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The effects of gold thioglucose on the central nervous system of chicks (Gallus domesticus)
TOXICOLOGY
AND
APPLIED
PHARMACOLOGY
(1976)
35223-228
The Effects of Gold Thioglucose on the Central Nervous System of Chicks (Gal/us Domesticus) MICHAEL J. GENTLE Poultry Research Centre, Edinburgh EH9 3JS, Scotland Received March 13,1975; accepted August 241975
The Effectsof Gold Thioglucoseon the Central NervousSystemof Chicks GENTLE, M. J. (1976).Toxicol. Appl. Pharmacol. 35,223228. The effectsof gold thioglucoseon the central nervoussystem,growth, and laying performancewerestudiedfollowing a singleintramuscularinjection of the drug in 2- and21-day-old chicks(Gallus domesticus). Injection of gold thioglucosein dosesof 1-O.1mg/gbody weightgaveriseto a largenumber of small perivascularlesionsdistributed mainly in the telencephalon and diencephalon.None of the birds which recoveredfrom a singledoseof 0.2 mg/gof gold thioglucoseshowedanyeffect on growth rate or laying performancecomparedto the control birds. (GaZlus domesticus).
Single injections of an LD50 of gold thioglucose (GTG) have been shown to induce in several strains of mice and rats lesions in the ventromedial region of the hypothalamus
and in other areas of the brain (Liebelt and Perry, 1967). This hypothalamic damage results in the animal becoming hyperphagic and obese. Carpenter et al. (1969) found that in quail there was considerable variability in the LD50 of the drug and no evidence of either histological or functional damage to the hypothalamus. Simkins and Pensach (1970) and Svacha and Reid (1973) found LD50 values to be 0.5 mg/g body weight for 5- to 7-day-old broiler chicks. The last two authors reported that the surviving birds showed some impairment in the regulation of body temperature as well as moderate increasesin growth rate and food consumption but did not report whether these increases were significant. Svacha and Reid (1973) also found that GTG injected into mature white Leghorn hens did not influence body weight or food intake. No changes in food intake or brain damage were found in brown Leghorn males (Wood-Gush and Gower, personal communication) following the injection of GTG in doses from 0.3 to 1.1 mg/g. One hypothesis for the mode of topic action of the drug in mammals is that the glucose becomes attached to specific glucoreceptor neurons in the hypothalamus and, because of the high toxicity of the drug, cell death results (Mayer and Marshall, 1956). One reason for the failure to find brain damage in adult birds following the injection of GTG may be due to a failure of the GTG to penetrate the blood-brain barrier. To avoid this difficulty chicks were used in the present experiment in order to study effects induced before the blood-brain barrier is fully developed. Previous reports using GTG in chicks Copyright 0 1976 by Academic Press, Inc. All rights of reproduction in any form reserved. Printed in Great Britain
223
MlCHAELJ.GENTLE
224
(Simkins and Pensach, 1970: Svacha and Reid, 1973) are confined to broiler strains. Broiler chickens have been specially bred to have a high growth rate, so that it might be difficult to induce hyperphagia becausethey might already be near their upper limit of growth and food intake. In the present experiment the birds usedwere from a laying strain which had not been bred for a fast growth rate and would hence have more potential to increasetheir food intake.
