Carbohydrate Metabolism

7 . 0 TOXICITY ItfPACT ON CARBOHYDRATE ~ A ~ L I W

7.1. I NTkOIflJCTION

Carbohydrate plays a very imwrtant role in the energy

production of all living organisms. It provides energy to

organism& as fast as it is required for metabolic activities.

Generally carbohydrate is stored as glycogen in the animal body

and converted into glucose when it is needed. Besides being

oxidised to supply energy, excess carbohydrates are transformed

into fats and proteins. Thus carbohydrates function as fuel to

the body. Carbohydrate metabolism is affected when the organism

is under stresa (Grant. and Schoettger, 1972; Mazeaud &.,

1977 ) .

Though extensive work has been done on the carbohydrate

metabolism in invertebrates and many vertebrates, very little

attempt has been made on frogs. Literature on the toxicity of

heavy metals on biochemical composition of frogs is meagre.

Hodeon (1976) reported the effect of zinc on the production

and utilization of lactic acid and glycogen and accumulation of

zinc in the gills of S& m. Sastry and Gupta (1978)

etudied the effect of mercuric chloride on S h ~ d and

reported that glycogen levels were fluctuated after the

treatment. Similar observat.ions were made in the carbohydrate

metabolism of testudineus treated with lindane. disyston

and furadan (Bakt,havathsalam, 1980). Changes in the carbohydrate

metab~liam were reported in lindane treated ~vstua

( &unaswamy, 1987 .

In carbohydrate metabolism, only little work haa been done

under experiamntal condition in frogs. Therefore in the present

study an attempt hae been made to investigate the changes in the

glucoee and glycogen contents of both tissue and blood at

sublethal and median lethal treatments of copper in the green

frog h hexadactvla-

7.2. OBSEHVATI ONS

7.2.1. Glvconen kud ef Ilver. muscle nnd kldnev:

The mean control glycogen level af liver. muscle and kidney

of hexAdactvla was 21.55. 1.57 and 1.35 mg/g wet weight of

tiasue respectively. When Ilana was treated with

sublethal doee of copper. liver showed a decreased level of

glycogen than the control value. But a gradual increase of

glycogen content was found throughout the period of treatment.

The percent change over control wae -14.69, -5.35, -3.35, -1.77

and 5.34 at 24,48,72. 96 and 120 hrs of treatment respectively

(Table.4.,Fig.4.).

Muscle glycogen was found to have increased gradually at all

hours of sublethal treatnrente except at 120 hrs which ahowed a

mild decline. At the same time the muecle glycogen was greater

than the control value at all houre of treatment. The percent

increase over control was +1.01. +67.05, +126.97, +180.15 and

+162.81 at 24, 48, 72, 96 and 120 hre respectively (Table.5.

Fig.5).

Kidney glycogen was found decreased than the control at all

hours of sublethal treatment. The percent change over control at

24 hrs wee -4.61 which wae followed by -6.76 (48 hrs.), -7.86 (72

hrs). -14.50 (96 hrs) and -17.60 (120 hrs) (Table.G..Fig.G.l.

When Bans hexadactvla was treated with median lethal dose.

liver showed increased glycogen level at 24 hrs (23.50 mg/g wet

tissue) and 48 hre (22.88 mg/g wet tissue) of treatment. Whereas

it was found to have decreased at later periods of treatment and

reached 13.18 mg/g wet tissue at 120 hrs. The percent change over

control was +7.21, +5.44, -25.65, -29.24 and -38.24 at 24, 48,

72. 96 and 120 hra respectively (Table.4.,Fig.4.).

Muscle showed decreased glycogen level than the control

value at all houra of median lethal treatment. The percent

change over control wee -26.26, -3.41, -6.58, -4.41 and -0.83 at

24, 48, 72, 96 and 120 hre reepectively (Table.5.. Fig.5.).

Kidney glycogen levels were lower than the control animal at

all hours of the median lethal treatment. But at 96 and 120 hre

a erlight increase was found than the earlier period of treatment.

The percent change over control was -27.63. -29.05. -40.00,

-13.74 and -2.40 at 24. 48, 72, 96 and 120 hrs respectively

(Table.6.,Fig.6.)

7.2.2. Glucoec h Y d Qf hlood:

The mean blood glucose level of all hours was found to be

12.39 mg/ml of blood. When Bans was treated with

dose of copper, it was found to be +32.47, +43.16.

+56.27. +73.94 and +68.40 (percent change over control) at 24.

