Environmental Toxicology and Pharmacology 29 (2010) 4449

Contents lists available at ScienceDirect

Environmental Toxicology and Pharmacology

journal homepage: www.elsevier.com/locate/etap

Inuence of cypermethrin toxicity on ionic regulation and gill Na+ /K+ -ATPase
activity of a freshwater teleost sh Cyprinus carpio
L. Suvetha, M. Ramesh , M. Saravanan
Unit of Toxicology, Department of Zoology, School of Life Sciences, Bharathiar University, Coimbatore 641046, Tamil Nadu, India

a r t i c l e i n f o a b s t r a c t

Article history: The effects of acute and sublethal toxicity of cypermethrin, a synthetic pyrethroid insecticide on plasma
Received 3 December 2008 electrolytes (Na+ , K+ and Cl ) levels and gill Na+ /K+ -ATPase activity in a common carp Cyprinus carpio
Received in revised form were examined. The 24 h LC50 value of cypermethrin for C. carpio was 1.86 ppm. During acute exposure,
14 September 2009
cypermethrin caused adverse effects in the form of hyponatreima, hypokalemia and hypochloremia and
Accepted 20 September 2009
inhibition of gill Na+ /K+ -ATPase activity. In sublethal exposure to cypermethrin (0.186 ppm for 35 days),
Available online 26 September 2009
plasma Na+ was decreased throughout the exposure period except 7th day whereas plasma K+ level
was increased up to 28th day, then declined. However, plasma Cl level was decreased. Likewise, gill
Keywords:
Cypermethrin
Na+ /K+ -ATPase activity was decreased as the exposure period extended. The present study indicates that
Electrolytes cypermethrin was highly toxic to freshwater sh and ion levels in blood as measured by specic ion
Enzyme activity concentrations (Na+ , K+ and Cl ) and changes in gill Na+ /K+ -ATPase activity may represent a sensitive
Cyprinus carpio and useful nonspecic biomarkers of chemical exposure and effects.
Acute and sublethal toxicity 2009 Elsevier B.V. All rights reserved.

1. Introduction Cypermethrin, -cyano-3-phenoxybenzyl-3,2,(2-dimethyl 2-2-

Due to rapid industrialization, application of large number of
pesticides to control pests and uses of synthetic fertilizers, freshwa-
ter resources are easily polluted and the water quality is unsuitable
for drinking and irrigation purposes which in turn affect the aquatic
fauna. Among the various kinds of pesticides used in India, synthetic
pyrethroid pesticides have largely used by the farmers due to their
lower mammalian toxicity when compared to other organophos-
phorus and organochlorine pesticides. Unfortunately, application
of these synthetic derivatives of pyrethrins is highly toxic to a num-
ber of non-target organisms such as bees, freshwater sh and other
aquatic organisms even at very low concentration (Oudou et al.,
2004; Begum, 2005; El-Sayed et al., 2007). Indiscriminate discharge
of these pesticides from agricultural run off and in aquaculture
operation may be washed into nearby water bodies and affects
non-target organisms such as sh and prawn which are of great
economics importance to humans (Adhikari et al., 2004). Among
the aquatic animals sh are highly sensitive to the pyrethroids pes-
ticides due to their neurotoxic effects and the pesticides are lethal
to sh at a minimum concentration (101000 lower) than the cor-
responding values for other groups of mammals and birds (Koprucu
and Aydin, 2004).
Corresponding author. Tel.: +91 422 2422222; fax: +91 422 2422387.
E-mail address: mathanramesh@yahoo.com (M. Ramesh).
dichlorovinyl)-2,2-cyclopropane carboxylate, a synthetic pyre-
throid widely used to control cotton boll worm (Heliothis armigera)
and to eradicate the larvae of mosquitoes and milk shes dur-
ing pond preparation in urban and agricultural environments
(David et al., 2004; Collins and Cappello, 2006). As per USEPA
cypermethrin is a type of cyanophenoxybenzyl pyrethroid and is
categorized as restricted because of its high toxicity to sh (ETN,
1996). The presence of low levels of cypermethrin in the aquatic
environment adversely affects the populations of Atlantic salmon
by disturbing the reproductive functions (Moore and Waring,
2001).
Fish in general are exposed to pyrethroids through their gills,
which are multifunctional and complex organs with which they
make intimate contact with their ambient water (Wendelaar Bonga,
1997). Due to their lipophilicity, pyrethroids easily permeate
through the gill, which is a contributing factor in the sensitivity of
the sh to aqueous pyrethroid exposures (Mishra et al., 2005). There
are many reports on the impact of pyrethroids on sh like biochem-
ical changes, hematological alterations, enzymatic consequences,
histopathological alterations, etc. (Gupta et al., 2000; Viran et al.,
2003; Das and Mukherjee, 2003; Koprucu and Aydin, 2004; Atif
et al., 2005; Cengiz and Unlu, 2006; Parvez and Raisuddin, 2006;
El-Sayed and Saad, 2007; Pimpao et al., 2007).
Blood is a pathophysiological reector of the whole body, so
blood parameters are important in diagnosing the structural and
functional status of the animal exposed to the toxicant. Electrolytes
of body uids have various functions the most important of which