METHODS The birds were female white Leghorn chicks which were bred at the Poultry Research Centre. They were housed in Perspex boxes under continuous light and maintained at 33°C by using infrared heaters. Food and water were always available. The number of birds used, their ages,and the dosesof gold thioglucose injected are shown in Table 1. Two groups of birds were injected at 2 and 21 days of age and were TABLE 1 SUMMARYOFTHENUMBEROF
CHICKS USED,THEIRAGE,THEDRUGDOSAGEANDTHESUBSEQIJENTFATEOFTHECHICKS
Number injected Total number Age of birds used (days) Drug injected and dose at eachdose 20
10
2 0.1-1.0 mg/g GTG in O.lmg steps 21 0.1-1.0 mg/g GTG in O.lmg steps 2 Salinecontrol
10
10
21 Salinecontrol
10
45
21 0.2 mg/g GTG
45
21
21 Salinecontrol
21
20
2 2
Treatment Died or killed in 24 hr, brain removed Died or killed in 24 hr, brain removed Killed in 24 hr, brain removed Killed in 24 hr, brain removed Body weight recorded at IO-day intervals up to 230 days Body weight recorded at IO-day intervals up to 230days
not allowed to survive for more than 24 hr after injection. These birds were used to determine the LD50 of the drug and to study its effects on the central nervous system. Each bird was given a single im injection of either GTG in oil (Solganal, Schering Corporation) or saline.To determine the approximate LD50 value, two birds were injected with the samedoseof the drug at 2 days and another group at 21 days of age. The dosesranged from 0.1 to 1.Omg/g body weight in O.l-mg steps.To study the effects of the GTG on the central nervous system, the surviving birds were killed after 24 hr using ether, and the brains of those that had died as well as those that were killed were dis-
GOLD
THIOGLUCOSE
IN CHICKS
225
sected out and fixed. Half the brains, i.e., one for each age and concentration, were fixed in aqueous Bouin and the other half in 10 % formol-saline. Those fixed in Bouin were embedded in paraffin wax, sectioned at 8 pm and stained with hematoxylin/eosin to study the general anatomy. Those fixed in formol-saline were frozen, cut at 50 pm, and stained using the Weil (1946) technique to look for demyelination. The third group of birds was injected with either 0.2 mg/g of GTG or saline at 21 days of age, and the animals were allowed to live for 230 days in order to see if the brain damage induced by the drug had any effect on the birds’ growth or laying performance. The birds were injected at 21 days of age because there is normally a higher mortality in the younger chicks. The use of 45 birds in the experimental group was to ensure equal numbers of control and experimental birds at the end of the 230-day period.
RESULTS
Mortality All birds injected at 2 days of age with doses of 0.3 mg/g or more became very sick or died within 24 hr. In the al-day-old birds the results were similar. Of the 45 birds injected with 0.2 mg/g, 20 died within 3 days of injection and a further 12 died or were killed in extremis during the next 230 days. In the 21-day-old control birds none died in the first 3 days following injection, but six died or were killed before reaching 230 days of age. It was not possible to determine the LD50 with any accuracy, as the mortality varied from group to group. The 45 birds that were injected at 21 days of age with 0.2 mg/g were injected in small groups. The groups consisted of 5, 12, 14, and 14 birds, of which 1, 6, 11, and 2 died within 3 days after injection. Pathological Changes Damage to the central nervous system following the injection of GTG was very extensive and took the form of a large number of small lesions. These were distributed around blood vessels (Fig. 1). The lesions consisted of a focal disseminated edema in the neuropile of the grey matter. The nuclei of the cells in the lesion area were smaller, irregularly shaped, and more deeply stained than the normal cells, indicating cell death, so that there was perivascular necrosis in addition to the edema. They were concentrated mainly in the telencephalon and anterior diencephalon. Fewer lesions were found in the posterior diencephalon. Although there were some lesions in the cerebellum there were few in the mesencephalon and medulla oblongata. The distribution of the lesions reflected that of the fine capillary blood supply to the brain. The extent of the lesions depended upon the dose of the drug and was greatest at high doses but was still extensive at 0.2 mg/g (Fig. 1). There were no clear pathological differences between the 2- and 21day-old chicks. There was no clear evidence of demyelination. Growth, Sexual Maturity, and Rate of Lay The mean weights of the surviving 13 control birds and 12 treated with GTG are shown in Fig. 2; there were no significant differences. Nor was there any significant
226
MICHAEL
J. GENTLE
difference in the date of onset of laying (mean values of 155.8 k 5.8 and 159.2 + 7.5 days) or in the number of eggs laid in the 25-day period after the onset of lay (means 12.1 & 1.6 and 11.5 & 1.4eggs).
