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Malic enzyme activity in the developing rat brain in relation to thyroid status
Ins.f. Devf. Neuroscience. Voi. 7, No. 2, pp. 203-208.1989. Printed in Great Britain.
073&5748/89 $03.00+0.00 PergamonPresspk. @ 1989ISDN
MALIC ENZYME ACTIVITY IN THE DEVELOPING RAT BRAIN IN RELATION TO THYROID STATUS U. R.
THAKARE,
Radiation Medicine Centre, BARC,
D. H. SHAH and
U.
VKAYAN
Tata Memorial Centre Annexe. Parel, Bombay-400 012, India
(Received 5 Mny 1988; revised 13 October 1988; accepred14 October 1988)
Abatraet-Malic enzyme activity in the soluble fraction of neonate brains from mothers fed with propyithiouracil (0.015% w/v) in drinking water from day 12 of the gestation period was significantly lowered (PcO.01) as compared to the offspring of normal mothers. Supplementation of triiodothyronine to the neonates from experimental mothers restored the malic enzyme activity to normal levels. However, administration of triiodothyronine to adult control rats did not influence malic enzyme activity in the brains of these animals. Our data suggest that during the initial critical period of brain maturation, malic enzyme is under the control of thyroid hormones. The response of malic enzyme towards thyroid hormones is lost once the brain has matured. Key WO&:
Malie enzyme, Neonate, Triiodothyronine,
Propylthiouracil.
Involvement of thyroid hormones in development, differentiation and maturation of the brain has been well-documented. Shanker et af. I3 have clearly demonstrated the importance of a critical period in early development of the brain of mice when there is an absolute dependency on thyroid hormone for proper myelination as well as for normal growth and differentiation.3 The brains of hypothyroid rats are known to be deficient in myelin lipids.‘*‘” Malic enzyme (ME) is one of the sets of four to six enzymes whose activity correlates positively with the rate of de novo synthesis of fatty acids. ‘**This enzyme is believed to be involved in fatty acid saturation and elongation. The precise role of ME in affecting thyroid hormone action at the cellular level is unknown. Although enhanced hepatic ME activity is a measure of hormonal action in liver, brain ME activity of an adult rat is unresponsive to thyroid hormones.” We report here the activity of ME (EC 1.1.1.40 malate NADP+ oxidoreductase enyzme) during development of neonate brains from normal and hypothyroid mothers. We have further evaluated the response of this enzyme to the supplementation of triiodothyronine (T3) to the neonates from mothers fed with propylthiouracil (PTU). EXPERIMENTAL
PROCEDURES
Chemicals
Malic acid and NADP were obtained from Sigma Chemicals, U.S.A. and ~-propylthioura~il (PTU) was a product of Fluka Laboratories, Switzerland. Method
Wistar rats weighting about 200-300 g were used for mating and immediately after conception the mothers were separated into control and experimental groups with each animal housed indi~dually. The mothers in the experimental group were given PTU (0.015% w/v) in their drinking water from day 12 of the gestation period and after delivery until the litters were killed as described by Wysocki and Segal. ” Estimation
of malic enzyme
activity in the developing
liver and brain
Litters from control and experimental groups were killed at 9,15,20,25 and 30 days after birth. Livers and brains were removed, weighed and homogenized as described previously.” Malic enzyme activity in the soluble fraction of the homogenate was measured according to the method Abbreviations: ME, Tj, triiodothyronine.
malic enzyme; NADP,
nicotinamide adenine dinucleotide phosphate; PTU,
203
propylthiouracil;
204
U. R. Thakare
et al.
of Hsu and Lardy.h ME activity was expressed an nmoles of NADPH produced/min/mg All the litters were monitored on alternate days after birth for weight gain.
protein.