1382-6689/$ see front matter 2009 Elsevier B.V. All rights reserved.
doi:10.1016/j.etap.2009.09.005
 L. Suvetha et al. / Environmental Toxicology and Pharmacology 29 (2010) 4449
are to contribute a majority of the osmotic ally active particles,
to provide buffer systems and mechanisms for the regulations of
pH (acidbase balance). In addition, they provide proper ionic bal-
ance for normal neuromuscular irritability and tissue functions.
In the regulation of osmolarity of a system, sodium, potassium
and calcium ions play a signicant role to keep the hyper osmotic
properties of freshwater shes. In freshwater sh the physiological
regulations of major electrolytes is very sensitive to environment
stressors and are commonly altered in response to pollutants
including pesticides (McDonald et al., 1989).
Na+ /K+ -ATPase is a highly conserved membrane enzyme essen-
tial for ion homeostasis at the cellular and organism levels (Dang et
al., 2000). In aquatic organisms particularly in sh gills, the Na+ /K+ -
ATPase is expressed abundantly in the tubular system of chloride
cells and plays a major role in the maintenance of ion balance (Metz
et al., 2003; Grosell et al., 2004; Kalay, 2006). Cypermethrin insec-
ticides inhibit both Mg2+ - and Na/K-dependent ATPases, which are
likely lead to a reduction in nerve impulse transmission and impair-
ment of ionic regulation (Larsson et al., 1985; Reddy and Philip,
1994).
In India, cypermethrin pesticide is widely used to control pests
which are attacking cotton, cabbage, brinjal, sugarcane, and wheat,
etc. Further it is also used in many sh culture operations against
lice infection. The aim of the present investigation is to examine
the acute toxicity of cypermethrin to Cyprinus carpio, a highly cul-
tivated shes to assess the potential risk of the cypermethrin to
the aquatic ecosystem at acute and sublethal levels and to evalu-
ate the relationships of the ionic regulation and gill Na+ /K+ -ATPase
activity.
2. Materials and methods
2.1. Fish and water
Fingerlings of C. carpio in the weight range of 8.0 0.4 g and body length of
8.5 1 cm were obtained from Tamil Nadu Fisheries Development Corporation
Limited, Aliyar Fish Farm, Aliyar, Tamil Nadu, India. They were safely brought
to the laboratory and acclimatized for 20 days in a large cement tank (1000 l
capacity) prior to the experiment. During the acclimatization period, sh were
fed ad libitum with rice bran and groundnut oil cake in the form of dough once
daily. Water was renewed (one third of the water) daily and feeding was with-
held 24 h before the commencement of the experiment. In the present study tap
water free from chlorine was used and the water had the following physico-
chemical characteristics (APHA, 1998); temperature (27.5 2 C), pH (7.2), dissolved
oxygen (6.4 mg l1 ), total hardness (90 mg l1 , as CaCO3 ), salinity (0.4 0.02 ppt).
Before the start of the experiment, sh were randomly divided into two groups
which were housed in 200-l aquaria with tap water which was continuously
aerated.
2.2. Toxicant
Technical grade pyrethroid insecticide, cypermethrin (25% EC) manufactured
by Rallis India Ltd., Agrochemicals Division, Mumbai, India, and supplied by Sakthi
Agroservice Ltd., Coimbatore, India, was used for evaluation of its toxicity to sh.
The stock solution of cypermethrin was prepared by dissolving in acetone (0.5%)
and appropriate amount of tap water.
2.3. Determination of 24 h LC50 value of cypermethrin
Static acute toxicity (24 h) test was conducted to determine the LC50 value of
cypermethrin toxicity considering the limitations of laboratory facilities. Different
concentrations of the cypermethrin like 0.017, 0.019, 0.021, 0.023, 0.025 ppm were
prepared from the stock and used to nd out the LC50 value for 24 h. For each con-
centration 10 sh randomly selected from the stock were introduced and kept in
separate glass tanks (120 cm 80 cm 40 cm). To each concentration a control (toxi-
cant free water) with three replicates was maintained. Equal volume of acetone was
maintained in the control aquarium. The mortality/survival of sh was recorded
after 24 h. The dead sh were removed from the tank immediately. Feeding was
withheld during the bioassay experiment. The concentration at which 50% mortal-
