Histopathological Alteration Due to the Effect of Lead Nitrate on the Testis of Freshwater Fish, Channa Striatus (Bloch, 1793)

Introduction

At present a large amount of metals are introduced through human activities into the natural environment. Globally, heavy metal pollution is becoming a serious issue of increasing concern to the survival of living organisms, including fish. Cadmium, mercury and lead are toxic to most aquatic organisms even in low concentrations¹ that produce histological alterations including liver damage,² respiratory dysfunctions,³ testicular and ovarian alterations.^4,5,6 From recent few years, lead has gained considerable attention among the scientific community due to its potential human health hazards.⁷ Popularity of lead is due to its dense ductile, highly malleable and corrosion resistant properties⁸ as it is used in manufacturing of storage batteries, paints, dyes, pigments, coloured inks, pipes, soldering materials, cement manufacturing, mining, smelting, etc.⁹ Histopathological studies are being widely used as reliable biomarkers for determining the health of the contaminants exposed fishes.^10,11 These pathological studies indicated the damage of cell membranes which increases the injury to more critical targets.¹² Lead is also known to accumulate in different tissues in fishes.¹³ Reproductive impact of lead exposure is spreading worldwide¹⁴ almost affecting all characteristic of reproduction.¹⁵ Therefore, the present study was carried out to investigate the testicular histopathological alterations in Channa striatus, induced by chronic exposure to lead nitrate.

Materials and Methods

Live specimen of fish, Channa striatus (average length 20-25 cm and weight 50-60 gms) were collected from different fish markets of Bhopal, Madhya Pradesh for experimentation. Fishes were divided into four groups as Group I, Group II, Group III and Group IV, and were exposed to sub lethal concentrations (0, 8, 18 and 28 mg/l, respectively) of lead nitrate (Ranbaxy India Ltd.) for a total period of 90 days. Exposure concentration was decided on the basis of 96 hrs LC₅₀ value of lead nitrate. LC₅₀ values at 95% confidence limits were calculated by using the software “Trimmed Spearman Karber method”, version-1.5¹⁶ for different exposure period. To maintain apposite lead nitrate concentration throughout the experimentation duration of 90 days, water of each aquarium was changed on every alternate day. At regular interval of 30 days (at the end of 30^th, 60^th and 90^th days of experimentation), fishes from each groups were harvested, and the testis of all the four groups were removed aseptically. Tissues were preserved in aqueous Bouin’s fixative for 48 to 72 hours.¹⁷ After fixation, 5-6 μm thick sections were cut with the help of rotatory microtome and stained with Ehrlich’s haematoxylin and eosin (H&E) for histopathological examination.

Results

Histological sections of the testis of the experimental fishes are shown in Fig. 1 to Fig. 10. Histological sections of testis of control fish (Group I) showed typical histological picture similar to that of a normal fish, characterised by a number of seminiferous tubules irregularly arranged inside and spermatozoa in the germinal epithelium (Fig. 1). In case of Group II fishes, damaged walls of seminiferous tubules and indistinguishable leydig’s cells on 30^th day (Fig. 2), disorganization of seminiferous tubules, vacuolization and indistinguishable leydig’s cells on 60^th day of lead nitrate exposure (Fig. 3), while disorganization of seminiferous tubules, deshaped seminiferous tubules, necrosis and centralization of spermatozoa on the 90^th days of exposure (Fig. 4) were observed. Deshaped and damaged seminiferous tubules, clumping of seminiferous tubules, necrosis and indistinguishable leydig’s cells on 30^th day (Fig. 5), while deshaped seminiferous tubules and necrosis on 60^th day (Fig. 6) and damaged walls of seminiferous tubules, centralization of spermatozoa and disappearance in the number of leydig’s cells on the 90^th day of lead nitrate exposure (Fig. 7) were observed in case of Group III fishes. In case of Group IV fishes, the common histoarchitectural alterations observed on the 30^th day of lead nitrate exposure were clustering of seminiferous tubules, disorganization of seminiferous tubules, damaged walls of seminiferous tubules and centralization of spermatozoa (Fig. 8). The alterations were more severe as disorganization of seminiferous tubules, deshaped and damaged walls of seminiferous tubules, centralization of spermatozoa, severe reduction in the number of spermatozoa and indistinguishable leydig’s cells on the 60^th and 90^th days (Fig. 9 & 10, resp.) were observed.