Response of ME to T3 Half the number of neonates from experimental and control groups of mothers were administered TJ (0.5 kg/g body wt; dissolved in 0.1 N NaOH and diluted with saline to the required volume) intraperitoneally on days 2, 4, 6 and 8 after birth and the other half, serving as pair matched controls, were administered solvent (0.1 N NaOH-saline) alone on respective days. On day 9 all the neonates were killed, the livers and brains removed. weighed, homogenized and assayed for ME activity as described previously. I4 Proteins were estimated according to the method of Lowry et al.” Results were expressed as mean -+ S.D. Statistical analysis of the results was performed according to Student’s t-test. Two-way analysis of variance at significance level of PC 0.01 was applied for the growth of control and experimental neonates.
RESULTS
Body weight As early as 2 days after birth (Fig. 1) the mean weight of neonates from the experimental group of mothers was significantly (P~0.01) lowered (5.26kO.59 g; n =30) as compared to controls (8.06 + 1.27 g; n = 30). The difference became more marked on day 15 after birth when the mean body weight of the controls was 37.67 + 2.52 g as compared to a value of 16.0 + 1.55 g in neonates from the experimental group of mothers. On day 30 the difference was further enhanced (P
Liver weight The liver weight in the two groups of neonates differed significantly throughout the period of the study (PC 0.001). The weight of livers from control neonates was 0.47 ? 0.015 g (n = 20) on day 9 after birth, increasing to 4.01+ 0.61 g over a period of 30 days, while during the same period liver weights of neonates in the experimental group of mothers showed a marginal increase from 0.37 + 0.042 g to 0.89 rt 0.089 g (n = 20) (Table 1).
Brain weight The weight of neonate brains from control and experimental groups did not differ significantly on day 9 after birth as shown in Table 1 (0.661tO.063 g in control group and 0.610 kO.053 g in experimental group; n = 20). However, during the next 21 days the difference in brain weight increased significantly (l.lSO-+ 0.033 g in the experimental group and 1.570 2 0.083 g in the control group; P-CO.01).
Days after
birth
Fig. 1. A graphical representation of the weight of control and experimental neonates as a function days after birth. The experimental protocol followed is as described in the text. O-----O CON, D-_cI PTU. Each value is a mean of 30 animals.
of
*rr = Number
of animals.
0.038
0.048
0.37 k 0.042
0.47 k 0.015
Liver wt
Brain wt/ body wt
Brain wt (n=20)8
0.053
PTU
0.612
9 days
0.057
0.077
0.39kO.032
0.70 -t 0.014
0.038
PTU
CON
0.038
1.6820.12
F+Tu
o.u72
0.54~0.084
CON
0.022
3.55kO.062
0.061
0.72kO.097
1.22kOGt3
PTU
at different
25 days
mothers
1.47+.0&W
and PTU-fed
1.17-tO.038
20 days
from control-
1.4220.035
of the neonates
I.112
15 days
in grams,
1.37 20.035
CON
1. Brain and liver weight,
0.66 2 0.063
CON
Table
0.018
PTU
0.054
0.89 kO.089
1.18kO.033
30 days
4.01 _‘O.tXl
1.5720.083
CON
days after birth
206
U.
R. Thakare
et al.
iaoa = Liver
b = brain
2 2 g 140E”
5
10
15 days
20 aftor
25
30
birth
5
10
15 day8
20 attar
25
30
birth
Fig. 2. Malic enzyme activity in liver and brain during development of neonates from control and experimental groups of mothers are expressed as a function of days after birth. (a) Liver. (b) Brain. G----0 CON, X-X PTU. Expressed value is a mean of 18 values. Bars = 2 1 S.D.