48, 72, 96 and 120 hre respectively. In median lethal treatment.

blood glucoee was found more elevated than the sublethal

treatment. It attained the maxlmum of 25.59 mg/ml at 96 hrr, and

fell to 22.44 mg/'ml of blood at 120 hrs. The percent change over

control was +51.29, +67.45, +88.09, +168.39 and +79.09 at 24, 48,

72, 96 and 120 hra reepectively (Table.7.. Fig.7.).

7.2.3. Glucoec level ef liver. muecle and k-:

The mean liver glucose level of control animal was 3.24 mg/g

wet tissue in Bans -. In sublethal treatment, it was

found lower than the control value. But an increasing trend was

obeerved upto 96 hre and a sudden fall uae recorded at 120 hre.

The percent change over control wae -46.61. -32.53, -25.31, -4.19

and -66.67 at 24, 48, 72, 96 and 120 hrs respectively

(Table.8., Fig.€!.).

The m a n of muscle glucose in control Bans hexadactvla wae

0.88 m/g wet tissue. In sublethal treatment muscle glucoee was

found lower than c o n t r o l . The pe rcen t change over c o n t r o l w a s

-20.65, -27.78, -40.91, -59.30 and -45.78 a t 24, 48, 72, 96 and

120 h r s r e spec t ive ly (Tab le .9 . . F ig .9 . ) .

The mean of c o n t r o l kidney glucoee was 1.55 mg/g wet

t i s s u e . When Barn hexadastvaA was t r e a t e d with s u b l e t h a l dose ,

kidney showed a gradual increaee i n the glucose l e v e l than t h e

control throughout t h e per iod of treatment. The percent change

over con t ro l was found t o be +5.49. +15.38, +37.01. +45.70 and

+58.12 a t 24, 48, 72, 96 and 120 h r s r eepec t ive ly

(Table. 10.. Fig. 10. ) .

When EUma hesadactvla was t r e a t e d with median l e t h a l dose,

l i v e r ehowed lower g lucose l e v e l than the c o n t r o l . But an

increas ing t r end was found throughout t h e treatment period. The

percent change over c o n t r o l was found t o be -56.50, -51.51,

-45.99, -38.71 and -33.33 a t 24, 48, 72, 96 and 120 h r s

r e spec t ive ly (Tab le .8 . , Fig.8 . ) .

The muscle glucose l e v e l was found g r e a t e r than the con t ro l

value i n median l e t h a l t rea tment . The percent change over

con t ro l was +7.61 a t 24 h r s and it was found increased t o +12.22.

+42.05 a t 48 and 72 h r s . But a t 96 and 120 h r s it was found

decreased t o +13.95 and +8.43 respect ively (Table.9. . F ig .9 . ) .

Kidney glucose l e v e l was a l s o found higher than t h e control

value in median l e t h a l treatment. The percent change over

at 24 hrs was +176.83 and it waa followed by

+7&.&5, +89.61, +127.15 and +102.70 at 48, 72, 96 and 120 hrs

reepectively (Table. 10. Fig. 10)

7.3. DISCUSSION

ChanPeeinliver.muscleandkidnevplvcoaenlevels:

In the preeent study of sublethal dose of treatment of

copper on BBna gradual elevation in the liver

glycogen was observed at all hours of treatment. which is lower

than the control. It indicates the possible glycogenesis while

glycogenolysis occurred at the sublethal treatment. In median

lethal liver glycogen level was found higher than the

control at 24 and 48 hre of treatment. At the later period it

decreased more gradually than the control value. It may be due

to the reason that at early hours of median lethal treatment

glycogen was synthesised in the liver and at the later period

break down of glycogen (Glycogenolysis) occurred to meet energy

crieie. The rate of glycogenolysis in median lethal treatment

wae found higher than the sublethal treatment.

In this regard Raja~manickam ( 1985) reported a well marked

decreaee in the liver glycogen in lindane treated

d. A decmaeed level of glycogen was reported in methyl

parathion treated (Rao and Rao, 1979).

When hexadactvLa was t r e a t e d with sub le tha l dose muscle

ehowed a g radua l e l e v a t i o n upto 96 h r and a s l i g h t fall

was obeerved a t 120 h r s . ' h ie might be due t o g l y ~ o g e n e s i ~ b,

glycogen w a s s y n t h e a i s e d - A t 120 h r a of sub le tha l

t r e a t m n t , e sudden f a l l i n t h e muscle g lucose waa a l s o observed.

This i n d i c a t e s t h e g lycogeno lys i s . In t h i s con tex t a s i g n i f i c a n t

i n c r e a s e i n muscle glycogen was r e p o r t e d i n endosulfan t r e a t e d

Dunctatua (Murthy and f riyamada k v i . 1982).

