Acta pharmacol. et toxicol. 1983, 53, 121-124.

From the Interdisciplinary Brain Research Center, Jawaharlal Nehru Medical College, Aligarh Muslim University, Aligarh-202001, India

Organophosphate Metasystox-induced Increment of Lipase Activity and Lipid Peroxidation in Cerebra1 Hemisphere:

Diminution of Lipids in Discrete Areas of the Rat Brain BY

Fakhrul Islam, K. Tayyaba and Mahdi Hasan (Received July 23, 1982; Accepted April 8, 1983)

Abstract: The effects of the pesticide, metasystox (0,O-dimethyl-S-2 or MetasystorB (ethylsulphinyl) ethylthiophosphate), on various lipid fractions in the discrete areas of the brain were studied. The daily intraperitoneal administration of 4 mg/kg body weight of metasystox for 10 days has depleted the levels of total lipids, phospholipids, cholesterol, esterified fatty acids and gangliosides in cerebral hemisphere, cerebellum, brain stem and spinal cord. But the lipase and lipid peroxidation measured in the cerebral hemisphere were significantly increased. It is possible that the inhibition of the lipid levels in the discrete areas of the brain may be due to the increase activity of lipase and lipid peroxidation.

Key-words: Metasystox - lipid peroxidation - lipase activity - total lipids - phospholipids - cholesterol - esterified fatty acids - gangliosides - rats.

Organophosphates are widely used as insecticides to increase the yield of agricultural products and also to control the vectors of malaria and the organism interfering with human food (Casida & Baron 1976). During the past several years num- erous studies have been conducted on metasystox because of its use as an agricultural insecticide (Kenneth & George 1962). Another organophos- phate compound DDVP (O,O-dimethyl-2:2-di- chlorovinyl phosphate), has been shown to alter the levels of total lipids, phospholipids and cholesterol in different parts of the brain (Tayyaba & Hasan 1980). No information is available on the effect of metasystox on the levels of lipids, lipase and lipid peroxidation in different regions of the central nervous system (CNS). Tissues most susceptible to lipid peroxidation appear to be those with low mitotic rates such as the brain (Barber & Wilbur

1959). It has also been reported that among the different tissues from the normal rat, the brain showed a considerably higher rate of lipid peroxi- dation than liver, kidney, spleen and heart homo- genates (Kartha & Krishnamurthy 1978). In view of these reports the present study was undertaken to demonstrate the possible effects of metasystox on the levels of lipid peroxidation, lipase activity and lipids in different regions of the CNS.

Materials and Methods

Twelve adults male albino rats weighing 200 f 20 g were used for this study. They were allowed free access to pellet diet (Hindustan Lever Laboratory Feeds, India) and water, and were divided into two groups, each comprising 6 animals. A freshly prepared solution of metasystox R (0,O-dimethyl-S-2 (ethylsulphinyl) ethylthiophosphate, Bayer India Ltd.) 4.0 mg per kg body weight was injected

122 FAKHRUL ISLAM ET AL.

intraperitoneally daily for 10 days to each rat of the experimental group. The control group received equal volume of physiological saline intraperitoneally con- currently. Over night fasted animals were stunned and killed by decapitation, brain and spinal cord were rapidly removed and cleaned of adhering blood in an ice cold petridish. Cerebrum, cerebellum, brain stem and spinal cord were dissected out. Cerebrum was divided into its two hemispheres by sagittal section. One hemisphere was used for the estimation of lipids. The other hemisphere was used for the estimation of lipid peroxidation and lipase activity.

Different parts of the brain were weighed and homo- genized in a glass homogenizer with ch1oroform:meth- anol(2:l) according to the method of Folch era/ . (1951). Each homogenate was shaken periodically for an hour at room temperature. Further isolation of lipids was carried out as described by Islam ef al. (1980). Total lipids were estimated according to the method of Woodman & Price (1972). The method of Marinetti (1962) was used for the estimation of phospholipids. The colorimetric method of Bloor era/. (1922) and Pollet et al. (1978) was used for the estimation of cholesterol and gangliosides respectively. Esterified fatty acids were estimated according to the method of Stern & Shapiro (1953).

For the evaluation of lipid peroxidation 10% (W/V) homogenate of one cerebral hemisphere was prepared in chilled 0.15 M KCI and 1.0 ml of this homogenate was used for the estimation of lipid peroxidation as described by Utley ef al. (1967). The rest of the homogenate was centrifuged at 5000 r.p.m. for 15 min. at room tempera- ture and 1.0 ml ofsupernatant was used for theestimation of lipase according to the method of Tietz & Fiereck (1966) as described in detail by Haider et a/. (1981).

All the reagents used were of analytical grade. Lipase substrate, thymolphthalein and thiobarbituric acid were purchased from Sigma Chemical Co. St. Louis, Mo., U.S.A. The data were analysed using Student's t-test. Significant differences between means of treated and control groups were calculated and P values were ob- tained. P values less than 0.05 were considered to be significant.

