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Comparison of the chronic effects of p,p′-DDT and α-chlordane on brain amines
EUROPEAN JOURNALOF PHARMACOLOGY20 (1972) 114-117. NORTH-HOLLANDPUBLISHINGCOMPANY
Short communication COMPARISON OF THE CHRONIC EFFECTS OF p,p'-DDT AND a - C H L O R D A N E ON B R A I N A M I N E S Pavel D. HRDINA, Radhey L. SINGHAL, David A.V. PETERS and George M. LING Department of Pharmacology, Faculty of Medicine, University of Ottawa, Ottawa, Canada
Accepted 26 June 1972
Received 24 May 1972
P.D. HRDINA, R.L. SINGHAL,D.A.V. PETERS and G.M. LING, Comparison of the chronic effects ofp,p'-DDT and a-chlordane on brain amines, European J. Pharmacol. 20 (1972) 114-117. Chronic exposure to p,p'-DDT, 2.5 mg/100 g/day for 45 days, produced slight ataxia, a decrease in striatal acetylcholine and an increase in the concentration of brain-stem 5-hydroxyindolaceticacid. In contrast, no apparent neurotoxic symptoms or changes in the levels of striatai acetylcholine or brain-stem 5-hydroxyindolaceticacid were noted after treatment with the same dose of a-chlordane. Our results demonstrate that under the experimental conditions used, a-chlordane in equivalent doses lacks the chronic effects of p,p'-DDT on the central nervous system. p,p'-DDT Brain
a-Chlordane Acetylcholine
1. INTRODUCTION Recent studies have shown that the acute neurotoxic effect of p,p'-DDT in rats (tremor, convulsions and hyperpyrexia) are associated with marked changes in cortical and striatal acetylcholine (Hrdina et al., 1971a,b) and in the metabolism of brain-stem serotonin (Peters et al., 1972). These alterations were dose-dependent and were found to correlate well with the severity of neurotoxic symptoms. It has been suggested that an enhancement in the turnover rate of brain serotonin (5-HT) may underlie the marked increase in body temperature (Peters et al., 1972) whereas a decrease in brain acetylcholine (ACh) may be related to tremor and convulsions seen during acute DDT poisoning (Hrdina et al., 1971a). Since it is the chronic exposure to small doses of chlorinated hydrocarbon insecticides which actually presents a potential health hazard, we were prompted to examine the long-term effect of p,p'-DDT and achlordane on the levels of brain ACh and other biogenic amines. The purpose of the present study was
Neurotoxicity 5-HT
Rat 5-HIAA
(1) to investigate whether any apparent signs of neurotoxicity and/or alterations in brain ACh, 5-HT and 5-hydroxyindolacetic acid (5-HIAA) are seen after the chronic exposure of rats to relatively small doses of p,p'-DDT and (2) to compare the effects of p,p'DDT with those of a-chlordane 1, a chlorinated hydrocarbon insecticide from the cyclodiene group.
2. MATERIALS AND METHODS Groups of male Wistar rats weighing approximately 100-110 g were given daffy 0.5 mg or 2.5 mg/100 g of either p,p'-DDT or a-chlordane for a period of 20 or 45 days. Both compounds were dissolved in corn off and administered by the intramuscular route. Control animals received an equal volume of oil vehicle. At least 4 rats were used in each group. The animals were kept in individual cages and had unlimited acI
1,2,4,5,6,7,8,8-Octachloro-3a,4,7,7a-tetrahydro-4,7methanoindane.
115
P.D. Hrdina et al., Brain amines after chronic exposure to DDTand chlordane
cess to food and water. They were observed carefully for occurrence o f any apparent neurotoxic symptoms or changes in their gross behaviour. Colonic temperatures o f rats were measured using a rectal probe. Animals were killed at respective time intervals by immersion into liquid nitrogen using the "near-freezing" technique of Takahashi and Aprison (1964). After decapitation, brains were removed, striata and brainstems dissected and rapidly weighed. Brain-stem refers to that portion o f the brain which was left after the removal of olfactory bulbs, cerebellum and the
cortex. Acetylcholine was extracted with acid-ethanol according to the m e t h o d of Crossland (1961) and bioassayed on eserinized frog rectus preparation as described in an earlier communication (Hrdina et al., 1971 c). 5-HT and 5-HIAA were determined according to the m e t h o d of Curzon and Green (1972). The data were statistically analysed by using the Student's ttest and significant differences between the means were calculated as p values.
