–5/μm2); however, there is no evidence of polarisation at this level of organisation, at either stage of development. The surface of cell fragments, common in the human embryoin vitro, differs in having few microvilli and numerous cytoplasmic blebs. In conclusion, there are no obvious morphological signs of regionalisation in the plasma membrane of the human embryo before the 8-cell stage.
- Published
- 1995
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.