Figure 1: Testis structure of control fish: showing seminiferous tubules (ST) and spermatozoa (SZ) X 400. Click here to View figure

Figure 2: Testis of fish exposed to 8 mg/l Lead nitrate for 30 days showing damaged walls of seminiferous tubules (DWST) X 400.  Click here to View Figure

Figure 3: Testis of fish exposed to 8 mg/l Lead nitrate for 60 days showing disorganization of seminiferous tubules (DOST) and vacuolization (V) X 400. Click here to View Figure

Figure 4: Testis of fish exposed to 8 mg/l Lead nitrate for 90 days showing disorganization of seminiferous tubules (DOST), deshaped seminiferous tubules (DSST), necrosis (N) and centralization of spermatozoa (CSZ) X 400. Click here to View Figure

Figure 5: Testis of fish exposed to 18 mg/l Lead nitrate for 30 days showing damaged seminiferous tubules (DST) and necrosis (N) X 400. Click here to View Figure

Figure 6: Testis of fish exposed to 18 mg/l Lead nitrate for 60 days showing deshaped seminiferous tubules (DSST) and necrosis (N) X 400. Click here to View Figure

Figure 7: Testis of fish exposed to 18 mg/l Lead nitrate for 90 days showing damaged walls of seminiferous tubules (DWST) and centralization of spermatozoa (CSZ) X 400. Click here to View Figure

Figure 8: Testis of fish exposed to 28 mg/l Lead nitrate for 30 days showing disorganization of seminiferous tubules (DOST), damaged walls of seminiferous tubules (DWST) and centralization of spermatozoa (CSZ) X 400. Click here to View Figure

Figure 9: Testis of fish exposed to 28 mg/l Lead nitrate for 60 days showing disorganization of seminiferous tubules (DOST), damaged walls of seminiferous tubules (DWST), deshaped seminiferous tubules (DSST) and centralization of spermatozoa (CSZ) X 400. Click here to View Figure

Figure 10: Testis of fish exposed to 28 mg/l Lead nitrate for 90 days showing centralization of spermatozoa (CSZ) and reduction in the number of spermatozoa (RSZ) X 400. Click here to View Figure

Discussion

Similar results were seen by¹⁸ in case of Lepomis macrochirus exposed to endosulfan for 24, 48, 72, 96 hr, 1 and 2 week periods. The histopathological alterations in testes of fish showed connective tissue breakage, dilation of lumens, reduction in the size of primary spermatogonia, breakage of primary spermatocyte walls, greater increase in the size of seminiferous tubules and lumens, damaged sertoli cells and spermatids, damaged primary spermatogonia, decrease in the number of spermatozoa and spermatids, disruption of seminiferous tubule walls and disorganised structure of the testis. Cytolysis, vacuolation in seminiferous tubules, disorganized testicular architecture and extensive damage to germinal epithelium, necrotic cells and connective tissues in the testis of air-breathing fish, Heteropneustes fossilis due to the exposure of cadmium chloride for 45 days was also reported by.¹⁹ Loosening and indistinguishable arrangement of tissue, disorganization of the lobules, disintegration of the spermatogonia and interstitial tissue, epithelial vacuolization, disorganization of germ cells, reduction in the number of spermatogonia in the testis of African catfish Clarias gariepinus exposed to 0.05 mg/l of mercuric chloride for 60 days was observed by.²⁰ It is construed from our study that the testicular histopathological alterations increased in proportion to the increase of lead nitrate concentration and length of period of intoxication in Channa striatus.

Conclusion

From the present study it is concluded that the extent of histological changes in the testis of Channa striatus not only depends on the concentration of lead nitrate but also on the duration of exposure.

Acknowledgements

The authors are thankful to Head, Department of Zoology and Applied Aquaculture, Barkatullah University, Bhopal (M.P) India, for providing necessary laboratory facilities to carry out the present work successfully.

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