ME activity in the liver and brain during development ME activity in the liver. The effectiveness of the hypothyroid status of the neonates has been clearly demonstrated in Fig. 2a where hepatic ME activity in neonates from PTU-fed mothers remained negligible throughout the period of study. Hepatic ME activity in the neonates from the control group of mothers remained undetected for 20 days after which there was a sharp rise reaching a value of 177.64 * 35.17 nmoles of NADPH/min/mg protein. ME activity in the brain. ME activity in the neonate brains from the control and experimental groups has been depicted in Fig. 2b. There was a gradual increase in the enzyme activity from day 9 after birth with a value of 2.54 4 0.33 nmoles of NADPWmin/mg protein, to 16.54 f 1.36 nmoles of NADPH produced/min/mg protein on day 30 in neonates from control mothers, while there was a slow rise in activity in the neonate brains from experimental mothers ranging from 1.77 2 0.57 nmoles of NADPH/min/mg protein on day 9 to 8.612 1.15 nmoles of NADPWmin/mg protein on day 30 after birth (P~0.01). Response
of ME to T3 in the liver and brain
Supplementation of T3 on days 2, 4, 6 and 8 to control and experimental groups of neonates, enhanced hepatic ME activity when assayed on day 9 after birth (22.95 k 0.20 and 25.48 f. 0.24 nmoles NADPWmin/mg protein in control and experimental groups, respectively) as compared to undetected values of ME observed in livers of their respective controls (Fig. 3a). Administration of T3 to adult rats also enhanced hepatic ME activity as depicted in Fig. 3a. Administration of T3 also influenced ME activity in the neonate brains from control and experimental groups of mothers (Fig. 3b). Enzyme activity in neonate brains from PTU-fed mothers was restored to normal levels after supplementation of TS to these neonates (1.77 rt 0.57 and 2.47 -t 0.08 nmoles of NADPH produced/min/mg protein in experimental and Tj supplemented hypothyroid groups, respectively). However, T3 administration did not influence the ME activity in the brains of adult rats.
207
Malic enzyme in the developing brain
264
b = broin
a=liver
r
24
-1
g 601 E 2
50-
2 t 40_=
P P +
i = xl-
i
B
20
10 I
L-_ilJ_ 5
0
:
-
Neonates
Neonotes
Adult
Fig. 3. Histograms represent ME activity on day 9 after birth in brains and livers of control, hypothyroid and T3 supplemented neonatal rats and control and T3 administered adult rats. The results are expressed as a mean of 18 values 2 1 SD.
DISCUSSION Thyroid hormones have long been recognized as very important regulators of brain development and function. However, the variety of metabolic and hormonal effects induced by thyroid hormones in vivo makes it difficult to pinpoint which observations represent direct effects of thyroid hormone and which are indirect. Our findings of a significantly lowered body weight of experimental neonates at the time of birth indicate the adverse effect of PTU on total growth, an issue which has been controversial.’ Nevertheless, growth after birth is reported to be thyroid hormone dependent.” It is interesting to note that the weights of the brains of both groups of animals were comparable. However, the overall decrease in the weight of the brain in experimental neonates is significant as compared to control neonates as calculated by two-way analysis of variance (P~0.01). Hence, the ratio between the brain weight and body weight is relatively higher (0.048) in experimental neonates than that of control neonates (0.038) as indicated in Table 1. With age this ratio also increased but remained high in the experimental group as compared to the control group because of the lowered body weights in the former group. We have reported earlier I4 that brain ME activity is thyroid hormone dependent in the early phase of brain development and this is confirmed in this paper. Similar findings have been reported by Diez Guerra et ~1.~The decrease in ME activity in neonate brains from the experimental group of mothers and restoration of the activity after supplementation of T3 to these neonates, as well as no response of ME in the brains of normal adult rats after administration of T, to these animals, indicates dependency of ME on thyroid hormones during the period of brain development. However, this enzyme activity in the brain does not seem to be entirely dependent on thyroid hormone as a basal level of ME has been detected on day 9 after birth in neonates from the experimental group of mothers. On the other hand, hepatic ME activity appears to be completely under the control of T3 (Fig. 2a) as a negligible amount of hepatic enzyme activity was observed in neonates from PTU-fed mothers during the period of study. The relationship between thyroid status and hepatic ME activity has been extensively studied by several investigators.“6*1” Oppenheimer et ~1.” and Hemon’ however, have failed to show ME response in the neonate brains on supplementation with thyroid hormones.