ity of sh occurred after 24 h was taken as the medium lethal concentration (LC50 )
for 24 h, which was 1.86 ppm. The LC50 concentration for 24 h was calculated by the
probit analysis method of Finney (1978). One-tenth value of the LC50 concentra-
tion of cypermethrin for 24 h (0.186 ppm) was taken as the sublethal concentration
(Sprague, 1971).
45
2.4. Acute toxicity test
For the determination of acute toxicity test 60 ngerlings (20 in each batch) were
selected and exposed to the cypermethrin (1.86 ppm) toxicant. Controls without
toxicant were also conducted simultaneously. At the end of 24 h period sh from the
control and experiment aquaria were taken for the analysis of plasma electrolytes
(Na+ , K+ and Cl ) and Na+ /K+ -ATPase activity.
2.5. Sublethal toxicity studies
For sublethal toxicity tests 200 ngerlings were selected and divided into two
groups with 100 sh in each aquarium. Each group was exposed to sublethal concen-
tration of the pesticide (0.186 ppm). A similar set up was also maintained as control.
During sublethal studies, sh were fed ad libitum before water replacement. The
water in the aquarium was renewed for every 24 h. and the cypermethrin concen-
tration (0.186 ppm) was added daily in the insecticide group in order to maintain
constant concentration. Experiment was conducted for 35 days and no mortality
was observed during the above treatment period. At the end of 7th, 14th, 21st, 28th
and 35th days of exposure, sh were randomly selected from experiment and con-
trol aquarium for the analysis. Blood samples were collected from each group for
the plasma electrolytes assay and simultaneously gills were removed and used for
the Na+ /K+ -ATPase activity.
2.6. Blood collection and analysis of plasma electrolytes
Blood was drawn from cardiac region by cardiac puncture using plastic dis-
posable syringe tted with 26 gauge needle which was already moistured with
heparin and expelled into separate heparinised plastic vials immediately on ice.
Blood samples were centrifuged for 15 min at 10,000 rpm and plasma was collected
for the estimation of plasma electrolytes; Na+ , K+ and Cl according to the methods
described by Maruna (1958); Sunderman and Sunderman (1959) and Schoenfeld
and Lewellan (1964) respectively.
2.6.1. Sodium and potassium assay
Sodium and potassium were estimated by the method of Maruna (1958) using
Diagnostic Reagent kit supplied by Monozyme, In vitro Diagnostics, Secunderabad,
Hyderabad, India.
2.6.2. Chloride assay
Chloride was estimated by modied method of Tietz (1990), and Young et al.
(1975) using Diagnostic Reagent kit provided by, Prism Diagnostic Pvt Ltd., Mumbai,
India.
2.7. Estimation of gill Na+ /K+ -ATPase activity
After drawing the blood, sh were washed with double distilled water and
blotted dry with absorbent paper. Then the gills were separated from the con-
trol and intoxicated sh and 100 mg gill tissue from each was weighed. They were
homogenized with ice-cold 1.0 ml of 0.1 M TrisHCl buffer (pH 7.4) using a Teon
homogenizer, and then centrifuged at 1000 rpm at 4 C for 15 min. The supernatant
was used for the estimation of Na+ /K+ -ATPase activity following the method of
Shiosaka et al. (1971).
2.8. Statistical analysis
The data were analysed statistically at P < 0.05. To test their signicance the t
values were calculated by Students t-test.
3. Results
In the present study, 24 h LC50 of cypermethrin in freshwa-
ter sh C. carpio was 1.86 ppm. One-tenth of the acute 24 h LC50
(0.186 ppm) was taken for sublethal test studies. The Chi-square
test on the toxicity data indicates that the sh population used for
the experiments was homogeneous.
3.1. Acute toxicity on plasma electrolytes and gill Na+ /K+ -ATPase
The plasma Na+ , K+ and Cl electrolytes in the cypermethrin
exposed sh during acute exposure (1.86 ppm) are presented in
Table 1. During 24 h exposure period, the plasma electrolyte lev-
els in cypermethrin treated sh were lowered when compared to
that of their control group showing a maximum percent of Na+
(53.49%), followed by Cl (16.09%) and K+ (1.18%). The differences
in ionic level were statistically signicant in the case of plasma
Na+ and Cl levels. However, plasma K+ level was not signicantly
46 L. Suvetha et al. / Environmental Toxicology and Pharmacology 29 (2010) 4449

