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International Journal of Morphology

On-line version ISSN 0717-9502

Int. J. Morphol. vol.21 no.3 Temuco  2003

http://dx.doi.org/10.4067/S0717-95022003000300011 

Int. J. Morphol., 21(3):251-255, 2003.

Octopus mimus (MOLLUSCA: CEPHALOPODA) EMBRYO MISDEVELOPMENT DUE
TO CHRONIC EXPOSURE TO THE ORGANOSPHOSPHORUS PESTICICE PARATHON

DESARROLLO EMBRIONARIO ANÓMALO EN Octopus mimus (MOLLUSCA: CEPHALOPODA)
DEBIDO A EXPOSICIÓN CRÓNICA AL PESTICIDA ORGANOFOSFORADO PARATHION
*Jorge L. Gutiérrez-Pajares; **Alberto Olivares & ***Eduardo Bustos-Obregón


GUTIÉRREZ-PAJARES, J. L.; OLIVARES, A. & BUSTOS-OBREGÓN, E. Octopus mimus (Mollusca: Cephalopoda) embryo misdevelopment due to chronic exposure to the organophosphorus pesticide Parathion. Int. J. Morphol., 21(3):251-255, 2003.

SUMMARY: Pesticides are of ample worldwide use. At present there is a growing concern about seawater contamination by these chemicals. Developing aquatic organisms are particularly at risk. For this reason, early development of Octopus mimus embryos under exposure to commercial Parathion was examined.

Concentrations over 0.4 mM of the pesticide altered formation of the embryonic disc at the blastula stage, resulting in abnormal gastrulation and arrest of further development. This toxic effect may be due to the blockade of DNA synthesis elicited by organophosphoric pesticides causing diminished number of blastodermic cells at the time of epiboly, thus hindering the start of invagination. Seawater contamination seems, therefore, of relevance for survival of exposed aquatic species.

KEY WORDS: 1. Octopus mimus; 2. Parathion; 3. Early embryo development 


INTRODUCTION

Pesticides are used for many purposes through out the world. In aquaculture some treatments have been introduced to control infestations such as sea lice (Copepoda: Caligidae). These treatments include formaldehyde, organophosphates (OPPs) and hydrogen peroxide (Roth et al., 1993). In addition, many pesticides that could drain into seawater have been introduced to the environment due to human activities, like agriculture.

Among pesticides, OPPs are commonly used because of their less bioaccumulative properties compared to organochlorine pesticides. The increasing environmental concern with regard to these compounds has determined the development of high-resolution techniques to detect this kind of pesticide (Albanis & Hela, 1995; Okumura & Nishikawa, 1995; Lambropoulou & Albanis, 2001). In fact, OPPs have been found in groundwater, surface waters, lagoons and drinking water (Driss et al., 1993). In Chile, 0.1 mg/L of OPPs have been found in seawater (D. G. T. M. y M. M., 1994).

Currently, there is a growing concern about searching for bioindicators of pesticide contamination and the effects of pesticides on non-target organisms in aquatic environments. Measurements of hemolymph cholinesterase activities in a tropical scallop have been suggested to assess seasonal, sublethal pesticide exposure (Owen et al., 2002). Recently, a research group from Japan suggested that common squid could be considered as a suitable indicator for monitoring pesticides pollution in water (Ueno et al., 2003). High concentration of organochlorines have been found in eels and clams in Moulay Bousselham lagoon (Mehdaoui et al., 2000). High concentration of a pesticide residue present in the Pensacola estuary caused pathological changes at tissular and cellular levels in oysters, spot and shrimp (Nimmo et al., 1975). Moreover organochlorine pesticides have been detected in marine mammals (Borrel & Aguilar, 1999).

Endpoints traditionally measured for adults may provide little prediction of the effects of pesticides on invertebrate embryo development in marine environment. Sea urchin embryo development has been used to monitor the quality of seawater (Quiniou et al., 1999) and to investigate toxic effects of methoxychlor (Green et al., 1997). Nevertheless, embryos from other species could be also exposed to OPPs. In that sense, the aim of the present study was to characterize the effects of Parathion on early embryo development of Octopus mimus, an important economic resource in the II Region of Chile.

MATERIAL AND METHOD

Species: Octopus mimus is present mainly in the North of Chile and can be easily cultured in laboratory under supervised conditions (Cortez et al., 1999).

Reproduction of Octopus mimus could occur in any month of the year. However, its reproduction increases in spring and summer, which may be related to a longer light photoperiod more than higher water temperature (Zúñiga et al., 1995). Clusters of eggs are deposited in the walls of the female's shelter in captivity. Octopus embryo development, comprising the period between fertilization and hatching, lasts 50 days approximately but depends strongly on water temperature in an inverse way. (Castro et al., 2002; Caverivière et al., 1999).

Experimental procedure: Egg masses of Octopus mimus, obtained from a brood stock maintained as described in Castro et al. were used for the rearing experiments conducted in filtered artificial seawater. Eggs were incubated at 17.5 - 19C with constant aeration. Commercial Parathion (cP), 80% w/v (Anasac, Santiago, Chile), was added to obtain 0, 0.2, 0.4 and 0.8 mM concentrations in seawater. Four time intervals of exposure (47, 96, 120 and 312 hours) were used. Seawater (control and treated groups) was renewed every 48 hours. Each cP treatment group included 70 - 100 embryos.

Embryos were fixed with glutaldehyde and then examined and photographed under light microscopy. Embryos were considered abnormal when a clear evidence of delayed development was observed.