208
U. R. Thakare
ef al.
Although no clearly defined function for malic enzyme has been established in the brain, this enzyme plays an important role in lipogenesis through the production of NADPH required for fatty acid synthesis and hence, for myelination process. Increase in hepatic ME activity at the time of weaning (day 21) has been related to its role in lipid biosynthesis.” Decreased activity of this enzyme in the brain during the developmental period might therefore affect lipogenesis which may lead to impairment of myelination in the brain and hence, dysfunction. Studies are in progress to correlate ME activity and myelination in the neonate brain during development. REFERENCES 1. Back D. W., Wilson S. B., Norris S. M. and Goodridge A. G. (1986) Hormonal regulation of lipogenic enzymes in chick embryo hepatocytes in culture. /. biof. Chem. 261, 12555-12561. 2. Balazs R., Brooksbank B. W. L., Davison A. N., Eayrs J. T. and Wilson D. A. (1%9) The effect of neonatal thyroidectomy on myelination in the rat brain. Brain Res. 15, 219-232. 3. Bass N. H., Pelton E. W. and Young E. (1977) Thyroid Hormones and Bruin Development (ed. Grave G. D.), pp. 199-214. Raven Press, New York. 4. Diez Guerra J., Aragon M. C., Gimenez C. and Valdivieso F. (1981) Effect of thyroid hormones on the Malic enzyme activity in rat brain during development. Devl Neurosci. 4, 130-133. in the 5. Hemon P. (1968) Malate dehydrogenase (decarboxylating) (NADPH) and glycerophosphateoxidase developing rat. Biochim. Biophys. Acta 151, 681-683. 6. Hsu R. Y. and Lardy H. A. (1969) Malic enzyme. Merh. Enzym. 13, 230-235. 7. Liu L. and Nicoll C. S. (1986) Postnatal development of dependence on thyroid hormones for growth and differentiation of rat skeletal structures. Growth 50, 472484. 8. Lobato M. F., Francisco M. R., Moreno J. and Garcia-Ruiz J. P. (1986) Nutritional and hormonal regulation of malic enzyme synthesis in rat mammary gland. Biochem. J. 236, 4414l5. 9. Lowry 0. H.. Rosebrough N. J., Farr A. L. and Randall R. A. (1951) Protein measurement with the Folin phenol reagent. 1. biol. Chem. 193.263-275. J. M.. Silva E.. Schwartz H. L. and Surks M. I. (1977) Stimulation of hepatic 10. Oooenheimer &&hondrial-glycerophospate dehydrogenase and Malic enzyme by L-triiodothyronine. J. Clin Invest. 59, 517-527. 11. Schwartz H. L. (1983) Molecular Basis of Thyroid Hormone Action (eds Oppenheimer J. H. and Samuels H. H.), p. 414. Academic Press, New York. 12. Schwartz H. L. and Oppenheimer J. H. (1978) Ontogenesis of 3.5,3’-triiodothyronine receptors in neonatal rat brain. Endocrinology
103,943-948.
13. Shanker G., Amur S. G. and Pieringer R. A. (1985) Investigations on myelinogenesis in vitro. Neurochem. Res. 10, 617-625. 14. Thakare U. R., Vijayan U., Sushila L. and Shah D. H. (1987) Malic enzyme response to triiodothyronine in the brains of neonatal rats. Life Sci. 41, 2823-2826. 15. Vernon R. G. and Walker D. G. (1986) Changes in activity of some enzymes involved in glucose utilization and formation in developing rat liver. Bioche’m. J. lii6, 321-329. _ 16 Walravens P. and Chase H. P. (1969) Influence of thyroid hormone on formation of myelin lipid. J. Neurochem. 16, 1477-1484. 17. Wysocki S. J. and Segal W. (1972) Influence of thyroid hormones on enzyme activities of myelinating rat central nervous tissues. Eur. J. Biochem. 28, 183-189.