PL
0
100 Pm
0
20 Pm
1. Sagittal sections of 21-day-old chick brain 24 hr after injection of a single dose of 0.2 mg/g of gold thioglucose, the sections were stained with hematoxylin/eosin. The distribution of the lesions as well as the histological details of the lesions are shown. Abbreviations used: BC, optic chiasma; BV, blood vessel; C, cerebellum; D, diencephalon; MO, medulla oblongata; PL, perivascular lesion; T, telencephalon. FIG.
GOLD
THIOGLUCOSE
227
IN CHICKS
LAYING
b
IO
20
30
40
50
60
70
so
90
100
110
l&J
DAYS AFTER COLDTHKXLUCOSE
130
140
INJECTIONS
150
1bu
(@2mg/g
170
180
190
ml
2x1 2x3
233
1
FIG. 2. The mean body weights (*SE) of the surviving 13 control birds(0. . . .e) and 12 experimental birds, which at 21-days-old had received a single injection of 0.2 mg/g of gold thioglucose (0-O). DISCUSSION
The highly variable nature of the mortality in the chick following injection of GTG is similar to that seen in quail (Carpenter et al., 1969) and is difficult to explain. The white Leghorn chicks used in the present experiment were more sensitive to GTG (100 % mortality with doses in excess of 0.2 mg/g) than the broiler chicks used by Simkins and Pensack (1970) and Svacha and Reid (1973), in which 0.5 mg/g allowed 62 % survival and no immediate mortality was found after doses of 0.4 mg/g. The failure of GTG to induce hyperphagia or obesity in the present experiment supports the results obtained by previous workers on birds (Carpenter et al., 1969; Simkins and Pensack, 1970). Moderate growth and appetite stimulation was reported by Svacha and Reid (1973) in male broiler chickens at 15 weeks of age, but they did not report whether these increases were significant. The birds in the present experiment were found to have a large number of small lesions in the central nervous system 24 hr after the injection of GTG, whereas both Carpenter et al. and Svacha and Reid found no damage in the central nervous system. This difference between broiler chicks (Svacha and Reid, 1973) and white Leghorns may be due to genetic factors similar to those used to explain the variability of response to GTG in different strains of mice (Liebelt and Perry, 1967). The finding of a large number of lesions in the brain of the chicken is similar to that in suckling mice, in which those surviving 1 or 2 days after injection with GTG showed
228
MICHAEL
J. GENTLE
necrotic foci in the spinal cord, medulla oblongata, hypothalamus, and limbic system (Perry, 1966). The extrahypothalamic distribution of lesions in mice treated with GTG may be associated with enhanced permeability of the blood-brain barrier, as described by Perry and Liebelt (1961). A similar explanation may apply to the results of the present experiment in which the blood-brain barrier is not fully developed in both the 2- and 21-day-old chicks and the lesions appeared to be perivascular. There seems to be general agreement that a GTG injection which results in massive obesity in mammals induces a lesion in the ventromedial region of the hypothalamus (Liebelt and Perry, 1967). In the birds used in the present study there were relatively few lesions in the ventromedial hypothalamus in the region where Lepkovsky and Yasuda (1966) were able to induce hyperphagia following electrolytic lesions. It therefore seems probable that the absence of hyperphagia in the birds was due to the failure of the GTG to induce lesions in this region. However. the binding of GTG to specific glucoreceptors, as envisaged by Mayer and Marshall (1956), cannot be ruled out by the diffuse nature of the lesions nor by the lack of any behavioral effects, for recent work on the chicken (Lepkovsky et al., 1965; Richardson, 1970) and the duck (Evans, 1969) suggeststhat glucose does not play a major role in the control of feeding. ACKNOWLEDGMENTS I wish to thank Dr. P. Moore-Robinson of the ScheringCorporation (EssexChemieAG, Switzerland)for thegenerousdonationof goldthioglucose,Mrs. I. Wilsonand MissI. Karlsson for their technicalassistance, and Dr. P. A. L. Wight for his help with the Weil technique. REFERENCES J. W., STEIN, C. M., SILVERSTEIN, A. AND VAN TIENHOVEN, A. (1969).The effect of gold thioglucoseon food consumptionand reproduction of the Japanesequail (Cotuvnix
CARPENTER,
coturnix japonica). EVANS,
Poult. Sci. 48, 574-578.