Muscle glycogen was found t o dec rease g radua l ly a t a l l hours

of median l e t h a l t r ea tmen t when compared w i t h c o n t r o l . I t might

be due t o t h e maximum u t i l i z a t i o n of glycogen f o r more muscular

a c t i v i t y needed a f t e r copper i n t o x i c a t i o n - Singh and S r iva t sava

(1982) r e g o r t e d d e p l e t i o n i n muscle glycogen i n formathion

t r e a t e d AetcroDneustee fossllle . . . A n o t a b l e dec rease i n t h e

muscle glycogen was obeerved i n Iinna hexadactvla t r e a t e d wi th

metacid 50 (Hamanujam, 1989) and nuvacron t r e a t e d Bans

(Hajam. 1987). Bakthavathsalam (1980) a l s o

r epor t ed a r a p i d d e p l e t i o n of muscle and l i v e r glycogen when t h e

f i s h wae exposed t o p e s t i c i d e s . Treatment o f Arecol ine decreased

the g lycogen c o n t e n t of l i v e r and muscle i n frog (Dragojevic and

Dida, 1978).

Kidney g lycogen was found t o have dec reased a t s u b l e t h a l and

median l e t h a l t r e a t m e n t s when compared t o t h e c o n t r o l . But a t 9 6

and 120 h r s of median l e t h a l t r ea tmen t a slow s tep-up w a s

noticed. It may be due to the glycogen synthesis at the later

period of median lethal treatment. The reason for decrement may

be due to the increased utilization of glycogen reserve to

the energy loss by copper loading and excretion, This

finding gains support from Jeyachandran and Caockalingam (1987)

and Isaiarasu and Haniffa (1987). Carbohydrate is the main and

imdiate energy precursor to animals under atress (Uminger,

1980). Reduction in glycogen level was an indication of the

onset of hypoxia involving utilization of stored glycogen by the

fish Sarotherodon exposed to endosulfan (Vasanthi and

Ramaeamy . 1987 ) . c h n g w j i n n l u c o s e l e v e l i n b l o o d :

In the present investigation, hyperglycemic condition was

observed in hexadactvla treated with both sublethal and

median lethal treatments. It might be due to glycogenolyais of

liver to liberate more glucose and the transportation of the sams

through blood to needy tissues in order to overcome the energy

demand caused by stress. kcline in the liver glycogen also

supports thia view. In thie context HcLeay and Brown (1975) have

suggested that the elevated blood glucose level has to be

correlated with mobilization of tissue glycogen. Hazeaud a d.

(1977) reported that the atresaful stimuli elicit rapid secretion

of both glucocorticoids and catocholaminea from the adrenal

tissue and these hormones produce hyperglycemia in fieh. Thie

may be applicable to amphibians also. Alloxan treatment produced

hyperglycemia and beta cell damage in I h a niDiene (Seiden,

1945). Wright (1959) also found that in the bullfrog, glucagon

administration produces rapid rise in blood sugar concentration.

~ ~ ~ ~ r g l y c e m i a was reported in Channe treated with

methyl parathion (Srivatsava and Singh, 1980).

C h a n n e a i n W l e v e l i n l i v e r . m u s c l e a n d k i d n e v :

Glucose is required readily to meet any energy demand and

hence the level of glucose is disturbed or altered whenever

organisms are brought under stress. In the present investigation

~ignificant decline in the liver glucose was observed in both

treatments. A t the same periods of both treatments, decline in

the liver glycogen was also observed. But at sublethal treatment

muscle glycogen had increased and glucose was found to have

decreased. The reduction in the liver glucose may probably be

due to the extensive glycolysis to counteract the loss of energy

due to the metal treatment. It may also be due to the reason

that glycogenolytic activity of liver produced glucose and it

mobilized through blood to muscle where it was stored as

glycogen. The increaeed muscle glycogen and decreased glucose

contente at sublethal treatments supports this view. Increaeed

blood glucose ale0 eupports this view. Rajamanickanr. (1985)

suggested a goseible transportation of glycogen as glucose from

liver t.0 muscle in lindane expomd nmffsnlabica.

~akthavathsalm (1980) also made a similar report in lindane

treated teatudineua.

Huacle glucose was found increased than the control value at

median lethal treatment. A gradual elevation was found upto 72

hrs and a alight fall was observed at 96 and 120 hrs of

treatment. The increaae in muscle glucose may probably be due to

glycogenol~tic activity of muscle. This is evidenced by the

continuous depletion of muscle glycogen at median lethal

treatment. The fall at 96 and 120 hrs may be due to utilization

for energy.