3. RESULTS Table 1 Effects of chronic treatment with a-chlordane or p,p'-DDT on ACh content of rat striatum. ACh (nmol/g) Treatment
None
(controls)
a-Chlordane 0.5 mg/100 g/day 2.5 mg/100 g/day p,p'-DDT 0.5 mg[100 g/day 2.5 mg/100 g/day
20 days
45 days
46.8 ± 2.0 * (100)
52.3 -* 2.4 (100)
51.7 ± 5.7 (110) 43.7 ± 2.8 (93)
46.1 -+ 2.6 (90) 50.3 -* 2.3 (96)
44.9 ± 0.5 (96) 51.5 ± 2.8 (109)
42.0 ~ 1.3 t (80) 39.6 ± 3.3 1 (76)
After 45 days o f p,p'-DDT treatment, significant decreases in the amount of striatal ACh were noted in rats receiving either 0.5 or 2.5 mg/100 g dose o f this insecticide (table 1). In contrast, no such changes were observed in brain ACh o f animals receiving either 0.5 or 2.5 mg/100 g dose oftx-chlordane. Similarly, tz-chlordane failed to produce any significant change in the levels o f brain-stem 5-HT or 5-HIAA (table 2). However, a significant increase in 5-HIAA was noted in animals receiving the higher dose (2.5 mg/100 g) of p,p'-DDT for 45 days. In this group of rats, slight ataxia also was observed during the last 5 - 6 days of the experimental period. No other signs o f neurotoxicity or changes in gross behaviour were recorded in animals treated with either insecticide. Administration of p,p'-DDT or a-chlordane in a dose o f 0.5 or 2.5 mg/100 g for 20 days did not produce any changes in the concentration o f striatal ACh or brain-stem 5-HT and 5-HIAA (tables 1 and 2).
* Mean ± S.E.M. based o n at least 4 animals in each group. t Statistically significant difference when compared with the control values (/9 < 0.05). Table 2 Effects of chronic exposure to two different doses of c~-chlordaneor p,p'-DDT on 5-HT and 5-HIAA levels in the brain-stem of rats. 5-HIAA (~tg/g/wet wt.)
5-HT (tag/g/wet wt.) Treatment
20 days
45 days
20 days
45 days
1.02 ± 0.07 *
0.86 ± 0.03
0.74 :~ 0.06
0.70 ± 0.04
cz-Chlordane 0.5 mg/kg/day 2.5 mg/kg[day
0.93 ± 0.04 0.89 ± 0.02
0.93 ± 0.03 0.86 ± 0.05
0.78 ± 0.09 0.84 ± 0.05
0.76 ± 0.04 0.76 ± 0.05
p,p'-DDT 0.5 mg/kg/day 2.5 mg/kg/day
1.04 ± 0.02 0.86 ± 0.05
0.88 ± 0.05 0.84 ± 0.05
0.80 ± 0.11 0.73 ± 0.09
0.84 ± 0.13 0.94 ± 0.08 t
None
(controls)
* Means t S.E.M. based on at least 4 rats in each group.
t Statistically significant difference when compared with the control values (p < 0.05).