073&5748/89 $03.00+0.00 PergamonPresspk. @ 1989ISDN
MALIC ENZYME ACTIVITY IN THE DEVELOPING RAT BRAIN IN RELATION TO THYROID STATUS U. R.
THAKARE,
Radiation Medicine Centre, BARC,
D. H. SHAH and
U.
VKAYAN
Tata Memorial Centre Annexe. Parel, Bombay-400 012, India
(Received 5 Mny 1988; revised 13 October 1988; accepred14 October 1988)
Abatraet-Malic enzyme activity in the soluble fraction of neonate brains from mothers fed with propyithiouracil (0.015% w/v) in drinking water from day 12 of the gestation period was significantly lowered (PcO.01) as compared to the offspring of normal mothers. Supplementation of triiodothyronine to the neonates from experimental mothers restored the malic enzyme activity to normal levels. However, administration of triiodothyronine to adult control rats did not influence malic enzyme activity in the brains of these animals. Our data suggest that during the initial critical period of brain maturation, malic enzyme is under the control of thyroid hormones. The response of malic enzyme towards thyroid hormones is lost once the brain has matured. Key WO&:
Malie enzyme, Neonate, Triiodothyronine,
Propylthiouracil.
Involvement of thyroid hormones in development, differentiation and maturation of the brain has been well-documented. Shanker et af. I3 have clearly demonstrated the importance of a critical period in early development of the brain of mice when there is an absolute dependency on thyroid hormone for proper myelination as well as for normal growth and differentiation.3 The brains of hypothyroid rats are known to be deficient in myelin lipids.‘*‘” Malic enzyme (ME) is one of the sets of four to six enzymes whose activity correlates positively with the rate of de novo synthesis of fatty acids. ‘**This enzyme is believed to be involved in fatty acid saturation and elongation. The precise role of ME in affecting thyroid hormone action at the cellular level is unknown. Although enhanced hepatic ME activity is a measure of hormonal action in liver, brain ME activity of an adult rat is unresponsive to thyroid hormones.” We report here the activity of ME (EC 1.1.1.40 malate NADP+ oxidoreductase enyzme) during development of neonate brains from normal and hypothyroid mothers. We have further evaluated the response of this enzyme to the supplementation of triiodothyronine (T3) to the neonates from mothers fed with propylthiouracil (PTU). EXPERIMENTAL
PROCEDURES
Chemicals
Malic acid and NADP were obtained from Sigma Chemicals, U.S.A. and ~-propylthioura~il (PTU) was a product of Fluka Laboratories, Switzerland. Method
Wistar rats weighting about 200-300 g were used for mating and immediately after conception the mothers were separated into control and experimental groups with each animal housed indi~dually. The mothers in the experimental group were given PTU (0.015% w/v) in their drinking water from day 12 of the gestation period and after delivery until the litters were killed as described by Wysocki and Segal. ” Estimation
of malic enzyme
activity in the developing
liver and brain
Litters from control and experimental groups were killed at 9,15,20,25 and 30 days after birth. Livers and brains were removed, weighed and homogenized as described previously.” Malic enzyme activity in the soluble fraction of the homogenate was measured according to the method Abbreviations: ME, Tj, triiodothyronine.
malic enzyme; NADP,
nicotinamide adenine dinucleotide phosphate; PTU,
203
propylthiouracil;
204
U. R. Thakare
et al.
of Hsu and Lardy.h ME activity was expressed an nmoles of NADPH produced/min/mg All the litters were monitored on alternate days after birth for weight gain.
protein.