RESULTS AND DISCUSSION

Continuous economical and industrial development has contributed to increase social benefits and quality of human beings. However, detrimental effects on the environment where human activities take place have also increased in a parallel way. Coastal zones should have special consideration because multiple technologies have been established thereby such as ports and fisheries. In addition, wastage water is usually sent into seawater. All these produce changes in biological communities whose habitats are inter- and sub tidal zones, marine sediments and water column (CPPS-PNUMA, 1999).

Octopus embryo development is clearly affected by chronic exposure to cP in seawater in a dose dependent way as shown in Fig. 1. Octopus egg is telolecytic and surrounded by a thick chorion. The animal pole, from which the embryo develops, is very small. During blastula formation, the blastodermic cells decrease in size and form an embryonic disc in the animal pole (Fig. 2A). This normal pattern, as described in detail by Castro et al., is altered by addition of cP to the seawater in concentrations greater than 0.4 mM (Fig. 2B).


Fig. 1. Percentage of abnormal octopus embryos chronically exposed to Parathion at different time intervals.

Gastrulation requires the migration of blastodermic cells over the yolk in the process of epiboly (Fig. 2C), as documented in eggs of Tremoctopus violaceus (Boletzky, 1988). This process is completely abolished in embryos treated with the higher dose of cP (Fig. 2D).


47 h
96 h
120 h
312 h
Fig. 2. Octopus mimus embryo development. The white triangle in C shows blastodermic cells migration during gastrulation. In H bar indicates 500 µm.

Embryo developmental arrest (Fig. 2F) may be due to cP interruption of cell proliferation. Rodriguez & Bustos-Obregón (2000) in in vitro culture of mouse seminiferous tubules observed decreased DNA synthesis as effect of OPPs. In accordance with this, Pesando et al. (2003) found that OPPs slowed the rate of early mitotic cycles, affected nuclear and cytoskeletal status and DNA synthesis in sea urchin embryos before gastrulation. On the other hand, the lower cP dose used in this study delayed embryo development (Fig. 2H) compared to control (Fig. 2G). This could be explained by a partial lack of epibolic displacement accounted for a decreased number of blastodermic cells available at the time of gastrulation.

OPPs are known inhibitors of the active sites of acetylcholinesterase (AChE) and some reports show the presence of this enzyme in mollusks. AChE has been demonstrated by cytochemical methods in the heart of Sepia officinalis L. (Cephalopoda) (Kling, 1986). In another sea mollusk, AChE has been detected in cholinergic and non-cholinergic neurons and in hemolymph. In this species Paraoxon (derivative of Parathion by oxidation) markedly reduced the growth of axons and neuronal survival in culture (Srivatsan, 1999). Even though it is not known at present if AChE plays a role in mollusk early embryo development, prior to neuronal stage, acetylcholine receptor has been detected in sea urchin oocytes and early embryos which regulate intracellular calcium (Buznikov & Rakich, 2000). Thus, the classical way of cP toxicity cannot be currently excluded in the early embryo of this species.

Pesticides exert deleterious effects on different embryos in aquatic habitats. Gorge & Nagel (1990) found abnormalities during embryo development and hatching and also mortality of fertilized zebra fish eggs exposed for 35 days to lindane, atrazine or deltamethrin. Parathion has the same effect in this species (Lara et al., 2002). As found in the present work, methoxyclor elicited delayed segmentation and abnormal gastrulation of fertilized sea urchin eggs (Green et al.).

Seawater contamination by OPPs remains as a relevant subject in terms of survival of marine species (Badawy & el-Dib, 1984a), which are exposed to accidental spill of these toxics (Badawy et al., 1984b).

ACKNOWLEDGMENTS

We would like to thank University of Chile (DID) for the financial support of this research. J.L. G.-P. was a fellow from Programa Latino-americano para Capacitación en Reproducción Humana (PLACIRH), PLC-239/2000. 


GUTIÉRREZ-PAJARES, J. L.; OLIVARES, A. & BUSTOS-OBREGÓN, E. Desarrollo embrionario anómalo en Octopus mimus (Mollusca:Cephalopoda) debido a exposición crónica al pesticida organofosforado Parathion. Int. J. Morphol., 21(3):251-255, 2003.

RESUMEN: Los pesticidas son de amplio uso mundial. Actualmente hay gran preocupación por la contaminación marina con estas sustancias. El desarrollo de los organismos acuáticos está especialmente comprometido. Por esta razón, se estudió el desarrollo temprano de embriones de Octopus mimus expuestos a Parathión comercial.

Se encontró que concentraciones sobre 0.4 mM del pesticida alteran la formación del disco embrionario al estado de blástula resultando en una gastrulación anormal y detención posterior del desarrollo.

Este efecto tóxico puede ser debido al bloqueo de la síntesis de ADN resultando en una disminución del número de células blastodérmicas al momento de la epibolia, y por tanto impiden el inicio de la invaginación. De esta manera, la contaminación marina parece ser de importancia para la sobrevida de las especies acuáticas expuestas a ella.

PALABRAS CLAVE: 1.Octopus mimus; 2. Parathión; 3. Desarrollo embrionario temprano. 


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Correspondence to:
Prof. Dr. Eduardo Bustos-Obregón
Laboratorio de Biología de la Reproducción
Facultad de Medicina. Universidad de Chile.
Independencia 1027.
Santiago - CHILE

E-mail: ebustos@machi.med.uchile.cl

Received : 15-07-2003
Acepted : 21-08-2003 


* Unit of Reproduction. Faculty of Biological Sciences. P. Catholic University of Chile.
** Department of Aquaculture. University of Antofagasta.
*** Laboratory of Biology of Reproduction (ICBM). Faculty of Medicine. University of Chile.

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