116
P.D. Hrdina et al., Brain amines after chronic exposure to DD T and chlordane
Average colonic temperatures in each experimental group increased by approximately 1.5°C when compared to the control group on the 45th day of treatment with both insecticides (table 3). There was no significant change in weight gain of animals treated with either p,p'-DDT or a-chlordane for 20 or 45 days over that of control animals. Table 3 Colonic temperature of rats treated with a-chlordane or p,p'DDT for 20 or 45 days. Colonic temperature (0°C) Treatment Control a-Chlordane 0.5 mg/100 g/day 2.5 mg/100 g/day p,p'-DDT 0.5 mg/100 g[day 2.5 mg/100 g/day
20th day
45th day
37.0 ± 0.1 *
36.5 ± 0.1
37.0 ± 0.1 37.5 ± 0.2
38.2 +-0.2 1" 38.0 ± 0.1 I"
37.5 ± 0.2 37.0 -+0.2
38.0 ± 0.1 1" 38.3 ± 0.1 "~
* Means ± S.E.M. based on at least four animals in each group. 1"Statistically significant difference when compared with the control values (p < 0.05).
4. DISCUSSION Our results demonstrate that long-term exposure of rats to relatively small doses of p,p'-DDT is capable of producing changes in the levels of striatal ACh that are qualitatively similar to those observed during acute poisoning with this insecticide (Hrdina et al., 1971a). Desi et al. (1966) and Farkas et al. (1968) reported that when rats were administered daily with 2 mg/100 g dose of DDT, marked changes in the EEG pattern (increases in both frequency and amplitude) were observed after four weeks, and slight ataxia after five weeks of insecticide treatment. They speculated that the observed EEG changes might result from an accumulation of acetylcholine caused by DDT. While our results on the occurrence of ataxia agree with those of the above investigators, we found a decrease rather than an increase in brain ACh after chronic treatment with p,p'-DDT. Central nervous system excitation has been found to be associated with a lowering of brain ACh (Aprison et al., 1968). It is therefore
possible that the DDT-induced EEG changes which are presumably due to increased irritability of the central nervous system (Farkas et al., 1968) may be associated with decreased brain ACh levels observed in the present study. Stohlman et al. (1950) reported that the chronic toxicity of dietary chlordane in rats was greater than that seen with DDT. However, in our experiments, chronic treatment with a-chlordane failed to produce ataxia or any significant change in the concentration of striatal ACh, which is in contrast to the effects exerted by p,p'-DDT. It is of interest that a-chlordane, when administered chronically, was found to be as effective as p,p'-DDT in enhancing the gluconeogenic potential of both kidney cortex and liver (Kacew et al., 1972). It would seem therefore, that during chronic exposure, different tissues exhibit different degree of sensitivity to various insecticides. Recently, Peters et al. (1972) demonstrated that administration of an acute dose (60 mg/100 g) of p,p'-DDT resulted in an increased turnover rate of brain-stem 5-HT which correlated well with marked increase in body temperature. The present study shows that moderate increase in body temperature which resulted from 45 days of treatment with either a-chlordane or p,p'-DDT was accompanied by enhanced levels of 5-HIAA only in the group of rats receiving the higher dose (2.5 rag/100 g) of p,p'-DDT. It is conceivable that an increased excretion of 5HIAA due to an adaptation to the long-term treatment with insecticides may have prevented the expected accumulation of this 5-HT metabolite in the brain-stem. It is concluded that, under the experimental condi. tions used, chronic treatment with a-chlordane fails to produce ataxia as well as alterations in brain ACh and 5-HIAA levels seen after the daily administration of same doses of p,p'-DDT. ACKNOWLEDGEMENTS This study was assisted by funds provided by the Public Health Research Grant (Project No. 605-7-711) of the National Health Grants Program of the Government of Canada. The authors wish to thank Dr. P. Polen of Velsicol Laboratories, Chicago, Illinois, for the generous supply ofp,p'-DDT and a-chlordane. REFERENCES Aprison, M.H., T. Kariya, J.N. Hingtgen and M. Toru, 1968,
P.D. Hrdina et al., Brain amines after chronic exposure to DDT and chlordane
Neurochemical correlates of behaviour: changes in acetylcholine, norepinephrine and 5-hydroxytryptamine concentrations in several discrete brain areas of the rat during behaviourai excitation, J. Neurochem. 15, 1131 - 1139. Crossland, J., 1961, Biologic estimation of acetylcholine, in: Methods in Medical Research, Vol. 7, ed. J.H. Quastel (Year Book Medical Publishers, Chicago) pp. 125-129. Curzon, W.E. and A.R. Green, 1972, Rapid method for the determination of 5-hydroxytryptamine metabolism, Brit. J. Pharmacol. 39, 653-655. Desi, I., I. Farkas and T. Kemeny, 1966, Changes of central nervous function in response to DDT administration, Acta Physiol. Acad. Sci. Hung. 30, 275-282. Farkas, I., I. Desi and T. Kemeny, 1968, The effect of DDT in the diet on the resting and loading electrocorticogram record, Toxicol. Appl. Pharmacol. 12, 518-525. Hrdina, P.D., R.L. Singhal, D.A.V. Peters and G.M. Ling, 197 la, Role of brain acetylcholine and dopamine in acute neurotoxic effects of DDT, European J. Pharmacol. 15, 379-382.