Response of ME to T3 Half the number of neonates from experimental and control groups of mothers were administered TJ (0.5 kg/g body wt; dissolved in 0.1 N NaOH and diluted with saline to the required volume) intraperitoneally on days 2, 4, 6 and 8 after birth and the other half, serving as pair matched controls, were administered solvent (0.1 N NaOH-saline) alone on respective days. On day 9 all the neonates were killed, the livers and brains removed. weighed, homogenized and assayed for ME activity as described previously. I4 Proteins were estimated according to the method of Lowry et al.” Results were expressed as mean -+ S.D. Statistical analysis of the results was performed according to Student’s t-test. Two-way analysis of variance at significance level of PC 0.01 was applied for the growth of control and experimental neonates.
RESULTS
Body weight As early as 2 days after birth (Fig. 1) the mean weight of neonates from the experimental group of mothers was significantly (P~0.01) lowered (5.26kO.59 g; n =30) as compared to controls (8.06 + 1.27 g; n = 30). The difference became more marked on day 15 after birth when the mean body weight of the controls was 37.67 + 2.52 g as compared to a value of 16.0 + 1.55 g in neonates from the experimental group of mothers. On day 30 the difference was further enhanced (P
Liver weight The liver weight in the two groups of neonates differed significantly throughout the period of the study (PC 0.001). The weight of livers from control neonates was 0.47 ? 0.015 g (n = 20) on day 9 after birth, increasing to 4.01+ 0.61 g over a period of 30 days, while during the same period liver weights of neonates in the experimental group of mothers showed a marginal increase from 0.37 + 0.042 g to 0.89 rt 0.089 g (n = 20) (Table 1).
Brain weight The weight of neonate brains from control and experimental groups did not differ significantly on day 9 after birth as shown in Table 1 (0.661tO.063 g in control group and 0.610 kO.053 g in experimental group; n = 20). However, during the next 21 days the difference in brain weight increased significantly (l.lSO-+ 0.033 g in the experimental group and 1.570 2 0.083 g in the control group; P-CO.01).
Days after
birth
Fig. 1. A graphical representation of the weight of control and experimental neonates as a function days after birth. The experimental protocol followed is as described in the text. O-----O CON, D-_cI PTU. Each value is a mean of 30 animals.
of
*rr = Number
of animals.
0.038
0.048
0.37 k 0.042
0.47 k 0.015
Liver wt
Brain wt/ body wt
Brain wt (n=20)8
0.053
PTU
0.612
9 days
0.057
0.077
0.39kO.032
0.70 -t 0.014
0.038
PTU
CON
0.038
1.6820.12
F+Tu
o.u72
0.54~0.084
CON
0.022
3.55kO.062
0.061
0.72kO.097
1.22kOGt3
PTU
at different
25 days
mothers
1.47+.0&W
and PTU-fed
1.17-tO.038
20 days
from control-
1.4220.035
of the neonates
I.112
15 days
in grams,
1.37 20.035
CON
1. Brain and liver weight,
0.66 2 0.063
CON
Table
0.018
PTU
0.054
0.89 kO.089
1.18kO.033
30 days
4.01 _‘O.tXl
1.5720.083
CON
days after birth
206
U.
R. Thakare
et al.
iaoa = Liver
b = brain
2 2 g 140E”
5
10
15 days
20 aftor
25
30
birth
5
10
15 day8
20 attar
25
30
birth
Fig. 2. Malic enzyme activity in liver and brain during development of neonates from control and experimental groups of mothers are expressed as a function of days after birth. (a) Liver. (b) Brain. G----0 CON, X-X PTU. Expressed value is a mean of 18 values. Bars = 2 1 S.D.