In the present study, an increase in the kidney glucose was

observed when f(ana hexadactvla was treated with both sublethal

and median lethal doses of copper. The lncrease Ln the glucose

level may be due to glycogenolys~s and gluconeogenesis in kidney.

Reduction in the kidney glycogen content at the same hours of

both treatment8 supports the view that by glycogenolysis the

stored glycogen is converted into glucose to meet the energy

demand. A decreasing trend in the aminoacid level in kidney

supports the view that gluconeogeneais could have occurred.

Continuous decreaae in the level of free aminoacids clearly

suggest their conversion into glucose through gluconeogeneeis to

meet the energy requirement of the f iah (Gupta at al. , 1987 . The aminoecids can be converted into keto acids to produce energy

through gluconeogenesie Aminoacids are believed to cope with

the atress through gluconeogeneals (Natarajan a d.. 19E3).

Kabeer Ahmed eLt al. (1976) also arrlved at a slmllar conclusion.

It is documented that malathion produced hyprglycemla In rat by

rapld mobilization af amlnclaclds and fats from t h e cellular

stores t Honnegowda and Garg , 1985 ) .

The changes In the glycogen and glucose levels are

statletlcally significant at 1% level.

7 3. I'ONC'LI.)'::IJNS

1 . In llver glucogenesla and glycugrnolysls had o:curred In

auhlethal treatment. I n the same ~erlod. ~ltlllzatron and

moblllzatlon of glu-.osr through blrtod t r m u s - l e h e d alsc

occurred which resulted In reduced gluc-ose ievel In llver.

In median lethal trratment,the rate of gly-ogenolysls was

more than sublethal treatment. Glucose thus produced ha3

undergone glycolysis to yield energy.

2. In m~~scle. due t.o glycogenesis, increased glycogen and

reduced glucose level were observed in the sublethal

treatment. In median lethal treatment glycogenolytic

activity was higher which resulted in decreased glycogen and

increased glucose levels.

3. In kidney, at sublethal t.reatment , glycogenolysis occurred

which reeulted in contintlous depletion of glycogen content.

In median lethal treatment, at the earlier period

glycopenolysie occurred, but at the later period

glycogenssie also occurred. But glucose level was found

high and it may be due to gluconeogene~~is and

glycogenolyeis.

4. Blood glucose levels were found increased at both treatmenta.

-6

Eff

ect

of v

ario

us t

reat

ren

ts and

trea

tpen

t pe

riod

s of

cop

per on g

lyco

gen

leve

l (

Wg

ast tissue)

in

the

kidney

of m

. Eac

h va

lrlt

is

the

aver

age

of

10 individuals (x

i SB). values in

pare

nthe

we reyreeent percent

change o

ver

cun

tro

l.

Tre

atm

ent

peri

ods

(Hou

rs)

48

72

96

Con

trol

1.

52

t 0.

043

1.48

t

0.04

2 1.

40

t 0.

053

1.31

t

0.02

8 1.

25

i

0.07

3

Suble t

ha 1

1.

45

t 0.

040

1.36

+

0.02

9 1.

29

2 0.

023

1.12

t

0.03

4 1.

03 i

0.02

2 (

- 4

.61)

(-

6.7

6)

(-

7.

86)

(-

14.5

0)

(-

17-8

0)

Hed

ian

leth

al

1.10

t

0.04

5 1.

05

? 0.

055

0.84

r

0.05

3 1

.13

t 0.

026

1.22

t

0.05

7 (

- 27

63

) (-

29

.05

) (-

40

DO

) (-

13.7

4)

(- 2

.4

)

a' -

0

Analysia of v

arim

r:e

-- - -

-

-- .

--

- -

Tre

atm

ent

L.

2.Cd;K;

1.32

18

59.4

720*

Per

iod

4 0.

8951

0.

2238

10

.068

0*

T~c

atm

ent x

per

iod

8 1 .

65

7

0. :!7

0 9.

3121

*

Err

r~r.

12

6 2.

8004

0.

0222

--

* Highly

sig

nific

ant

at.

1% le

vel

.

.7

Eff

ect

of various t

reat

pen

ta a

nd t

reat

men

t periods

of copper o

n glumge level (W.1)

in t

ha

blood

of

m. Kac

h value

ie

the average of 10 individuals (

X f SH),

values in

mn

tbs-

8

-sent

per

cen

t ch

snge

ove

r co

ntr

ol.