Table 1
Changes in the plasma electrolytes (Na+ , K+ and Cl ) and gill Na+ /K+ -ATPase activity in a freshwater sh Cyprinus carpio treated with acute concentration of cypermethrin
(1.86 ppm; 24 h).

Sl. no. Parameters Control Experiment Percent change t value

1 Sodium (mmol/L) 138.34 1.10 64.56 1.79 53.49 35.010*

2 Potassium (mmol/L) 9.58 0.19 9.47 0.42 1.18 0.233
3 Chloride (mEq/L) 104.9 1.67 88.05 1.74 16.09 6.962*
4 Na+ /K+ -ATPase (g/h/g) 10.31 0.34 6.88 1.03 33.26 3.139*

Values are means SE of ve individual observations; () denotes percent decrease over control.
*
Values are signicant at P < 0.05.

changed. The inhibition of gill Na+ /K+ -ATPase activity in the intox-
icated sh is statistically signicant when compared to control
sh.
3.2. Sublethal toxicity on plasma electrolytes
Plasma Na+ level in the control sh ranged from 133.65 to
155.97 mmol/L (Fig. 1). In the cypermethrin treated sh, at the
end of 7th day plasma Na+ level was signicantly higher (+5.61).
However, at the end of 14, 21 and 28th days, plasma Na+ was sig-
nicantly lower than the control group (Fig. 1). Plasma K+ level was
higher in cypermethrin treated sh at 7th, 21st and 28th day, but
not at 14th and 35th days (Fig. 2). Regarding to the plasma Cl
level, it was signicantly decreased in cypermethrin treated sh
when compared to unexposed one during entire period of exposure
(Fig. 3).
3.3. Sublethal toxicity on gill Na+ /K+ -ATPase
Inhibition of Na+ /K+ -ATPase activity in the gills of pesticide
exposed sh seen throughout the experimental period showing
a direct relationship with the exposure period (Fig. 4). A maxi-
mum inhibition was recorded at the end of 35th day (43.81). The
Na+ /K+ -ATPase activity in the gills of control group of ngerlings
ranged from 8.75 to 10.31 g/h/g.
4. Discussion
Contamination of aquatic environments by pesticides poses a
serious threat to aquatic life forms. The 24 h LC50 value of cyper-
methrin to the ngerlings of C. carpio was found to be 1.86 ppm
indicating that cypermethrin is toxic to sh. The toxicity of cyper-
methrin varies depending on the sh species (Borges et al., 2007).
Smith and Stratton (1986) reported a list of toxic effects of cyper-
methrin on different sh species as follows (LC50 ): Salmo salar
96 h 2 g l1 ; Salmo gairdneri 96 h 6 g l1 ; Gambussia afnis 24 h
9 g l1 , and Cyprinodon macularius 24 h 10 g l1 and 48 h 6 g l1 .
Bradbury and Coast (1989) reported 96 h cypermethrin toxicity
(LC50 ) to C. carpio 0.91.1 g l1 ; Salmo trutta 1.2 g l1 ; S. gaird-
neri 0.5 g l1 ; Scardinius erythropthalmus 0.4 g l1 , Tilapia nilotica
2.2 g l1 . Das and Mukherjee (2003) reported that the 96-h LC50
to be 0.139 ppm for Labeo rohita and Ylmaz et al. (2004) reported
that the 96-h LC50 value for guppy was 9.43 g l1 . Aydin et al.
(2005) reported that the high toxicity of pyrethroids in sh to a
combination of three factors, a sensitive central nervous system,
rather slow hydrolytic detoxication and the route of exposure
(direct absorption via the gills into the blood stream). Further
pyrethroids may also secondarily induce on osmotic imbalance that
contributes to their toxicity. Due to neurotoxic effect cypermethrin
might have affected the synthesis process regulation in nerve cells
and particularly the transmembrane sodium ux, particularly in