A. J. (1969). Some effects of protamine zinc insulin on juvenile ducks. J. Physiol.
(London)
203,84P-86P.
S., LEN, R., KOIKE, T. AND BONTHILET, R. (1965).Effects of protamine zinc insulin on chickens.Amer. J. Physiol. 208, 589-592. LEPKOVSKY, S.AND YASUDA, M. (1966).Hypothalamic lesions,growth and body composition of malechickens.Poult. Sci. 45, 582-588. LIEBELT, R. A. AND PERRY, J. H. (1967).Action of gold thioglucoseon the central nervous system.In Handbook ofPhysiology, Section 6: Alimentary canal (F. Code, Ed.), pp. 271286. American PhysiologicalSociety, Washington, D.C. MAYER, J. AND MARSHALL, M. B. (1956). Specificity of gold thioglucosefor ventromedial hypothalamic lesionsand hyperphagia.Nature (London) 178, 1399-1400. PERRY, J. H. (1966).Central nervouslesionsand weightgain in sucklingCBA/Ki micetreated with gold thioglucose.Anat. Rec. 154,400. PERRY, J. H. AND LIEBELT, R. A. (1961). Extra-hypothalamic lesionsassociatedwith gold thioglucoseinducedobesity. Proc. Sot. Exp. Biol. Med. 106, 55-57. RICHARDSON, A. I. (1970).Blood glucoselevelsand food intake in the domesticchicken.Brit. LEPKOVSKY,
Poult. Sci. 11, 501-504.
K. L. AND PENSACK, J. M. (1970).Effect ofgold thioglucoseon survival, feedconsumption and body weight of broilers.Poult. Sci. 49, 1341-1345. SVACHA, A. J. AND REID, B. L. (1973).Effect of gold thioglucosein the domesticfowl. P&t. SIMKINS,
Sci. 52, 926-930. A. (1946). Textbook
WEIL,
of Neuropathology.
Kimpton, London.
AND
APPLIED
PHARMACOLOGY
(1976)
35223-228
The Effects of Gold Thioglucose on the Central Nervous System of Chicks (Gal/us Domesticus) MICHAEL J. GENTLE Poultry Research Centre, Edinburgh EH9 3JS, Scotland Received March 13,1975; accepted August 241975
The Effectsof Gold Thioglucoseon the Central NervousSystemof Chicks GENTLE, M. J. (1976).Toxicol. Appl. Pharmacol. 35,223228. The effectsof gold thioglucoseon the central nervoussystem,growth, and laying performancewerestudiedfollowing a singleintramuscularinjection of the drug in 2- and21-day-old chicks(Gallus domesticus). Injection of gold thioglucosein dosesof 1-O.1mg/gbody weightgaveriseto a largenumber of small perivascularlesionsdistributed mainly in the telencephalon and diencephalon.None of the birds which recoveredfrom a singledoseof 0.2 mg/gof gold thioglucoseshowedanyeffect on growth rate or laying performancecomparedto the control birds. (GaZlus domesticus).