ME activity in the liver and brain during development ME activity in the liver. The effectiveness of the hypothyroid status of the neonates has been clearly demonstrated in Fig. 2a where hepatic ME activity in neonates from PTU-fed mothers remained negligible throughout the period of study. Hepatic ME activity in the neonates from the control group of mothers remained undetected for 20 days after which there was a sharp rise reaching a value of 177.64 * 35.17 nmoles of NADPH/min/mg protein. ME activity in the brain. ME activity in the neonate brains from the control and experimental groups has been depicted in Fig. 2b. There was a gradual increase in the enzyme activity from day 9 after birth with a value of 2.54 4 0.33 nmoles of NADPWmin/mg protein, to 16.54 f 1.36 nmoles of NADPH produced/min/mg protein on day 30 in neonates from control mothers, while there was a slow rise in activity in the neonate brains from experimental mothers ranging from 1.77 2 0.57 nmoles of NADPH/min/mg protein on day 9 to 8.612 1.15 nmoles of NADPWmin/mg protein on day 30 after birth (P~0.01). Response
of ME to T3 in the liver and brain
Supplementation of T3 on days 2, 4, 6 and 8 to control and experimental groups of neonates, enhanced hepatic ME activity when assayed on day 9 after birth (22.95 k 0.20 and 25.48 f. 0.24 nmoles NADPWmin/mg protein in control and experimental groups, respectively) as compared to undetected values of ME observed in livers of their respective controls (Fig. 3a). Administration of T3 to adult rats also enhanced hepatic ME activity as depicted in Fig. 3a. Administration of T3 also influenced ME activity in the neonate brains from control and experimental groups of mothers (Fig. 3b). Enzyme activity in neonate brains from PTU-fed mothers was restored to normal levels after supplementation of TS to these neonates (1.77 rt 0.57 and 2.47 -t 0.08 nmoles of NADPH produced/min/mg protein in experimental and Tj supplemented hypothyroid groups, respectively). However, T3 administration did not influence the ME activity in the brains of adult rats.
207
Malic enzyme in the developing brain
264
b = broin
a=liver
r
24
-1
g 601 E 2
50-
2 t 40_=
P P +
i = xl-
i
B
20
10 I
L-_ilJ_ 5
0
:
-
Neonates
Neonotes
Adult
Fig. 3. Histograms represent ME activity on day 9 after birth in brains and livers of control, hypothyroid and T3 supplemented neonatal rats and control and T3 administered adult rats. The results are expressed as a mean of 18 values 2 1 SD.
DISCUSSION Thyroid hormones have long been recognized as very important regulators of brain development and function. However, the variety of metabolic and hormonal effects induced by thyroid hormones in vivo makes it difficult to pinpoint which observations represent direct effects of thyroid hormone and which are indirect. Our findings of a significantly lowered body weight of experimental neonates at the time of birth indicate the adverse effect of PTU on total growth, an issue which has been controversial.’ Nevertheless, growth after birth is reported to be thyroid hormone dependent.” It is interesting to note that the weights of the brains of both groups of animals were comparable. However, the overall decrease in the weight of the brain in experimental neonates is significant as compared to control neonates as calculated by two-way analysis of variance (P~0.01). Hence, the ratio between the brain weight and body weight is relatively higher (0.048) in experimental neonates than that of control neonates (0.038) as indicated in Table 1. With age this ratio also increased but remained high in the experimental group as compared to the control group because of the lowered body weights in the former group. We have reported earlier I4 that brain ME activity is thyroid hormone dependent in the early phase of brain development and this is confirmed in this paper. Similar findings have been reported by Diez Guerra et ~1.~The decrease in ME activity in neonate brains from the experimental group of mothers and restoration of the activity after supplementation of T3 to these neonates, as well as no response of ME in the brains of normal adult rats after administration of T, to these animals, indicates dependency of ME on thyroid hormones during the period of brain development. However, this enzyme activity in the brain does not seem to be entirely dependent on thyroid hormone as a basal level of ME has been detected on day 9 after birth in neonates from the experimental group of mothers. On the other hand, hepatic ME activity appears to be completely under the control of T3 (Fig. 2a) as a negligible amount of hepatic enzyme activity was observed in neonates from PTU-fed mothers during the period of study. The relationship between thyroid status and hepatic ME activity has been extensively studied by several investigators.“6*1” Oppenheimer et ~1.” and Hemon’ however, have failed to show ME response in the neonate brains on supplementation with thyroid hormones.
