Titikksha Das and Mamata Goswami
INTRODUCTION
Contamination of water by arsenic compounds and its
toxicological effect on aquatic organism is a major world-
wide problem. Geogenic processes and anthropogenic dis-
turbances are the two main causes of dispersal of arsenic
in aquatic environment (Bears et. al., 2006; Gonazalez et.
al., 2006). Several countries including Argentina, Bangla-
desh, Chile, China, India, Japan, Mexico, Mongolia, Nepal,
Poland, Taiwan, Vietnam, and some part of United States
have been reported with high concentration of arsenic in
groundwater (Anowar et. al., 2002; Mitra et. al., 2002;
Smith et. al., 2001; Chowdhury et. al., 2000). A correlation
has been found between chronic arsenic poisoning and
many health effects including cancers, melanosis, hyper-
keratosis, restrictive lung disease, peripheral vascular dis-
ease, gangrene in leg, skin, lung, bladder, liver, diabetes
mellitus, hypertension and ischemic heart disease (Ana-
war et. al., 2002). It is evident that arsenic exposure has
multiple effects at the molecular level for instance liver
chromosomal DNA fragmentation, expression of certain
proteins, differential expression of genes involved in cell
cycle regulation, signal transduction, stress response,
apoptosis, cytokine production, growth-factor and hor-
mone-receptor production (Hossain et. al., 2003; Tabellini
et. al., 2005; Ahmed et. al., 2008; Sangeeta et. al., 2012
Paruruckumani et al., 2015).
Both in laboratory and eld studies histopathologi-
cal investigations have been long recognised as reli-
able biomarkers of stress in sh and in the evaluation
of the health of sh exposed to contaminants. The gills,
liver and kidney are the common primary target organs
for many chemicals primarily because of their vital
role within the body (Chowdhury et. al., 2000; Hossain
et. al., 2000, Paruruckumani et al., 2015).
In this work, we studied the toxicity effect of sodium
arsenite in the gill and liver tissues of fresh water sh
Channa punctatus and for the rst time observed the his-
topathological as well as surface ultrastructural changes
on it. We also estimated a critical value of concentration
of sodium arsenite above which shes are likely to be
killed. A commonly useful measure of toxicity LC50 is
used for this purpose. The goal of this study was, rstly,
to observe any histological changes, arsenic could bring
to the vital organs of living animal and secondly, to sub-
stantiate the role of arsenic as a toxic environmental
agent which can cause many severe health effects.
MATERIALS AND METHODS
For the present study healthy and disease free shes
Channa punctatus (weight 22-50 gm) were collected from
local markets in Guwahati. After disinfection with a dip of
2% potassium permanganate (KMnO
4
) solution the shes
were acclimatised in aquaria for two weeks before initia-
tion of experiment. The water provided in the aquaria was
from the tap water in the laboratory and was changed
on the following day. The shes were fed everyday with
sh food available in the market. Proper aeration was
done during these periods. Sodium Arsenite (NaAsO
2
),
molecular weight-129.91 Merck, India (Ltd.) was procured
for performing the experiment. A stock solution was pre-
pared with water from which the test concentration was
prepared by dilution. The control group of shes were
kept in similar conditions without adding sodium arsen-
ite. Fishes were exposed to 5 different concentration of
Sodium Arsenite of 5, 15, 25, 35 and 45 ppm. The toxicity
bioassay was performed in semi-static system in triplicate
with 10 specimens exposed for each concentration in each
set in accordance with the standard methods of acute tox-
icity bioassay procedures (APHA, 2005).
Fishes were transferred to each aquarium and exposed
to ve different concentrations such as 5, 15, 25, 35 and
45 ppm of sodium arsenite. In all cases, control groups
of shes were maintained. Each experimental trial was
carried out for a period of 96 hours. The mortality rate of
the sh was recorded at logarithmic time intervals that
is, after 6, 12, 24, 48, 72 and 96 hours of exposure. The
test media was renewed daily during the experimental
period. The data obtained in course of the investigation
were analysed statistically to see whether there is any
in uence of different treatment concentrations on the
mortality of the sh. Fishes were exposed to sub lethal
concentration i.e. 12ppm of sodium arsenite along with
a control group for 96 hours. At the end of the exposure
period, shes were randomly selected for histopatho-
logical examinations. Gill, liver, tissues were isolated
from normal and experimental sh. Physiological saline
solution (0.75% NaCl) was used to rinse and clean the
tissue. They were xed in aqueous Bouins solution for
24 hours, processed through graded series of alcohols,
cleared in xylene and embedded in paraf n wax. Sec-
tions were cut at 4 micron thickness and stained with
Hematoxylin and eosin stain. Histopathological lesions
were examined and photographed with the help of com-
puter attached Bright Field Microscope (Leica DM 3000).
Gills and liver tissues of both the control and treated
groups were rapidly removed and processed routinely
for scanning electron microscopic studies. Gills and liver
tissues were cut into small pieces of 1 mm thickness and
xed in 2.5 % glutaraldehyde prepared in cacodylate
(sodium phosphate) buffer adjusted to pH 7.4 for 24
hours and afterward washed in phosphate buffer for 15
min. After dehydration in ascending series of acetone,
samples were immersed in Tetra Methyl Silane for 10
minutes at 4 degree centrigrate. Then they were brought
to room temperature to dry. The specimens were mounted
on Aluminium Stubs coated with gold and observed
BIOSCIENCE BIOTECHNOLOGY RESEARCH COMMUNICATIONS HISTOPATHOLOGICAL AND ULTRASTRUCTURAL CHANGES IN THE GILL AND LIVER 435