Tre

atm

ent periods

(Hou

rs)

48

72

96

Sublethal

15.91t0.787

17.8820.752

19.94i0.065

21.36t0.033

21.10i0.052

(+32.47)

(+43.16)

(+56.27)

(+73

.94)

(+68.40)

tkdian lethal 10.17

t 0.074

20.68

5 0.561

24.00

t 0.98

25.59

t 0.076

22.44

t 0.085

(+ 51.29)

(C

97.45)

(+

RB

.09)

(+lUB..39)

(+7

9.0

9)

Analy

sis

of variance

Sour

ce

d f

3s

me

F --

--

- -

Reylicatlnne

9 54.0105

6.0

01

2

1.8607

Treatment

L.

2519.9310

1259.9650

390.6693

Per iu

l 4

374.4664

93.6166

29.0271*

Treatment

R period 8

182-4301

22.8

0%

'l7.O7O6*

Error

126

406.3682

:i. :!2!51

--

-- * H

ighly significant at 1%

lev

el.

Table.8

Eff

ect

of v

ario

us

trea

tau

nts

and

trea

tmen

t periods

of c

oppe

r on

plu

cow

level (.9/p

wet

tissue) in

tbe

liver

of m

.

Each

valu

e is t

he a

vera

ge

of 10 individual8 (x

k SE).

valu

es

in

parenthceee

mw

sen

t ye

rcen

t ch

ange

we

r co

ntr

ol.

Con

trol

3.54

i 0.078

3.32

?: 0.078

3.24 i 0.067

3.10

+ 0.062

3.00

+ 0.082

Sublethal

1.89 2 0.066

2.24

+_ 0.077

2.42 t

0.073

2.97 t

0.085

1.00

0.040

(-46.61)

(-32.53)

(-25.31)

(- 4.19)

(-66

.67)

Median lethal

1.54 t

0.078

1.61

t 0.067

1.75

0.056

1.90 t 0.063

2.00

*

0.066

(- 58

.50)

(- 51.51)

(-

45.99)

(-

33.71)

(-33

.33)

8

rd

Sou

rce

df

as

ma

F --

- - -

--- - - -.

-.

Replications

9

0.2654

0.0295

0.5461

Treatment

2 59.9872

3.9

93

6

555.3913

Period

4 6.9491

1.7373

32.1690*

Treatment, x

per

iod

8 17.5981

;!. 1998

40.7331*

Err

or

126

6.8046

0.0540

P

* Highly significant

at U level.

Median lethal

"Sublethal

4.7{

Control

TREATMENT PERIOD (HMJRS)

F i g 4 Levels of Glycogen (mg/g wet tissue) in the llver of

R 3 a hexadactyla treated wlth sublethal and median lethal doses of copper for varlous hours.

1 ? j

Median le thal

I , ,Sublethal 3

0

V E

c 1 : 4 4

,;' Control / ;

U I i : 1 I

TREATMENT PERIOD (HOURS)

3 Levels of Glycogen (mg/g w e t t i s s u e ) in the muscle of

R z a hQxmdactyla t r e a t e d w i t h sublethal and median le thal doses of copper for varlous hours.

4 3

- 1 : <

, Median lethal

- 4

Control

TRECYCMENT PERIOD (HWRS)

Fig. 6 Levels of Glycogen (mg/g wet tissue) in the kidney of R z hexadactyla treated w l t h sublethal and medlan lethal doses of copper f o r various hours.

, Median lethal

A .I

E \ 0 'Sublethal

TFECFiMENT PERIOD (HOURS)

Flg. 7 Levels of Glucose (mg/ml) in the blood of R- hexadactyla treated with sublethal and medlan lethal doses of copper lor V ~ ~ I O U S hours.

i 4 Median lethal

t 'Sublethal

m . , 'Control

TRECITMENT PERIOD (HOURS)

Fig. 8 Levels of Glucose (mg/g net tissue) in the liver of Rena hexadactyla treated nith sublethal and median

lethal dosar of copper for various hours.

'1 Median l e t h a l

TRE(Tt)4ENT PERIOD (HOURS)

Fig. q L e v e l s of Glucose (mg/g wet tissue) in the muscle of

hcxrdactylr t reated with sublethal and median lethal doses of copper for various hours.

2

Sublethal

Median lethal

- I - 4 t 1 I m E Y

8 I , Control c 4 j

TREfiTHENT PERIOD (HOURS)

Fig. 10 Levels of Glucose (mg/g ne t t ~ s s u e ) in the kldney of

R z hexadactyla treated with sublethal and median lethal doses of copper for various hours.