Figs. 14. Plasma sodium, potassium, chloride and gill Na+ /K+ -ATPase levels of control and cypermethrin treated sh (0.186 ppm; 35 days).
 L. Suvetha et al. / Environmental Toxicology and Pharmacology 29 (2010) 4449
the neurohormonal system (Collins and Cappello, 2006). However
Saha and Kaviraj (2008) concluded that the cypermethrin is a mix-
ture of its isomers and the toxicity depends on the ratio of its cis
and trans isomers.
During acute treatment, the sh showed changes in its behav-
ioral attitude, drastic variation in opercular beat rates, loss of its
equilibrium, sluggish and settlement at the bottom, etc. These may
ultimately lead to respiratory distress and paralysis. The dead sh
was covered with a layer of mucous on the surface of the body. The
hypersensitivity of sh to pyrethroid toxicity may be due to species
specic differences in pyrethroid metabolism, however high sen-
sitivity of the piscine nervous system to these pesticides might be
the possible reason (Begum, 2005).
Generally physiological and biochemical measurements will
be more useful in setting priorities for determining the chemi-
cals for which more comprehensive hazard assessment is needed.
Alteration in physiological and biochemical parameters of toxi-
cant treated sh has recently emerged as an important tool for
water quality assessment in the eld of environmental toxicology.
Blood being the medium of intercellular and intracellular transport,
which comes in direct contact with various organs and tissues of
the body, the physiological state of an animal at a particular time is
reected in its blood. Freshwater teleost shes maintain their nor-
mal physiological process and their body uid homeostasis with
the help of ion/osmoregulatory processes (Hwang and Lee, 2007).
Ion uptake from water is required to maintain internal acidbase
balance and ionic equilibrium between blood and tissues for those
ions that are continuously lost by diffusion across permeable parts
of the external body surface. In the present study during acute
treatment of cypermethrin toxicity plasma Na+ , K+ and Cl were
decreased whereas in sublethal treatment plasma Na+ (except 7th
day) and Cl levels decreased. However plasma potassium level
was increased (except 35th day).
The freshwater animals compensate their renal and surface loss
of ions, mainly sodium and chloride, by absorbing these ions from
the external medium through specialized surface structures (gills
in shes and skin in amphibia). Gills of freshwater sh contain the
machinery for the active transport of the electrolytes and play an
important role in the transport of respiratory gases and regulate
the osmotic and ionic balance. Toxic substances may cause damage
to gill tissues, thereby reducing the oxygen consumption and dis-
turbing the osmoregulatory function of aquatic organisms. Kabeer
Ahamed Sahib et al. (1981) reported a decrease in Na+ , K+ and Ca2+
ions in tissue of sh exposed to malathion. Similar works were
also reported by Moorthy et al. (1984). According to McCarty and
Houston (1976), the decrease in plasma electrolyte levels tended to
be coupled with increases in tissue concentration and particularly
in the case of the major extracellular electrolytes, sodium and chlo-
ride. In L. rohita Na+ , K+ and Ca2+ ions in gills, muscle and liver were
decreased when exposed to fenvalerate indicating that fenvalerate
might have altered the permeable properties of the cell membrane
and of deranged Na+ /K+ and Ca2+ ionic pumps due to the probable
consequences of tissue damage (Reddy and Philip, 1994). Further
osmoregulatory failure may be also a reason for decreased levels of
major plasma electrolytes.
Lipophilic materials are taken up in sh partly by the gills
(McKeown and March, 1978). The accumulation of these lipophilic
substances in the cell membranes may affect the permeability of
cell membrane and also the membrane located transport mech-
anisms (Payne et al., 1978). Pyrethroids insecticides due to their
lipophilic character have a high rate of gill absorption even at low
concentrations in the water which leads to the high sensitivity of
the sh to aqueous pyrethroid exposures, because sh are unable
to metabolize the pyrethroids efciently (Viran et al., 2003). Due to
the lipophilic nature of the insecticide, pyrethroids change the u-
idity of the lipid phase of the cell membrane, and the hydrophobic
47
interactions between proteins and lipids might have alter the pro-
tein congurations (controlling membrane permeability) affecting
the transport rates and enzyme activities. In the present investiga-
tion the decreased level of electrolytes during acute and sublethal
treatment might have resulted from the lipophilic nature of the
insecticide cypermethrin which may accumulated on the gill sur-
face either damage or alter the membrane permeability leading to
lesser intake of electrolytes into the body or efux of the same to
the exterior. Further renal dysfunction due to cypermethrin toxicity
may be another possible reason.
Elevation in the serum sodium, potassium, and chloride ion
Tilapia mossambicus exposed to atrazine might have attributed to
their consequent reduction in the tissues of sh thereby indicat-
ing the imbalances in the osmoregulation phenomenon (Prasad
and Reddy, 1994). Signicant increase of serum ionic levels like
Na+ and Mg2+ produced by cypermethrin is probably due to lesions