Results

Following the administration o f metasystox t o rats, signs such as hyperexcitability t o tactile stimuli, occasionally fasciculation, convulsions a n d ataxia were detected. After 6-7 days all the treated rats usually became lethargic. These signs were inten- sified with the passage of time. Rats of the control g r o u p did n o t show a n y abnormality.

The d a t a presented in tables 1 a n d 2 represent the effect of daily administration o f 4.0 mg/kg b o d y weight of metasystox R t o a d u l t male rats. T h e significant depleted levels of to ta l lipids, choleste-

ORGANOPHOSPHATE-INDUCED INCREMENT OF LIPASE ACTIVITY 123

Table 2. Elevation in the lipid peroxidation and the lipase activity following the administration of metasystox.

Lipid peroxidation Lipase activity (nanomol malonaldialdehyde International unit/g, formed/30 min., meanfS.D.) mean f S.D.)

Control Experimental Control Experimental

Cerebral hemisphere 2.4310.1 3.7f.0.2 0.3f0.0 0.4kO.O Percent elevation 57 50 P values <0.01 <0.01

rol and gangliosides were in the cerebral hemi- sphere and cerebellum but esterified fatty acids were decreased only in cerebral hemisphere. Levels of total lipids, phospholipids, cholesterol, esterified fatty acids and gangliosides were depleted in brain stem and spinal cord. Significantly increased activ- ity of lipase and increment in lipid peroxidation were observed in the cerebral hemisphere (table 2).

Discussion

In the present report organophosphate metasystox influenced the lipid metabolism in the brain. These observations can be very well explained on the basis of the findings of Majno & Karnovsky (1955) who reported changes in lipid synthesis in the brain following the administration of an organophos- phate. Ham & Rose (1969) have suggested that a reduced activity of lipase may be a factor contri- buting to the increased plasma lipids. Recently Haider et al. (1981) have reported that the de- creased concentration of lipids in cerebrum, cere- bellum and brain stem may also be due to increased activity of lipase in these regions. To date, no report dealing with the activity of this enzyme, in the different regions of the brain, after the administra- tion of metasystox/organophosphate is available in the literature. Hence, the reduction in the different lipid levels, irrespective of their regional variations, may be due to the increased rate of degradation, as is obvious from the present find- ings of increased lipase activity which is likely to enhance the rate of degradation of lipids, resulting in the lowering of the lipid concentration. Further- more, depletion in the phospholipid concentration is supported by the recent findings of Tayyaba &

Hasan (1980) and Nelson & Barnum (1960) who have observed reduction in the brain phospholipid concentration following organophosphate admin- istration.

Gangliosides are mainly localized in the follow- ing two fractions of the brain. A minor amount of ganglioside is localized in myelin matrix and the bulk is found in the outer membrane of the nerve endings (Ramsey & Nicholas 1972). It has been demonstrated that the distribution of gangliosides resembles that of y-aminobutyric acid (Lowden & Wolfe 1964). Ramsey & Nicholas (1972) have indicated that the binding of serotonin to synaptic vesicles may be mediated by gangliosides. It is also of interest that gangliosides can act as receptor substances for neurotoxins (Van Heyningen 1959; Doery & North 1961). There is no previous report on the levels of ganglioside in different regions of the CNS following metasystox administration. Whatever the specific role or roles of gangliosides in the brain, it is clearly evident by now that they serve an active rather than a passive function. At present the mechanism is unclear how decrement in the content of gangliosides is caused after the administration of organophosphates.

Lipid peroxidation involves the direct reaction of oxygen and lipid to form free radical intermediate and to produce semistable peroxide. Bio- membranes and subcellular organelles are the major sites of lipid peroxidation damage (Tappel 1970). Kartha & Krishnamurthy (1978) reported that among the different tissues from normal rat, the brain showed a considerably high degree of peroxidation, while the homogenate of other body organs showed comparatively low lipid peroxida- tion. This might be that the brain has the largest

124 FAKHRUL ISLAM ET AL.

a m o u n t o f lipids o f all body organs. Pathological free radical mechanism leading t o lipid peroxida- t ion a n d degradat ion o f phosphol ipids with loss o f membrane integrity a r e current ly considered as impor tan t factors in t h e development o f irrevers- ible brain cell d a m a g e dur ing ischaemic a n d o t h e r adverse condi t ions. Recently Haider et al. (1982) f rom this labora tory have repor ted tha t t h e in- crement in t h e brain lipid peroxidat ion also con- t r ibutes t o t h e decrement in t h e level o f t h e brain lipids. I t is clear tha t t h e peroxidat ion o f en- dogeneous lipids has been enhanced by metasystox adminis t ra t ion, and thus reduct ion in t h e level o f brain lipids is understandable .

A c k n o w l e d g e m e n t

G r a n t s Commission, New Delhi (India). This work was suppor ted b y the University

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