Single injections of an LD50 of gold thioglucose (GTG) have been shown to induce in several strains of mice and rats lesions in the ventromedial region of the hypothalamus
and in other areas of the brain (Liebelt and Perry, 1967). This hypothalamic damage results in the animal becoming hyperphagic and obese. Carpenter et al. (1969) found that in quail there was considerable variability in the LD50 of the drug and no evidence of either histological or functional damage to the hypothalamus. Simkins and Pensach (1970) and Svacha and Reid (1973) found LD50 values to be 0.5 mg/g body weight for 5- to 7-day-old broiler chicks. The last two authors reported that the surviving birds showed some impairment in the regulation of body temperature as well as moderate increasesin growth rate and food consumption but did not report whether these increases were significant. Svacha and Reid (1973) also found that GTG injected into mature white Leghorn hens did not influence body weight or food intake. No changes in food intake or brain damage were found in brown Leghorn males (Wood-Gush and Gower, personal communication) following the injection of GTG in doses from 0.3 to 1.1 mg/g. One hypothesis for the mode of topic action of the drug in mammals is that the glucose becomes attached to specific glucoreceptor neurons in the hypothalamus and, because of the high toxicity of the drug, cell death results (Mayer and Marshall, 1956). One reason for the failure to find brain damage in adult birds following the injection of GTG may be due to a failure of the GTG to penetrate the blood-brain barrier. To avoid this difficulty chicks were used in the present experiment in order to study effects induced before the blood-brain barrier is fully developed. Previous reports using GTG in chicks Copyright 0 1976 by Academic Press, Inc. All rights of reproduction in any form reserved. Printed in Great Britain
223
MlCHAELJ.GENTLE
224
(Simkins and Pensach, 1970: Svacha and Reid, 1973) are confined to broiler strains. Broiler chickens have been specially bred to have a high growth rate, so that it might be difficult to induce hyperphagia becausethey might already be near their upper limit of growth and food intake. In the present experiment the birds usedwere from a laying strain which had not been bred for a fast growth rate and would hence have more potential to increasetheir food intake.
METHODS The birds were female white Leghorn chicks which were bred at the Poultry Research Centre. They were housed in Perspex boxes under continuous light and maintained at 33°C by using infrared heaters. Food and water were always available. The number of birds used, their ages,and the dosesof gold thioglucose injected are shown in Table 1. Two groups of birds were injected at 2 and 21 days of age and were TABLE 1 SUMMARYOFTHENUMBEROF
CHICKS USED,THEIRAGE,THEDRUGDOSAGEANDTHESUBSEQIJENTFATEOFTHECHICKS
Number injected Total number Age of birds used (days) Drug injected and dose at eachdose 20
10
2 0.1-1.0 mg/g GTG in O.lmg steps 21 0.1-1.0 mg/g GTG in O.lmg steps 2 Salinecontrol
10
10
21 Salinecontrol
10
45
21 0.2 mg/g GTG
45
21
21 Salinecontrol
21
20
2 2
Treatment Died or killed in 24 hr, brain removed Died or killed in 24 hr, brain removed Killed in 24 hr, brain removed Killed in 24 hr, brain removed Body weight recorded at IO-day intervals up to 230 days Body weight recorded at IO-day intervals up to 230days
not allowed to survive for more than 24 hr after injection. These birds were used to determine the LD50 of the drug and to study its effects on the central nervous system. Each bird was given a single im injection of either GTG in oil (Solganal, Schering Corporation) or saline.To determine the approximate LD50 value, two birds were injected with the samedoseof the drug at 2 days and another group at 21 days of age. The dosesranged from 0.1 to 1.Omg/g body weight in O.l-mg steps.To study the effects of the GTG on the central nervous system, the surviving birds were killed after 24 hr using ether, and the brains of those that had died as well as those that were killed were dis-
GOLD
THIOGLUCOSE
IN CHICKS
225
sected out and fixed. Half the brains, i.e., one for each age and concentration, were fixed in aqueous Bouin and the other half in 10 % formol-saline. Those fixed in Bouin were embedded in paraffin wax, sectioned at 8 pm and stained with hematoxylin/eosin to study the general anatomy. Those fixed in formol-saline were frozen, cut at 50 pm, and stained using the Weil (1946) technique to look for demyelination. The third group of birds was injected with either 0.2 mg/g of GTG or saline at 21 days of age, and the animals were allowed to live for 230 days in order to see if the brain damage induced by the drug had any effect on the birds’ growth or laying performance. The birds were injected at 21 days of age because there is normally a higher mortality in the younger chicks. The use of 45 birds in the experimental group was to ensure equal numbers of control and experimental birds at the end of the 230-day period.
