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6. zoomedvet URL: http://www.zoomedvet.ru/articles/lechenie/azoksivet-kompleksnyy-vysokotekhnologich-nyy-immunomodulyator/
7. URL: http://www.vet.petrovax.ru
DOI: http://dx.doi.org/10.20534/ELBLS-17-2-19-22
Ismat S. Ahmadov, Department of Chemical Physics of Nanomaterials, Faculty Physics, Nargiz J. Agayeva, Narmina A. Sadigova, Department of Bioecology, Faculty of Biology, Baku State University, Baku, Azerbaijan E-mail: [email protected]
The impact of nanoparticles on the embrional and postembrional development in molluscs Lymnaea Auricularia
Abstract: In this article has been studied toxic effects of some nanoparticles on the embrional and postembrional development of mollusks. Experiments were carried out on Lymnaea auricularia mollusks. It was concluded, the embryonic and postembryonic stages of development of mollusks are very sensitive against nanoparticles.
Keywords: Food chain, mollusks, hatchability, egg clusters, embryo, nanoparticles, nanotoxicology.
1. Introduction. Using as model organisms, the mol- by this way. The adopting effect was 83% and absorption lusks living both in marine and fresh waters, the toxic influences of nanoparticles are comparatively investigated in them and the main aim in these studies is to determine the risk of nanomaterials. Based on reviews of scientific articles, there are no standart experimental approaches to determine the toxicity of nanoparticles and it needs a lot of experimental results in their organization. Generally it is known that, the main organs accumulated in water organisms of nanoparticles are glands of digestive system and their cells. The main targets of nanoparticles are the endosomal-lizosomal system and mitochondrias. Nanoparticles both directly and by the products are separated from them, create immunetoxicity, oxadizing stress and damage cell proteins, biologycal membranes and DNA [1]. Maria Noyel and her colleagues (2014) studied the bioaccumulation of copper oxide nanoparticles ( 65CuO) which were modified with isotope, by keeping in water and feeding its mollusk Lymnaea stag-nalis living in freshwater basin. They identified that, the mollusks effectively adopt CuO nanoparticles included
speed 0.61 g g-1 d-1 at concentration of CuO nanoparticles (< 100 nmol g-). At higher values of concentration the effectiveness decreased to 50%. It is interesting that, in a day CuO nanoparticles were cleaned up from snail's body. It has been identified that, TiO nanoparticles at > 1.0 mg/kg concentrations stimulate the immune system in Octopus vulgaris (Mollusca: Cephalopoda) snail. In 4 hours after nanoparticle injection the increase of circulating hemosit number, lizosome activity, the concentration of nitrogen oxide were observed. Return to these parameters norm happened in 24 hours [2]. In order to identify the influences of biotransformation of nanoparticles in marine ecosystems Milca O. Montes and his colleagues (2012) fed the median concentrations of Mytilus galloprovincialis with two various nanometals — CeO2 and ZnO being in 1 mg L-1 to 10 mg L-1 interval in laboratory condition. At 10 mg L-1 concentrations medians adopted 62 ^g L-1 Ce and 880 ^g L-1 Zn-calculated in dry weight of tissue. By Electron Scan Microscope it was determined that CeO2 nanoparticles remained in median
body, but ZnO nanoparticles were removed [2]. Based on these results of experiments it becomes clear that nanoparticles can make serious toxic effects in mollusks being important components of water ecosystems and in other invertebrates. Therefore to learn the exposing results of mollusks participating in food chain of nanoparticles is one of the actual problems. Taking this idea in mind the present investigation was undertaken to study the sequential events of the embryonic and postembryonic development of L. auricularia which were developed in solutions of nanoparticles.
2. Materials and methods. Experiments were carried out in Lymnaea auricularia mollusks — being the second component of simple food chain (phytoplank-ton-mollusk-fish) existing in water ecosystems. L. auricularia is found in freshwater lakes, ponds, and slow-moving
rivers with mud bottoms [3]. L.auricularia are the pulmonary breathing gastropod mollusks widely spread in still water basins. Water temperature must be ~19 0C, pH 6-7,1 for their survival and proliferation. These species live in hard water in Great Britain. The species can't live in polysaprobic waters much contaminated areas by higher organic substances, sulphides, bacterias, and lack of oxygen. L.auricularia are fed with plants and various detrits. The body sizes of Lymnaea auricularia depend on the development of their pelvises. Sometimes the height of the pelvis can develop up to 30 mm and width up to 25 mm, and the ratio of width to length is more than 0,75. But most species grow up to half of the maximum dimension in breeding. The width of the pelvis is 12-18 mm, and height 14-24 mm.