117
Hrdina, P.D., A. Maneckjee, S. Kacew, D.A.V. Peters and R.L. Singhal, 1971b, Acetylcholine and dopamine in rat striatum during acute DDT poisoning, Proc. Can. Fed. Biol. Soc. 14, 67. Hrdina, P.D., G.M. Ling and A. Maneckjee, 1971c, Desipramine (DMI): Effect on the levels of acetylcholine (ACh) in whole brain and in striatum of rats, European J. Pharmacol. 15,141-144. Kacew, S. and R.L. Singhal, 1972, Effects of chronic exposure to p,p'-DDT and a-chlordane on enzymes in liver and kidney cortex, Proc. Can. Fed. Biol. Soc., in press. Peters, D.A.V., P.D. Hrdina, R.L. Singhal and G.M. Ling, 1972, The role of brain serotonin in DDT-induced hyperpyrexia, J. Neurochem. 19, in press. Stohlman, E.F., W.T.S. Thorp and M.I. Smith, 1950, Toxic action of chlordane, Arch. Ind. Hyg. Occup. Med. 1, 13-19. Takahashi, R. and M.H. Aprison, 1964, Acetylcholine content of discrete areas of the brain obtained by a nearfreezing method, J. Neurochem. 11,887-898.
Short communication COMPARISON OF THE CHRONIC EFFECTS OF p,p'-DDT AND a - C H L O R D A N E ON B R A I N A M I N E S Pavel D. HRDINA, Radhey L. SINGHAL, David A.V. PETERS and George M. LING Department of Pharmacology, Faculty of Medicine, University of Ottawa, Ottawa, Canada
Accepted 26 June 1972
Received 24 May 1972
P.D. HRDINA, R.L. SINGHAL,D.A.V. PETERS and G.M. LING, Comparison of the chronic effects ofp,p'-DDT and a-chlordane on brain amines, European J. Pharmacol. 20 (1972) 114-117. Chronic exposure to p,p'-DDT, 2.5 mg/100 g/day for 45 days, produced slight ataxia, a decrease in striatal acetylcholine and an increase in the concentration of brain-stem 5-hydroxyindolaceticacid. In contrast, no apparent neurotoxic symptoms or changes in the levels of striatai acetylcholine or brain-stem 5-hydroxyindolaceticacid were noted after treatment with the same dose of a-chlordane. Our results demonstrate that under the experimental conditions used, a-chlordane in equivalent doses lacks the chronic effects of p,p'-DDT on the central nervous system. p,p'-DDT Brain
a-Chlordane Acetylcholine
1. INTRODUCTION Recent studies have shown that the acute neurotoxic effect of p,p'-DDT in rats (tremor, convulsions and hyperpyrexia) are associated with marked changes in cortical and striatal acetylcholine (Hrdina et al., 1971a,b) and in the metabolism of brain-stem serotonin (Peters et al., 1972). These alterations were dose-dependent and were found to correlate well with the severity of neurotoxic symptoms. It has been suggested that an enhancement in the turnover rate of brain serotonin (5-HT) may underlie the marked increase in body temperature (Peters et al., 1972) whereas a decrease in brain acetylcholine (ACh) may be related to tremor and convulsions seen during acute DDT poisoning (Hrdina et al., 1971a). Since it is the chronic exposure to small doses of chlorinated hydrocarbon insecticides which actually presents a potential health hazard, we were prompted to examine the long-term effect of p,p'-DDT and achlordane on the levels of brain ACh and other biogenic amines. The purpose of the present study was
Neurotoxicity 5-HT
Rat 5-HIAA
(1) to investigate whether any apparent signs of neurotoxicity and/or alterations in brain ACh, 5-HT and 5-hydroxyindolacetic acid (5-HIAA) are seen after the chronic exposure of rats to relatively small doses of p,p'-DDT and (2) to compare the effects of p,p'DDT with those of a-chlordane 1, a chlorinated hydrocarbon insecticide from the cyclodiene group.
