Role of Endosulfan Dosing in Behavioural Changes and Testicular Toxicity in Swiss Albino Mice

Introduction

Pesticides played significant role in bringing green revolution in India as well as in world. They are widely used to control agricultural pests and pests causing public health problems [1]. It is well established that the indiscriminate use of agrochemicals in modern agriculture not only poses severe health hazards for human beings but also has numerous other side effects on the environment including destruction of the biodiversity.

Endosulfan (Thiodon6,7,8,9,10,10-hexachloro- 1,5,5a,6,9,9a-hexa hydro-6,9-methano-2,4,3- benzodioxathiepin-3 oxide) is an organochlorine compound that is used as an insecticide and acaricide. It has become highly controversial agrochemical due to its acute toxicity, potential for bioaccumulation, and role as an endocrine disruptor. US Environmental Protection Agency (US EPA) classify it as Category-I: “Highly Acutely

Toxic” while WHO keep it in class –II “Moderately Hazardous” list. It is banned in most developed countries e.g., USA, European Union and some developing countries of Asia and Africa but is being still extensively used for control of vectors in public health in Australia, Brazil, India etc [2, 3, 4]. India is World’s largest consumer of endosulfan. It is used in agriculture to control insects, pests, whiteflys, aphids, leaf hopper, colrado; wood preservation, home gardening etc. It is used primarily to kill insects and mites on crops including tea, coffee, cotton, fruits, vegetables, rice and other cereals [5]. Because of its non-specific effect, it can affect beneficial species members also and thus posing a risk to biodiversity. Endosulfan belongs to group of endocrine disrupting chemicals and acts like xenoestrogen. Endocrine disrupting chemicals (EDCs) are a structurally diverse group of compounds that may adversely affect the health of human being, wildlife and fisheries or their progenies, by interaction with the endocrine system (Fossi et al., 2007). It has been suggested that EDCs pose a potential risk and can alter the hormone balance in humans and wildlife (Ishibashi et al., 2001). These environmental xenobiotics may impair the normal embryonic development and disrupt normal reproductive functions in adulthood [6, 7]. It persists in adipose tissues for decades and is biomagnified along food chain [8, 9, 10]. It is acutely neurotoxic to both insects and mammals, including human. It is also a xenoestrogen- a synthetic substance that imitates or enhances the effect of estrogens and can as endocrine disruptor, cause reproductive and developmental damage in both animals and humans. The present experiment aimed to study the effect of endosulfan on overall animal behavior as well as histopathology of testis.

Experimental Design

The experiments were conducted on swiss albino mice (Mus musculus) under controlled conditions of temperature (24± 5 oC), light (alternate 12 hrs light and 12 hrs dark), proper ventilation and cleaning at Mahavir Cancer Sansthan and Research Centre, Patna, India (Reg. 1129/bc/07/CPCSEA). The age group of mice selected for the study was 12 weeks old with 30±2 gm body weight (b.w.).The experiment was conducted in polypropylene cages. Cages of two size, (a) small= 26X19X13 cm and (b) large cage= 40X25X15 cm were used. Endosulfan was supplied orally (gavage method) as ‘endosel’ manufactured by Excel Industries Ltd, Mumbai, India at a concentration of 3.0 mg/Kg b.w. ; for 30 days. During treatment of endosulfan large cages were used, keeping two male and one female mice per cage. The control group of mice was supplied distilled water for drinking. The ‘treatment’ was supplied endosulfan 3mg/Kg b.w. daily by gavage method for five weeks. The variation in behavior from control was recorded daily. The mice were sacrificed after schedule treatment.

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Figure 1a: Light micrograph of normal testis of Swiss albino mice showing well organized seminiferous tubule visible with different stage of spermatogonia, no gap between seminiferous tubules, and proper layer of tunica albuginea.

Figure 1b: Enlarge view of seminiferous tubules showing different stages of spermatogonia, primary and secondary spermatocyte, spermatids and spermatozoa.

Results

The oral administration of endosulfan to the mice for 30 days exhibited behavioral and morphological changes like loss of body weight, fall of body hair, rough and loose skin, restlessness and aggressiveness, budging of eyes which gave treated mice ugly appearance. After 14 DAT, gradual loss of appetite and thrist, frequent rubbing of mouth with forelimbs was observed. After 30 DAT mice became dull and sluggish. Increased incidence of stilted gait, hunched posture, irregular breathing and decreased spontaneous activity in males were also observed. The inner anatomical observations suggested severe damage to histopathology and appreciable modifications in reproductive organ anatomy of male mice (Figure 2 and Figure 3). Sertoli cells of control mice show well organized nucleus. Nuclear membrane was also continuous (Figure 1a,1b). Sertoli cells of five weeks endosulfan administered mice show many osmiophilic granules condensed in cytoplasm. Degenerated tail of spermatozoa was also observed (Figure 2). Deshaped nucleus and degenerated mitochondria were clearly observed with many vacuolated spaces (Figure 3). Degenerated plasma membrane of sertoli cells was observed. Nourishing spermatozoa were clearly observed with degenerated acrosome.

Light micrograph showing longitudinal section of testis of endosulphan at 3mg/Kg body weight/day for 30 DAT

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Discussion

The degradation of seminiferous tubules, scattering of spermatogonia were observed. The number of sperms decreases in somniferous tubules. The degradation of wall of somniferous tubules was also observed. The adverse effects of endosulfan on semniferous tubules are similar to earlier reports [1, 11, 12, 13]. This may be due to degradation of sertoli cells which are principal structural element of the semniferous epithelium and provide physical support and an environment conducive to germ cell development and maturation [14].

Wall of seminiferous tubule is also degraded.

Conclusion

Endosulfan is important pesticide exclusively used around the globe. It can cause behavioural changes as well as testicular toxicity in mammals. The judicious use of Endosulfan is highly recommended.

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