208
U. R. Thakare
ef al.
Although no clearly defined function for malic enzyme has been established in the brain, this enzyme plays an important role in lipogenesis through the production of NADPH required for fatty acid synthesis and hence, for myelination process. Increase in hepatic ME activity at the time of weaning (day 21) has been related to its role in lipid biosynthesis.” Decreased activity of this enzyme in the brain during the developmental period might therefore affect lipogenesis which may lead to impairment of myelination in the brain and hence, dysfunction. Studies are in progress to correlate ME activity and myelination in the neonate brain during development. REFERENCES 1. Back D. W., Wilson S. B., Norris S. M. and Goodridge A. G. (1986) Hormonal regulation of lipogenic enzymes in chick embryo hepatocytes in culture. /. biof. Chem. 261, 12555-12561. 2. Balazs R., Brooksbank B. W. L., Davison A. N., Eayrs J. T. and Wilson D. A. (1%9) The effect of neonatal thyroidectomy on myelination in the rat brain. Brain Res. 15, 219-232. 3. Bass N. H., Pelton E. W. and Young E. (1977) Thyroid Hormones and Bruin Development (ed. Grave G. D.), pp. 199-214. Raven Press, New York. 4. Diez Guerra J., Aragon M. C., Gimenez C. and Valdivieso F. (1981) Effect of thyroid hormones on the Malic enzyme activity in rat brain during development. Devl Neurosci. 4, 130-133. in the 5. Hemon P. (1968) Malate dehydrogenase (decarboxylating) (NADPH) and glycerophosphateoxidase developing rat. Biochim. Biophys. Acta 151, 681-683. 6. Hsu R. Y. and Lardy H. A. (1969) Malic enzyme. Merh. Enzym. 13, 230-235. 7. Liu L. and Nicoll C. S. (1986) Postnatal development of dependence on thyroid hormones for growth and differentiation of rat skeletal structures. Growth 50, 472484. 8. Lobato M. F., Francisco M. R., Moreno J. and Garcia-Ruiz J. P. (1986) Nutritional and hormonal regulation of malic enzyme synthesis in rat mammary gland. Biochem. J. 236, 4414l5. 9. Lowry 0. H.. Rosebrough N. J., Farr A. L. and Randall R. A. (1951) Protein measurement with the Folin phenol reagent. 1. biol. Chem. 193.263-275. J. M.. Silva E.. Schwartz H. L. and Surks M. I. (1977) Stimulation of hepatic 10. Oooenheimer &&hondrial-glycerophospate dehydrogenase and Malic enzyme by L-triiodothyronine. J. Clin Invest. 59, 517-527. 11. Schwartz H. L. (1983) Molecular Basis of Thyroid Hormone Action (eds Oppenheimer J. H. and Samuels H. H.), p. 414. Academic Press, New York. 12. Schwartz H. L. and Oppenheimer J. H. (1978) Ontogenesis of 3.5,3’-triiodothyronine receptors in neonatal rat brain. Endocrinology
103,943-948.
13. Shanker G., Amur S. G. and Pieringer R. A. (1985) Investigations on myelinogenesis in vitro. Neurochem. Res. 10, 617-625. 14. Thakare U. R., Vijayan U., Sushila L. and Shah D. H. (1987) Malic enzyme response to triiodothyronine in the brains of neonatal rats. Life Sci. 41, 2823-2826. 15. Vernon R. G. and Walker D. G. (1986) Changes in activity of some enzymes involved in glucose utilization and formation in developing rat liver. Bioche’m. J. lii6, 321-329. _ 16 Walravens P. and Chase H. P. (1969) Influence of thyroid hormone on formation of myelin lipid. J. Neurochem. 16, 1477-1484. 17. Wysocki S. J. and Segal W. (1972) Influence of thyroid hormones on enzyme activities of myelinating rat central nervous tissues. Eur. J. Biochem. 28, 183-189.