2. MATERIALS AND METHODS Groups of male Wistar rats weighing approximately 100-110 g were given daffy 0.5 mg or 2.5 mg/100 g of either p,p'-DDT or a-chlordane for a period of 20 or 45 days. Both compounds were dissolved in corn off and administered by the intramuscular route. Control animals received an equal volume of oil vehicle. At least 4 rats were used in each group. The animals were kept in individual cages and had unlimited acI
1,2,4,5,6,7,8,8-Octachloro-3a,4,7,7a-tetrahydro-4,7methanoindane.
115
P.D. Hrdina et al., Brain amines after chronic exposure to DDTand chlordane
cess to food and water. They were observed carefully for occurrence o f any apparent neurotoxic symptoms or changes in their gross behaviour. Colonic temperatures o f rats were measured using a rectal probe. Animals were killed at respective time intervals by immersion into liquid nitrogen using the "near-freezing" technique of Takahashi and Aprison (1964). After decapitation, brains were removed, striata and brainstems dissected and rapidly weighed. Brain-stem refers to that portion o f the brain which was left after the removal of olfactory bulbs, cerebellum and the
cortex. Acetylcholine was extracted with acid-ethanol according to the m e t h o d of Crossland (1961) and bioassayed on eserinized frog rectus preparation as described in an earlier communication (Hrdina et al., 1971 c). 5-HT and 5-HIAA were determined according to the m e t h o d of Curzon and Green (1972). The data were statistically analysed by using the Student's ttest and significant differences between the means were calculated as p values.
3. RESULTS Table 1 Effects of chronic treatment with a-chlordane or p,p'-DDT on ACh content of rat striatum. ACh (nmol/g) Treatment
None
(controls)
a-Chlordane 0.5 mg/100 g/day 2.5 mg/100 g/day p,p'-DDT 0.5 mg[100 g/day 2.5 mg/100 g/day
20 days
45 days
46.8 ± 2.0 * (100)
52.3 -* 2.4 (100)
51.7 ± 5.7 (110) 43.7 ± 2.8 (93)
46.1 -+ 2.6 (90) 50.3 -* 2.3 (96)
44.9 ± 0.5 (96) 51.5 ± 2.8 (109)
42.0 ~ 1.3 t (80) 39.6 ± 3.3 1 (76)
After 45 days o f p,p'-DDT treatment, significant decreases in the amount of striatal ACh were noted in rats receiving either 0.5 or 2.5 mg/100 g dose o f this insecticide (table 1). In contrast, no such changes were observed in brain ACh o f animals receiving either 0.5 or 2.5 mg/100 g dose oftx-chlordane. Similarly, tz-chlordane failed to produce any significant change in the levels o f brain-stem 5-HT or 5-HIAA (table 2). However, a significant increase in 5-HIAA was noted in animals receiving the higher dose (2.5 mg/100 g) of p,p'-DDT for 45 days. In this group of rats, slight ataxia also was observed during the last 5 - 6 days of the experimental period. No other signs o f neurotoxicity or changes in gross behaviour were recorded in animals treated with either insecticide. Administration of p,p'-DDT or a-chlordane in a dose o f 0.5 or 2.5 mg/100 g for 20 days did not produce any changes in the concentration o f striatal ACh or brain-stem 5-HT and 5-HIAA (tables 1 and 2).