caused by the pyrethroid in the kidney, liver and muscle (Das and
Mukherjee, 2003). There are a few reports on the electrolyte levels
in sh blood upon exposure to pesticides. Rise in serum sodium,
calcium, potassium, magnesium and chloride levels was recorded
in L. rohita and Saccobranchus fossilis exposed to chlordane and Mys-
tus vittatus exposed to thiotex, dichlorovos and carbofuran (Verma
et al., 1979). Fletcher (1975) suggested that loss of water from the
circulation could in part account for the rise in plasma electrolytes
in Pseudopleuronectes americanus during stress.
In the present investigation the signicant increase in plasma
potassium level might be that these ions were transported from
other tissues to blood or their consequent reduction in the tis-
sues due to imbalances in the osmoregulation process. McKim et
al. (1970) and Watson and Beamish (1980) reported that during
sublethal treatment, the initial effects are transient and ion concen-
trations can return to normal values despite continued exposure to
toxic concentrations; the ionoregulatory hormones like cortisol and
prolactin may be responsible for the reversal of plasma electrolytes
to the normal levels during sublethal treatments through ion mobi-
lization or restriction. Further the initial increase in plasma sodium
level and the decrease in plasma potassium level during sublethal
treatment may be due to changes in the water ion equilibrium that
occurred in inner- and intercells under the effect of cypermethrin
as suggested by Hilmy et al. (1987). Alterations in the ionic regula-
tion of environmental organisms can be due to stressor effects on
the ion regulating organs (Gilles and Requex, 1983) or internal and
external sensory receptors involved with detection of changes in
osmotic conditions (Inman and Lock Wood, 1977) or the endocrine
system (Harman et al., 1980) or metabolism (Greenway, 1979) or
active transport processes (Jowett et al., 1981).
Pollution monitoring method using enzyme inducement or
enzyme depression in sh or other organisms has been proposed
for studying polluted environments. In freshwater organisms the
enzyme Na+ /K+ -ATPase is present in the basolateral membrane of
gill epithelial cells and actively involved in the electrolyte trans-
port across the gills (Parvez et al., 2006). Na+ /K+ -ATPase is a highly
conserved membrane enzyme essential for ion homeostasis at
the cellular and organism levels (Dang et al., 2000). Xenobiotics
can alter Na+ /K+ -ATPase activity due to disruption of energy pro-
ducing metallic pathways or interact directly with the enzyme
(Watson and Beamish, 1980). Addition of toxicants to the enzyme
reaction mixture causes a concentration dependent inhibition of
Na+ /K+ -ATPase activity (Davis and Wedemeyer, 1971; Watson and
Beamish, 1981; Stagg and Shuttle Worth, 1982). Cypermethrin
the pyrethroid insecticide may inhibit Mg2+ - and Na/K-dependent
ATPases in exposed animals resulting impairment of ionic reg-
ulation. The observation was supported by David et al. (2003)
indicating that Na+ /K+ -ATPase activity in tissues of the sh C. car-
pio was inhibited due to cypermethrin intoxication. In the present
study the signicant decrease of Na+ /K+ -ATPase activity during
48 L. Suvetha et al. / Environmental Toxicology and Pharmacology 29 (2010) 4449
acute and sublethal treatment might have resulted from the direct
toxicity of the cypermethrin on ATPase function. Pyrethroids due to
its absorption in the gill region might have inhibited the enzyme gill
Na+ /K+ -ATPase which results in the disruption of cellular and ionic
regulation and salt uptake. In the present investigation the signif-
icant alterations of plasma electrolyte levels may be attributed to
a change in the membrane permeability of the gill region due to
toxicity of the insecticide cypermethrin.
5. Conclusion
The present study conclude that the enzyme Na+ /K+ -ATPase
plays a major role in branchial ion transport and the mainte-
nance of osmotic and ionic homeostasis in fresh water sh, and
the inhibition of this enzyme activity by the toxicants might have
implications for the maintenance of osmotic and ionic homeostasis
and the measurement of this enzyme can be used as an pollutant-
induced damage to the ionic and osmoregulatory system in sh.
Therefore use of pesticides which are less persistent, development
of target specic pesticides and biological control may help to
reduce the risk to man and other organisms.
Conict of interest statement
No competing interests.
References
Adhikari, S., Sarkar, B., Chatterjee, A., Mahapatra, C.T., Ayyappan, S., 2004. Effects of
cypermethrin and carbofuran on certain hematological parameters and predic-
tion of their recovery in a freshwater teleost, Labeo rohita (Hamilton). Ecotoxicol.
Environ. Saf. 58, 220226.
APHA (American Public Health Association), 1998. Standard Methods for the Exam-
ination of Water and Wastewater, 20th ed. American Public Health Association,
Washington, DC.
Atif, F., Parvez, S., Pandey, S., Ali, M., Kaur Rehman, H.A., Khan, S., Raisuddin, S., 2005.
Modulatory effect of cadmium exposure on deltamethrin-induced oxidative
stress in Channa punctata Bloch. Arch. Environ. Contam. Toxicol. 49, 371377.
Aydin, R., Koprucu, K., Dorucu, M., Kopruc, S.S., Pala, M., 2005. Acute toxicity of syn-
thetic pyrethroid cypermethrin on the common carp, Cyprinus carpio L. Embryos