RESULTS
Mortality All birds injected at 2 days of age with doses of 0.3 mg/g or more became very sick or died within 24 hr. In the al-day-old birds the results were similar. Of the 45 birds injected with 0.2 mg/g, 20 died within 3 days of injection and a further 12 died or were killed in extremis during the next 230 days. In the 21-day-old control birds none died in the first 3 days following injection, but six died or were killed before reaching 230 days of age. It was not possible to determine the LD50 with any accuracy, as the mortality varied from group to group. The 45 birds that were injected at 21 days of age with 0.2 mg/g were injected in small groups. The groups consisted of 5, 12, 14, and 14 birds, of which 1, 6, 11, and 2 died within 3 days after injection. Pathological Changes Damage to the central nervous system following the injection of GTG was very extensive and took the form of a large number of small lesions. These were distributed around blood vessels (Fig. 1). The lesions consisted of a focal disseminated edema in the neuropile of the grey matter. The nuclei of the cells in the lesion area were smaller, irregularly shaped, and more deeply stained than the normal cells, indicating cell death, so that there was perivascular necrosis in addition to the edema. They were concentrated mainly in the telencephalon and anterior diencephalon. Fewer lesions were found in the posterior diencephalon. Although there were some lesions in the cerebellum there were few in the mesencephalon and medulla oblongata. The distribution of the lesions reflected that of the fine capillary blood supply to the brain. The extent of the lesions depended upon the dose of the drug and was greatest at high doses but was still extensive at 0.2 mg/g (Fig. 1). There were no clear pathological differences between the 2- and 21day-old chicks. There was no clear evidence of demyelination. Growth, Sexual Maturity, and Rate of Lay The mean weights of the surviving 13 control birds and 12 treated with GTG are shown in Fig. 2; there were no significant differences. Nor was there any significant
226
MICHAEL
J. GENTLE
difference in the date of onset of laying (mean values of 155.8 k 5.8 and 159.2 + 7.5 days) or in the number of eggs laid in the 25-day period after the onset of lay (means 12.1 & 1.6 and 11.5 & 1.4eggs).
PL
0
100 Pm
0
20 Pm
1. Sagittal sections of 21-day-old chick brain 24 hr after injection of a single dose of 0.2 mg/g of gold thioglucose, the sections were stained with hematoxylin/eosin. The distribution of the lesions as well as the histological details of the lesions are shown. Abbreviations used: BC, optic chiasma; BV, blood vessel; C, cerebellum; D, diencephalon; MO, medulla oblongata; PL, perivascular lesion; T, telencephalon. FIG.
GOLD
THIOGLUCOSE
227
IN CHICKS
LAYING
b
IO
20
30
40
50
60
70
so
90
100
110
l&J
DAYS AFTER COLDTHKXLUCOSE
130
140
INJECTIONS
150
1bu
(@2mg/g
170
180
190
ml
2x1 2x3
233
1
FIG. 2. The mean body weights (*SE) of the surviving 13 control birds(0. . . .e) and 12 experimental birds, which at 21-days-old had received a single injection of 0.2 mg/g of gold thioglucose (0-O). DISCUSSION
The highly variable nature of the mortality in the chick following injection of GTG is similar to that seen in quail (Carpenter et al., 1969) and is difficult to explain. The white Leghorn chicks used in the present experiment were more sensitive to GTG (100 % mortality with doses in excess of 0.2 mg/g) than the broiler chicks used by Simkins and Pensack (1970) and Svacha and Reid (1973), in which 0.5 mg/g allowed 62 % survival and no immediate mortality was found after doses of 0.4 mg/g. The failure of GTG to induce hyperphagia or obesity in the present experiment supports the results obtained by previous workers on birds (Carpenter et al., 1969; Simkins and Pensack, 1970). Moderate growth and appetite stimulation was reported by Svacha and Reid (1973) in male broiler chickens at 15 weeks of age, but they did not report whether these increases were significant. The birds in the present