* Mean ± S.E.M. based o n at least 4 animals in each group. t Statistically significant difference when compared with the control values (/9 < 0.05). Table 2 Effects of chronic exposure to two different doses of c~-chlordaneor p,p'-DDT on 5-HT and 5-HIAA levels in the brain-stem of rats. 5-HIAA (~tg/g/wet wt.)
5-HT (tag/g/wet wt.) Treatment
20 days
45 days
20 days
45 days
1.02 ± 0.07 *
0.86 ± 0.03
0.74 :~ 0.06
0.70 ± 0.04
cz-Chlordane 0.5 mg/kg/day 2.5 mg/kg[day
0.93 ± 0.04 0.89 ± 0.02
0.93 ± 0.03 0.86 ± 0.05
0.78 ± 0.09 0.84 ± 0.05
0.76 ± 0.04 0.76 ± 0.05
p,p'-DDT 0.5 mg/kg/day 2.5 mg/kg/day
1.04 ± 0.02 0.86 ± 0.05
0.88 ± 0.05 0.84 ± 0.05
0.80 ± 0.11 0.73 ± 0.09
0.84 ± 0.13 0.94 ± 0.08 t
None
(controls)
* Means t S.E.M. based on at least 4 rats in each group.
t Statistically significant difference when compared with the control values (p < 0.05).
116
P.D. Hrdina et al., Brain amines after chronic exposure to DD T and chlordane
Average colonic temperatures in each experimental group increased by approximately 1.5°C when compared to the control group on the 45th day of treatment with both insecticides (table 3). There was no significant change in weight gain of animals treated with either p,p'-DDT or a-chlordane for 20 or 45 days over that of control animals. Table 3 Colonic temperature of rats treated with a-chlordane or p,p'DDT for 20 or 45 days. Colonic temperature (0°C) Treatment Control a-Chlordane 0.5 mg/100 g/day 2.5 mg/100 g/day p,p'-DDT 0.5 mg/100 g[day 2.5 mg/100 g/day
20th day
45th day
37.0 ± 0.1 *
36.5 ± 0.1
37.0 ± 0.1 37.5 ± 0.2
38.2 +-0.2 1" 38.0 ± 0.1 I"
37.5 ± 0.2 37.0 -+0.2
38.0 ± 0.1 1" 38.3 ± 0.1 "~
* Means ± S.E.M. based on at least four animals in each group. 1"Statistically significant difference when compared with the control values (p < 0.05).
4. DISCUSSION Our results demonstrate that long-term exposure of rats to relatively small doses of p,p'-DDT is capable of producing changes in the levels of striatal ACh that are qualitatively similar to those observed during acute poisoning with this insecticide (Hrdina et al., 1971a). Desi et al. (1966) and Farkas et al. (1968) reported that when rats were administered daily with 2 mg/100 g dose of DDT, marked changes in the EEG pattern (increases in both frequency and amplitude) were observed after four weeks, and slight ataxia after five weeks of insecticide treatment. They speculated that the observed EEG changes might result from an accumulation of acetylcholine caused by DDT. While our results on the occurrence of ataxia agree with those of the above investigators, we found a decrease rather than an increase in brain ACh after chronic treatment with p,p'-DDT. Central nervous system excitation has been found to be associated with a lowering of brain ACh (Aprison et al., 1968). It is therefore
possible that the DDT-induced EEG changes which are presumably due to increased irritability of the central nervous system (Farkas et al., 1968) may be associated with decreased brain ACh levels observed in the present study. Stohlman et al. (1950) reported that the chronic toxicity of dietary chlordane in rats was greater than that seen with DDT. However, in our experiments, chronic treatment with a-chlordane failed to produce ataxia or any significant change in the concentration of striatal ACh, which is in contrast to the effects exerted by p,p'-DDT. It is of interest that a-chlordane, when administered chronically, was found to be as effective as p,p'-DDT in enhancing the gluconeogenic potential of both kidney cortex and liver (Kacew