and larvae. Aquat. Int. 13, 451458.
Begum, G., 2005. In vivo biochemical changes in liver and gill of Clarias batra-
chus during cypermethrin exposure and following cessation of exposure. Pestic.
Biochem. Physiol. 82, 185196.
Borges, A., Scotti, L.V., Siqueira, D.R., Zanini, R., Amaral, F., Jurinitz, D.F., Wassermann,
G.F., 2007. Changes in hematological and serum biochemical values in jundia
Rhamdia quelen due to sub-lethal toxicity of cypermethrin. Chemosphere 69,
920926.
Bradbury, S.P., Coast, J.R., 1989. Comparative toxicology of the pyrethroid insecti-
cides. Environ. Contam. Toxicol. 108, 13411377.
Cengiz, E.I., Unlu, E., 2006. Sublethal effects of commercial deltamethrin on the
structure of the gill, liver and gut tissues of mosquitosh, Gambusia afnis: a
microscopic study. Environ. Toxicol. Pharmacol. 21, 246253.
Collins, P., Cappello, S., 2006. Cypermethrin toxicity to aquatic life: bioassays for the
freshwater prawn, Palaemonetes argentinus. Arch. Environ. Contam. Toxicol. 51,
7985.
Dang, Z., Lock, R.A.C., Flik, G., Wendelaar Bonga, S., 2000. Na+ /K+ -ATPase immunore-
activity in branchial chloride of Oreochromis mossambicus exposed to copper. J.
Exp. Biol. 20, 370387.
Das, B.K., Mukherjee, S.C., 2003. Toxicity of cypermethrin in Labeo rohita ngerlings:
biochemical, enzymatic and haematological consequences. Comp. Biochem.
Physiol. 134, 109121.
David, M., Mushigeri, S.B., Philip, G.H., 2003. Alterations in the levels of ions in tissues
of freshwater sh, Labeo rohita exposed to fenvalerate. Pollut. Res. 2, 359363.
David, M., Mushigeri, S.B., Shivakumar, R., Philip, G.H., 2004. Response of Cypri-
nus carpio L. to sublethal concentration of cypermethrin: alterations in protein
metabolic proles. Chemosphere 56, 347352.
Davis, P.W., Wedemeyer, G.A., 1971. Na+ /K+ activated ATPase inhibition in rainbow
trout. A site for organochlorine pesticide toxicity. Comp. Biochem. Physiol. 40,
823827.
El-Sayed, Y.S., Saad, T.T., 2007. Sub-acute intoxication of deltamethrin-based prepa-
ration (Butox 5% EC) in monosex Nile tilapia, Oreochromis niloticus L. Basic Clin.
Pharmacol. Toxicol. 10, 17.
El-Sayed, Y.S., Saad, T.T., El-Bahr, 2007. Acute intoxication of deltamethrin in
monosex Nile tilapia, Oreochromis niloticus with special reference to the clin-
ical, biochemical and haematological effects. Environ. Toxicol. Pharmacol. 24,
212217.
ETN (Extension Toxicology Network), 1996. Pesticide information prole, revised,
available at http://www.extoxnet.orst.edu/pips/cypermet.htm.
Finney, D.J., 1978. Statistical Methods in Biological Assay, 3rd ed. Grifn Press, Lon-
don, UK, 508.
Fletcher, G.L., 1975. The effects of capture stress and storage of whole blood on the
red blood cells, plasma proteins, glucose and electrolytes of the winter ounder,
Pseudopleuronectes americanus. Can. J. Zool. 53, 197206.
Gilles, R., Requex, A., 1983. Interactions of chemicals and osmotic regulation
with the environment. In: Verberg, F.J., Verberg, W.B. (Eds.), The Biology of
Crustaceans. Environmental Adaptations, vol. 8. Academic Press, New York,
pp. 109177.
Greenway, P., 1979. Freshwater invertebrates. In: Maloig, G.M.O. (Ed.), Compara-
tive Physiology of Osmoregulation in Animals. Academic Press, New York, pp.
117173.
Grosell, M., McDonald, M.D., Walsh, P.J., Wood, C.M., 2004. Effects of copper expo-
sure in the marine gulf toadsh Opsanus beta. II. Copper accumulation, drinking
rate and Na+ /K+ -ATPase activity in osmoregulatory tissues. Aquat. Toxicol. 68,
263275.
Gupta, A., Agarwal, A.K., Shukla, G.S., 2000. Effect of quinalphos and cypermethrin
exposure on developing blood-brain barrier: role of nitric oxide. Ecotoxicol.
Environ. Saf. 8, 7378.
Harman, B.J., Jhonson, D.I., Greenwald, L., 1980. Physiological responses of Lake Erie
freshwater drum to capture by commercial shoreseine. Trans. Am. Fish. Soc. 109,
544551.
Hilmy, A.M., El Domiaty, N.A., Daabees, A.Y., Moussa, F.I., 1987. Short-term effects of
mercury on survival, behaviour, bioaccumulation and ionic pattern in the catsh
(Clarias lazera). Comp. Biochem. Physiol. 87, 303308.
Hwang, P.P., Lee, T.H., 2007. New insights into sh ion regulation and mitochondrion-
rich cells. Comp. Biochem. Physiol. 148, 479497.
Inman, C.B.E., Lock Wood, A.P.M., 1977. Some effects of methyl mercury and lindane
on sodium regulation on the amphipod, Gammarus deubeni during changes in
the salinity of its medium. Comp. Biochem. Physiol. 58C, 6775.
Jowett, D.E., Rhead, M.M., Bayne, B.L., 1981. In vivo changes in the activity of gill
ATPase and haemolymph ions of Carcinus maeas exposed to p,p -DDT at reduced
salinities. Comp. Biochem. Physiol. 69C, 399402.
Kabeer Ahamed Sahib, I., Jagannatha Rao, K.S., Ramana Rao, K.V., 1981. Effect of
malathion exposure on some physical parameters of whole body and on tissue
cations of teleost, Tilapia mossambica. J. Biol. Sci. 3 (1), 1721.
Kalay, M., 2006. The effect of cadmium on the levels of Na+ , K+ , Ca++ and Mg++ in
serum of Tilapia nilotica L. Ekoloji 15, 17.
Koprucu, K., Aydin, R., 2004. The toxic effects of pyrethroid deltamethrin on the
common carp Cyprinus carpio embryos and larvae. Pestic. Biochem. Physiol. 80,
4753.
Larsson, A., Haux, C., Johansson-Sjobeck, M.L., 1985. Fish physiology and metal pol-
lution: results and experience from laboratory and eld studies. Ecotoxicol.
Environ. Saf. 9, 250281.
