Abstract:In this work, five local sea urchin species found in European waters were studied. Four were regular species: Sphaerechinus granularis, Psammechinus miliaris, Echinus esculentus (Linnaeus, 1758) and the edible sea urchin Paracentrotus lividus; and one was an irregular species, Echinocardium cordatum. These five species of sea urchins have been studied regarding their fertility, toxicity of cryoprotecting agents, cryopreservation of different cell types and chilling injury. The baseline fertility is similar in … Show more
“…3 D–F, 4 D–F, 5 D–F, 6 D–F). These results are consistent with the LOEC (Lowest Observed Effect Concentration) values calculated by Paredes and Bellas 17 and Paredes et al 40 for DMSO, EG and PG.…”
Section: Discussionsupporting
confidence: 92%
“…Results have shown that exposing the eggs to low concentrations (0.5 M) of DMSO, EG and PG did not cause damage to the cells (Figs. 3 , 4 , 5 , 6 A–C) which is in agreement with prior data of CPA toxicity 17 , 40 . In the case of eggs, the NOEC (No Observed Effect Concentration) values calculated support the results obtained in this study, since after exposing those cells to low concentrations of CPA, the ultrastructure was similar to controls.…”
Section: Discussionsupporting
confidence: 91%
“…All methods were carried out in accordance with relevant guidelines and regulations of animal handling and care at the Universidade de Vigo. Animals handled in this study are out of scope of the regulations ECC/566/2015, nonetheless animal handling was done to the best standard and following guidelines from Paredes and Costas 65 and Paredes et al 40 . ARRIVE guidelines in experimental design to ensure reliability of research were followed.…”
This study examinates the challenges of cryopreserving sea urchin (Paracentrotus lividus) eggs, a task hindered by factors like low membrane permeability and high sensitivity to cryoprotective agents (CPAs). While successful cryopreservation has been achieved for some marine invertebrates, eggs remain problematic due to their unique characteristics. The study explores the impact of various CPAs and cryopreservation techniques on sea urchin eggs, employing scanning and transmission electron microscopy to analyze cellular damage. The findings reveal that exposure to low CPA concentrations (0.5 M) did not induce significant damage to eggs. However, high concentrations (3 M) proved highly detrimental. Every cryopreservation approach investigated in this study resulted in irreversible damage to the sea urchin eggs, rendering them nonviable for future use. The research sheds light on the importance of understanding the structural alterations induced by CPAs and cryopreservation methods. This knowledge is essential for refining cryopreservation methods, potentially paving the way for successful preservation of these challenging cells.
“…3 D–F, 4 D–F, 5 D–F, 6 D–F). These results are consistent with the LOEC (Lowest Observed Effect Concentration) values calculated by Paredes and Bellas 17 and Paredes et al 40 for DMSO, EG and PG.…”
Section: Discussionsupporting
confidence: 92%
“…Results have shown that exposing the eggs to low concentrations (0.5 M) of DMSO, EG and PG did not cause damage to the cells (Figs. 3 , 4 , 5 , 6 A–C) which is in agreement with prior data of CPA toxicity 17 , 40 . In the case of eggs, the NOEC (No Observed Effect Concentration) values calculated support the results obtained in this study, since after exposing those cells to low concentrations of CPA, the ultrastructure was similar to controls.…”
Section: Discussionsupporting
confidence: 91%
“…All methods were carried out in accordance with relevant guidelines and regulations of animal handling and care at the Universidade de Vigo. Animals handled in this study are out of scope of the regulations ECC/566/2015, nonetheless animal handling was done to the best standard and following guidelines from Paredes and Costas 65 and Paredes et al 40 . ARRIVE guidelines in experimental design to ensure reliability of research were followed.…”
This study examinates the challenges of cryopreserving sea urchin (Paracentrotus lividus) eggs, a task hindered by factors like low membrane permeability and high sensitivity to cryoprotective agents (CPAs). While successful cryopreservation has been achieved for some marine invertebrates, eggs remain problematic due to their unique characteristics. The study explores the impact of various CPAs and cryopreservation techniques on sea urchin eggs, employing scanning and transmission electron microscopy to analyze cellular damage. The findings reveal that exposure to low CPA concentrations (0.5 M) did not induce significant damage to eggs. However, high concentrations (3 M) proved highly detrimental. Every cryopreservation approach investigated in this study resulted in irreversible damage to the sea urchin eggs, rendering them nonviable for future use. The research sheds light on the importance of understanding the structural alterations induced by CPAs and cryopreservation methods. This knowledge is essential for refining cryopreservation methods, potentially paving the way for successful preservation of these challenging cells.
“…The history of cryopreservation of sea urchin sperm began in 1973 (9), and at present, sperm of about 15 species have been cryopreserved as recently reviewed (11,26-28,31). Cryopreservation of sperm of all animals involves dilution of concentrated semen into an extender solution and then mixing this cell suspension with a cryoprotectant such as dimethyl sulfoxide (DMSO).…”
Sea urchins have contributed greatly to knowledge of fertilization, embryogenesis and cell biology. However, until now, they have not been a genetic model organism because of the long generation times of commonly used species, and lack of tools for husbandry and genetic manipulation. We recently establishedLytechinus pictus, as a multigenerational sea urchin model, because of its relatively short generation time of 4-6 months and ease of laboratory culture. To take full advantage of this new multigenerational species, methods are needed to biobank and share mutantL. pictussperm. Here, we describe a new extender based on sperm ion physiology before spawning of sperm into seawater. This extender maintains sperm capable of fertilization for at least 5-10 weeks when stored at 0 °C. We use the extender, and the cryoprotectant dimethyl sulfoxide (DMSO), to cryopreserve sperm of bothL. pictus, and the widely used sea urchin,Strongylocentrotus purpuratus. The simple methods we describe work well for both species, achieving > 90% development and producing larvae that successfully undergo metamorphosis to juvenile sea urchins. Sperm of these two species can be frozen and thawed at least twice and still give rise to larvae that undergo metamorphosis.Main PointsSperm can maintain fertilizing capacityex vivofor 5-10 weeks when stored at 0°C.When freezing in liquid nitrogen no stepwise addition of cryoprotectant, or stepwise drop in temperature are required.A standard fertilization assay is presented to score cleavage stage sea urchin embryos produced by cryopreserved sperm.Sperm frozen and thawed more than once can produce larvae.
“…In all cases, protocols need to be established, shared and standardized within the scientific community to successfully obtain gametes from different mollusks, as well as sea urchins, which also present with external fertilization. Sea urchins are model organisms and, therefore, some species of sea urchins, such as Strongylocentrotus purpuratus or Paracentrotus lividus, are quite well studied and have many protocols in place; however, this is not the case for other sea urchins [6], as it has been shown in many cases that protocols would have to be adapted to particular species. Important knowledge about reproduction is not often well-known or standardized, such as contact time, the egg:sperm ratio, optimum parameters for embryo incubation and expected outcome after larval rearing, but is key information for marine invertebrates [2,6].…”
Artificial reproduction in aquatic animals usually involves the collection and handling of gametes both from males and females in a way that secures their quality and optimizes the fertilization event [...]
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