et al., 1972). It would seem therefore, that during chronic exposure, different tissues exhibit different degree of sensitivity to various insecticides. Recently, Peters et al. (1972) demonstrated that administration of an acute dose (60 mg/100 g) of p,p'-DDT resulted in an increased turnover rate of brain-stem 5-HT which correlated well with marked increase in body temperature. The present study shows that moderate increase in body temperature which resulted from 45 days of treatment with either a-chlordane or p,p'-DDT was accompanied by enhanced levels of 5-HIAA only in the group of rats receiving the higher dose (2.5 rag/100 g) of p,p'-DDT. It is conceivable that an increased excretion of 5HIAA due to an adaptation to the long-term treatment with insecticides may have prevented the expected accumulation of this 5-HT metabolite in the brain-stem. It is concluded that, under the experimental condi. tions used, chronic treatment with a-chlordane fails to produce ataxia as well as alterations in brain ACh and 5-HIAA levels seen after the daily administration of same doses of p,p'-DDT. ACKNOWLEDGEMENTS This study was assisted by funds provided by the Public Health Research Grant (Project No. 605-7-711) of the National Health Grants Program of the Government of Canada. The authors wish to thank Dr. P. Polen of Velsicol Laboratories, Chicago, Illinois, for the generous supply ofp,p'-DDT and a-chlordane. REFERENCES Aprison, M.H., T. Kariya, J.N. Hingtgen and M. Toru, 1968,
P.D. Hrdina et al., Brain amines after chronic exposure to DDT and chlordane
Neurochemical correlates of behaviour: changes in acetylcholine, norepinephrine and 5-hydroxytryptamine concentrations in several discrete brain areas of the rat during behaviourai excitation, J. Neurochem. 15, 1131 - 1139. Crossland, J., 1961, Biologic estimation of acetylcholine, in: Methods in Medical Research, Vol. 7, ed. J.H. Quastel (Year Book Medical Publishers, Chicago) pp. 125-129. Curzon, W.E. and A.R. Green, 1972, Rapid method for the determination of 5-hydroxytryptamine metabolism, Brit. J. Pharmacol. 39, 653-655. Desi, I., I. Farkas and T. Kemeny, 1966, Changes of central nervous function in response to DDT administration, Acta Physiol. Acad. Sci. Hung. 30, 275-282. Farkas, I., I. Desi and T. Kemeny, 1968, The effect of DDT in the diet on the resting and loading electrocorticogram record, Toxicol. Appl. Pharmacol. 12, 518-525. Hrdina, P.D., R.L. Singhal, D.A.V. Peters and G.M. Ling, 197 la, Role of brain acetylcholine and dopamine in acute neurotoxic effects of DDT, European J. Pharmacol. 15, 379-382.
117
Hrdina, P.D., A. Maneckjee, S. Kacew, D.A.V. Peters and R.L. Singhal, 1971b, Acetylcholine and dopamine in rat striatum during acute DDT poisoning, Proc. Can. Fed. Biol. Soc. 14, 67. Hrdina, P.D., G.M. Ling and A. Maneckjee, 1971c, Desipramine (DMI): Effect on the levels of acetylcholine (ACh) in whole brain and in striatum of rats, European J. Pharmacol. 15,141-144. Kacew, S. and R.L. Singhal, 1972, Effects of chronic exposure to p,p'-DDT and a-chlordane on enzymes in liver and kidney cortex, Proc. Can. Fed. Biol. Soc., in press. Peters, D.A.V., P.D. Hrdina, R.L. Singhal and G.M. Ling, 1972, The role of brain serotonin in DDT-induced hyperpyrexia, J. Neurochem. 19, in press. Stohlman, E.F., W.T.S. Thorp and M.I. Smith, 1950, Toxic action of chlordane, Arch. Ind. Hyg. Occup. Med. 1, 13-19. Takahashi, R. and M.H. Aprison, 1964, Acetylcholine content of discrete areas of the brain obtained by a nearfreezing method, J. Neurochem. 11,887-898.