Maruna, R.F.L., 1958. Quantitative estimation of sodium (Na+ ), potassium
(K+ ) in human serum by colorimetric method. Clin. Chim. Acta 2,
581585.
McCarty, L.S., Houston, A.H., 1976. Effects of exposure to sublethal levels of cadmium
upon water-electrolyte status in the goldsh, Carassius auratus. J. Fish. Biol. 9,
1119.
McDonald, D.G., Reader, J.P., Dalziel, T.R.K., 1989. The combined effects of pH and
trace metals on sh ionoregulation. In: Morris, R., Taylor, E.W., Brown, D.J.A.,
Brown, J.A. (Eds.), Acid Toxicity and Aquatic Animals. Cambridge University
Press, Cambridge, UK, pp. 221241.
McKeown, B.A., March, G.L., 1978. The effects of Bunker C oil and oil dispersant: part
2. Effects on the accumulation of chloride-labelled Bunker C oil in various sh
tissues. Mar. Environ. Res. 1, 119124.
McKim, J.M., Christensen, G.M., Hunt, E.P., 1970. Changes in the blood of brook trout,
Salvelinus fontinalis after short term and long term exposure to copper. J. Fish.
Res. Board Can. 27, 18831889.
Metz, J.R., Van den Burg, E.H., Wendelaar Bonga, S.E., Flik, G., 2003. Regulation of
branchial Na+ /K+ -ATPase in common carp Cyprinus carpio L. acclimated to dif-
ferent temperatures. J. Exp. Biol. 206, 22732380.
Mishra, D., Srivastava, S.K., Srivastava, A.K., 2005. Effects of the insecticide cyperme-
thrin on plasma calcium and ultimobranchial gland of a teleost, Heteropneustes
fossilis. Ecotoxicol. Environ. Saf. 60, 193197.
Moore, A., Waring, C.P., 2001. The effect of a synthetic pyrethroid pesticide on some
aspects of reproduction in Atlantic salmo, Salmo salar (L.). Aquat. Toxicol. 52,
112.
Moorthy, K.S., Kasi Reddy, B., Swami, K.S., Chetty, C.S., 1984. Changes in respiration
and ionic content in tissues of freshwater mussel exposed to methyl parathion
toxicity. Toxicol. Lett. 21, 287291.
Oudou, H.C., Alonso, R.M., Bruun Hansen, H.C., 2004. Voltammetric behaviour of the
synthetic pyrethroid lambda-cyhalothrin and its determination in soil and well
water. Anal. Chim. Acta 523, 6974.
Parvez, S., Raisuddin, S., 2006. Copper modulates non-enzymatic antioxidants in the
freshwater sh Channa punctata (Bloch) exposed to deltamethrin. Chemosphere
62, 13241332.
Parvez, S., Sayeed, I., Raisuddin, S., 2006. Decreased gill ATPase activities in the
freshwater sh Channa punctata (Bloch) exposed to diluted paper mill efuent.
Ecotoxicol. Environ. Saf. 65, 6266.
Payne, J.F., Kiceniuk, J.W., Squires, W.R., Fletcher, G.L., 1978. Pathological changes in
a marine sh after a six month exposure to petroleum. J. Fish. Res. Board Can.
35, 665667.
 L. Suvetha et al. / Environmental Toxicology and Pharmacology 29 (2010) 4449
Pimpao, C.T., Zampronio, A.R., Silva de Assis, H.C., 2007. Effects of deltamethrin on
hematological parameters and enzymatic activity in Ancistrus multispinis (Pisces,
Teleostei). Pestic. Biochem. Physiol. 88, 122127.
Prasad, T.A.V., Reddy, D.C., 1994. Atrazine toxicity on hydromineral balance of sh
Tilapia mossambicus. Ecotoxicol. Environ. Saf. 28, 313316.
Reddy, P.M., Philip, G.H., 1994. In vivo inhibition of AChE and ATPase activities in the
tissues of freshwater sh, Cyprinus carpio exposed to technical grade cyperme-
thrin. Bull. Environ. Contam. Toxicol. 52, 619626.
Saha, S., Kaviraj, A., 2008. Acute toxicity of synthetic pyrethroid cypermethrin to
some freshwater organisms. Bull. Environ. Contam. Toxicol. 80, 4952.
Schoenfeld, F.G., Lewellan, C.J., 1964. A colorimetric method for determination of
serum chloride. Clin. Chem. 10, 533539.
Shiosaka, T., Okuda, H., Fungi, S., 1971. Mechanisms of phosphorylation of thymidine
by the culture ltrate of Clostridium perfringens and rat liver extract. Biochim.
Biophys. Acta 246, 171183.
Smith, T.M., Stratton, G.M., 1986. Effects of synthetic pyrethroid insecticides on non-
target organisms. Residue Rev. 97, 93119.
Sprague, J.B., 1971. Measurement of pollutant toxicity to sh. III. Sublethal effects
and safe concentrations. Water Res. 5, 245266. Ylmaz, M., Gul, A., Erba, K., 2004. Acute toxicity of alpha-cypermethrin to guppy
Stagg, R.M., Shuttle Worth, T.J., 1982. The effects of copper on ionic regulation by the
gills of the seawater adapted ounder, Platichys esus L. J. Comp. Physiol. 149,
8390.
49
Sunderman, F., Sunderman, F.W., 1959. The rapid colorimetric estimation of potas-
sium. Am. J. Clin. Pathol. 29, 95.
Tietz, N.W., 1990. Clinical Guide to Laboratory Test, 2nd ed. W.B. Saunders Co.,
Philadelphia, p. 118.
Verma, S.R., Bansal, S.K., Gupta, A.K., Dalela, R.C., 1979. Pesticides induced haema-
tological alterations in a fresh water sh, Saccobranchus fossilis. Bull. Environ.
Contam. Toxicol. 22, 467474.
Viran, R., Erkoc, F.U., Polat, H., Kocak, O., 2003. Investigation of acute toxicity
of deltamethrin on guppies Poecilia reticulata. Ecotoxicol. Environ. Saf. 55,
8285.
Watson, T.A., Beamish, F.W.H., 1980. Effects of zinc on branchial ATPase activ-
ity in vivo in rainbow trout, Salmo gairdneri. Comp. Biochem. Physiol. 66,
7782.
Watson, T.A., Beamish, E.W.H., 1981. The effects of zinc on branchial adenosine
triphosphatase enzymes in vitro from rainbow trout, Salmo gairdneri. Comp.
Biochem. Physiol. 68, 167173.
Wendelaar Bonga, S.E., 1997. The stress response in sh. Physiol. Rev. 77,
592625.
(Poecilia reticulata, Pallas, 1859). Chemosphere 56, 381385.
Young, D.S., Pestaner, L.C., Gibberman, V., 1975. Effects of drugs on clinical laboratory
tests. Clin. Chem. 21, 1D432D.