experiment were found to have a large number of small lesions in the central nervous system 24 hr after the injection of GTG, whereas both Carpenter et al. and Svacha and Reid found no damage in the central nervous system. This difference between broiler chicks (Svacha and Reid, 1973) and white Leghorns may be due to genetic factors similar to those used to explain the variability of response to GTG in different strains of mice (Liebelt and Perry, 1967). The finding of a large number of lesions in the brain of the chicken is similar to that in suckling mice, in which those surviving 1 or 2 days after injection with GTG showed
228
MICHAEL
J. GENTLE
necrotic foci in the spinal cord, medulla oblongata, hypothalamus, and limbic system (Perry, 1966). The extrahypothalamic distribution of lesions in mice treated with GTG may be associated with enhanced permeability of the blood-brain barrier, as described by Perry and Liebelt (1961). A similar explanation may apply to the results of the present experiment in which the blood-brain barrier is not fully developed in both the 2- and 21-day-old chicks and the lesions appeared to be perivascular. There seems to be general agreement that a GTG injection which results in massive obesity in mammals induces a lesion in the ventromedial region of the hypothalamus (Liebelt and Perry, 1967). In the birds used in the present study there were relatively few lesions in the ventromedial hypothalamus in the region where Lepkovsky and Yasuda (1966) were able to induce hyperphagia following electrolytic lesions. It therefore seems probable that the absence of hyperphagia in the birds was due to the failure of the GTG to induce lesions in this region. However. the binding of GTG to specific glucoreceptors, as envisaged by Mayer and Marshall (1956), cannot be ruled out by the diffuse nature of the lesions nor by the lack of any behavioral effects, for recent work on the chicken (Lepkovsky et al., 1965; Richardson, 1970) and the duck (Evans, 1969) suggeststhat glucose does not play a major role in the control of feeding. ACKNOWLEDGMENTS I wish to thank Dr. P. Moore-Robinson of the ScheringCorporation (EssexChemieAG, Switzerland)for thegenerousdonationof goldthioglucose,Mrs. I. Wilsonand MissI. Karlsson for their technicalassistance, and Dr. P. A. L. Wight for his help with the Weil technique. REFERENCES J. W., STEIN, C. M., SILVERSTEIN, A. AND VAN TIENHOVEN, A. (1969).The effect of gold thioglucoseon food consumptionand reproduction of the Japanesequail (Cotuvnix
CARPENTER,
coturnix japonica). EVANS,
Poult. Sci. 48, 574-578.
A. J. (1969). Some effects of protamine zinc insulin on juvenile ducks. J. Physiol.
(London)
203,84P-86P.
S., LEN, R., KOIKE, T. AND BONTHILET, R. (1965).Effects of protamine zinc insulin on chickens.Amer. J. Physiol. 208, 589-592. LEPKOVSKY, S.AND YASUDA, M. (1966).Hypothalamic lesions,growth and body composition of malechickens.Poult. Sci. 45, 582-588. LIEBELT, R. A. AND PERRY, J. H. (1967).Action of gold thioglucoseon the central nervous system.In Handbook ofPhysiology, Section 6: Alimentary canal (F. Code, Ed.), pp. 271286. American PhysiologicalSociety, Washington, D.C. MAYER, J. AND MARSHALL, M. B. (1956). Specificity of gold thioglucosefor ventromedial hypothalamic lesionsand hyperphagia.Nature (London) 178, 1399-1400. PERRY, J. H. (1966).Central nervouslesionsand weightgain in sucklingCBA/Ki micetreated with gold thioglucose.Anat. Rec. 154,400. PERRY, J. H. AND LIEBELT, R. A. (1961). Extra-hypothalamic lesionsassociatedwith gold thioglucoseinducedobesity. Proc. Sot. Exp. Biol. Med. 106, 55-57. RICHARDSON, A. I. (1970).Blood glucoselevelsand food intake in the domesticchicken.Brit. LEPKOVSKY,
Poult. Sci. 11, 501-504.
K. L. AND PENSACK, J. M. (1970).Effect ofgold thioglucoseon survival, feedconsumption and body weight of broilers.Poult. Sci. 49, 1341-1345. SVACHA, A. J. AND REID, B. L. (1973).Effect of gold thioglucosein the domesticfowl. P&t. SIMKINS,
Sci. 52, 926-930. A. (1946). Textbook
WEIL,
of Neuropathology.
Kimpton, London.