Figure 1. Lymnaea auriculara eggs capsules
Their legs are nearly 18x11 mm. As the sizes of neurons in L.auricularia nervous system relatively bigger, they are used as the model object to study the functionalizing of animal nervous system in neurophysiology. L.auricularia are hermaphrodite, insemination is cross, development is straight and embryo development happens inside of egg membrane till the end. The eggs of L.auricularia mollusks are inside of the longish shape transparent, helmes slime mass.The amount of eggs in slime mass can be changed depending on the types of mollusks, age, physiological state, environmental food richness, temperature. They put their eggs as a capsules from 50 to 210 (Figure 1). In experiments the slime masses (complex with eggs) of egg clusters ofmollusks have been chosen. The mollusks were distributed into glasswares of 25 mm 3 volume. In a day we observed the mollusks put eggs inside the slime mass.
The egg clusters using for the experiments were chosen, and were put into glassware of 25 mm 3 volume in room temperature. In control approximately in 14-15 days we observed the mollusk babies getting out of the most eggs inside the slime mass.Though the get-
ting out of mollusk babies from the slime mass in test began in 14-15 days, appeared young snails were few compared to control. The appearing of young snails in test eggs, kept in solution which were added nanopar-ticles with various concentrations was compared to control variant. Embryonic and postembryonic development of mollusks was taken and morphological changes occurring in them were watched.
3. Results. It was interesting to know, in what period of development the influence of nanoparticles on the snails is strictly in embryonic or post-embryonic? In order to find out the answers of these questions, the egg capsules have been kept in the solution of nanopar-ticles and the development of the embryos has been observed in all period of development. On the first day of experiments were taken two days lading ofegg capsules as a control and they were in discrete form, the eggs were in spherical form and embryos are seeing as points. On the 7th day of development, the miniature snail which was possessed in all the structures found in a newly hatched eggs and its spinning was observed. On the tenth day of
the development a very slow jerking movement of the miniature snails is observed and the eyes are apparent.
Figure 2. The embrional and postembrional
On the 11, 12, 13-th days of development of snails have been seen normal and structured mouth, the velum and head. On the 14th days of development the snail moved freely in the jelly of the egg capsule, outside the egg. On the 15 th day of the development 90% of snails were moved freely in the glass beaker and have already escaped out of an egg. On the 23, 24, 25-th days of development, young snails are feeding and their excrements were observed in glass beaker. During 63 days of development of the snails can be observed the feeding, free movement, growth of the young snails. The results of experiment have given in Figure 2.
The egg clumps of mollusks were exposed to the solutions of the nanoparticles with the various concentrations (0,001%, 0,005%, 0,01%). In the first variant of experiments the eggs of snails were
development of molluscs in normal conditions
exposed to the solution of Fe3O4 with 0,001% concentration. A few hours after the exposition some changes are observed in the laid egg capsules, the snails periodically thrust its foots against the wall of egg and thus the shape of egg elongates. On the 7-th day of the exposition of eggs it became evident that the development of the embryos in the egg has happened differently compared with control. On the 10-th day of exposition in the solutions of nanoparticles the development of embryo in two of eggs (3,6%) has ceased completely, the development in 4 eggs (7,3%) fell behind than others, and their sizes are small compared with control. On the 15-th day of the exposition there is not any improvement in the eggs which the process of development was delayed, only from two eggs (3,6%) hatched molluscs.
Figure 3. The development of egg snails exposed in the solution of Fe3O4 nanoparticles with 0,001% concentration
On the 23-rd day of experiment one of the young 24, 25-th days mass death of young snails was observed, snails died, 3 were in the deadly state, heart beating, and the bodies of dead ones were outside of the shell. On feeding, weakness in movement were watched. On the the other side, there was no improvement in the embryo
state, delayed from the development inside the slime mass in eggs. On other days death among the young snails also continued, and the bodies of the individuals remained outside the shell. On the 63-rd day of experiment the young snails both in eggs and outside the eggs died. As the result of these experiments we can say that in test 12,73% of eggs remained fully undeveloped, and 87,27% of developped ones all died within 63 days. For the comparison it must be noted that in control young snails remained alive within the same day. The results of the experiments were presented in figure 3.
4. Discussion. There are different approaches for the determination toxicity of nanoparticles on aquatic organism. Understanding the toxic effects of nanoparticles on aquatic organism allows to evaluate the risk of the spread in the food chain of water ecosystems. The last review of research articles in the area of nanotoxicology indicates that the aquatic invertebrate
testing will be key in the evaluation of this risk [4; 5; 6; 7; 8;]. The most frequently tested engineered nanoparticles in invertebrate tests are C (60), carbon nanotubes, and titanium dioxide. As the test organism, in the majority of the studies have been used Daphnia magna. The results of these studies have indicated that acute toxicity takes place at the low concentration range of engineered nanoparticles, although some indications of chronic toxicity and behavioral changes of aquatic invertebrates have also been described at concentrations in the high microgram/L-1 range [8].
Conclusion
Based on the results of this study it can be concluded, the embrional and postembrional stages of development of mollusks Lymnaea auricularia are very sensitive against nanoparticles expozitions. The toxic effects of nanoparticles on the L.auricularia depend on their concentrations and exposure time of nanoparticles.
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