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Increased cytochrome oxidase activity of mesencephalic neurons in developing rats displaying methylmercury-induced movement and postural disorders
Neuroseience Letters, 89 (1988) 271 276
27 I
Elsevier Scientific Publishers Ireland Ltd.
NSL 05407
Increased cytochrome oxidase activity of mesencephalic neurons in developing rats displaying methylmercury-induced movement and postural disorders Richard H. Dyck 2 and John R. O'Kusky I IDepartment ~/" Pathology, Division o["Medical Microbiology and:the Kinsmen Laboratory O! Neurological Research, University qf British Columbia, Vancouver, B.C. (Canada) (Received 30 December 1987; Revised version received 16 March 1988; Accepted 18 March 1988)
Key words." Methylmercury; Neurotoxicit~; Development; Red nucleus; Cytochrome oxidase: Cerebral palsy Subcutaneous administration of the neurotoxin methylmercuric chloride to developing rats produced movement and postural disorders during the 4th postnatal week. Cytochrome oxidase histochemistry revealed an increase in the oxidative metabolic activity of small neurons within the magnocellular red nucleus (RMC) and the interrubral mesencephalon. A concurrent suppression of cytochrome oxidase activity in the large neurons and neuropil of RMC was apparent relative to controls. Decortication on postnatal day 3 did not alter the course of motor impairment or the cytochrome oxidase histopathology, suggesting that the role of neocortex in the pathogenesis of methylmercury-induced movement and postural disorders is minimal.
The neurotoxicity of methylmercury in humans during prenatal and early postnatal development can result in neurological disorders resembling cerebral palsy [1, 2]. Chronic postnatal administration of methylmercury to developing rats has been shown to produce a similar syndrome, including spasticity, ataxia and myoclonic jerking, which appears during the animal's fourth postnatal week [7 10]. Using this animal model, ultrastructural examination of the brain at the onset of neurological impairment has revealed a relatively selective degeneration of aspinous and sparsely-spinous stellate neurons in the cerebral cortex [7]. In the neocortex and caudate-putamen, neurochemical assays have shown a significant reduction in the specific activity of glutamate decarboxylase [9, 10], the enzyme which synthesizes 7aminobutyric acid (GABA). These studies indicate that inhibitory GABAergic interneurons in neocortex are highly susceptible to methylmercury toxicity. A consequent
Correspondence: R.H. Dyck, Kinsmen Laboratory of Neurological Research, 2255 Wesbrook Mall, Vancouver, B.C. Canada, V6T IW5. 0304-3940/88/$ 03.50 O 1988 Elsevier Scientific Publishers Ireland Ltd.
272 disinhibition of cortical efferent neurons may contribute to the observed motor impairment by increasing activity along corticospinal or cortico-bulbospinal pathways. Previous studies have demonstrated that changes in activity within discrete neuronal populations can be detected using an histochemical technique for the presence of the mitochondrial enzyme cytochrome oxidase (CO) [3, 14-16]. This hi stochemical method was used in the present study to investigate abnormal neuronal activity in brainstem nuclei of methylmercury-treated rats at the onset of neurological impairment. If increased activity of brainstem neurons is caused primarily by an increase in corticofugal activity, then neonatal decortication should reduce or eliminate methylmercury-induced changes. This hypothesis was tested by comparing CO histochemistry following methylmercury treatment of both normal and decorticate neonatal rats. Each of 6 litters of Sprague-Dawley rats were culled to 3 triplets of weight- and gender-matched pups on postnatal day (PND) 3. Individual pups within each triplet were randomly assigned as methylmercury-treated (MeHg), weight-matched control (WMC), or normal control (NC). The matched triplets in 3 litters received complete aspirative neocortical lesions on PND 3. At the time of surgery, pups were placed in a cotton-lined box and hypothermic anaesthesia was induced by cooling for 4-8 min at - 10°C. The utility and effectiveness of using hypothermia as an anaesthetic is reviewed in Popovic and Popovic [12]. Animals were monitored until a toe-pinch no longer elicited limb withdrawal or a crossed extensor reflex. Using this technique animals were completely immobilized for 6-8 min, while the surgical procedure required less than 4 min. Decortication was performed using the method of Kolb and Whishaw [5]. The frontal and parietal bones were removed with iris scissors leaving only a narrow strip of bone in the midline. The dura overlying the exposed cortex was excised and the neocortex was aspirated using a glass pipette under visual guidance with the aid of a surgical microscope. Frontal and posterior cortex was removed from the midline laterally to the rhinal fissure. The wound was sutured and the rats were allowed to warm under a heat lamp before being returned to the home cage. Beginning on PND 5, MeHg animals received daily subcutaneous injections of 0.01 M methylmercuric chloride in physiological saline (5 mg Hg/kg). Control animals received injections of an equivalent volume of physiological saline. WMC animals were periodically isolated in an incubator at 37°C to maintain body weight within _+5% of MeHg rats. At the onset o f neurological impairment (PND 20-26), an individual MeHg rat and its matched controls were injected i.p. with a lethal dose of sodium pentobarbital and perfused through the ascending aorta with ice-cold fixative containing 4% paraformaldehyde and 4% sucrose in 0.1 M phosphate buffer (pH 7.4) at a pressure of 120 mmHg for 15 rain. The brains were removed immediately and postfixed for 45 min, at which time they were transferred to 0.1 M phosphate buffer containing 4% sucrose (PBS) for 30 min. The 3 brains from each triplet were blocked and mounted on a brass freezing platform and quickly frozen with powdered dry ice. Coronal sections (40 pm) from all 3 brains were cut simultaneously through the brainstem and collected in cold PBS. Sections were processed for the presence of cytochrome oxi-
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Postnatal Day Fig. 1. Postnatal changes in body weight for methylmercury-treated (MeHg) and normal control (NC)
animals, including those animals receivingaspirative neocortical lesions (Decort.) on PND 3. Inset: abnormal weight gain in decorticate and cortex-intact animals, expressed as a percentage of corresponding norreal controls.
dase using the histochemical method of Wong-Riley [14]. Sections were incubated at 3T~C for 2 h in a PBS solution containing 0.03% cytochrome c (Sigma C-2506) and 0.05% 3,3'-diaminobenzidine (Sigma D-5637), washed in 3 changes of PBS for 15 rain each and then mounted on chrome alum-coated slides. The mounted sections were dehydrated briefly in absolute ethanol, followed by 5 rain in xylene and then coverslipped with Permount. Representative sections were stained for Nissl substance using 0.1% thionin in acetate buffer (pH 3.7). The first detectable clinical sign of methylmercury toxicity was an impaired rate of weight gain in M e H g relative to NC animals beginning around P N D 15 (Fig. I). Neonatal decortication alone resulted in an impaired rate of weight gain (Fig. 1); however, methy[mercury administration to decorticate or cortex-intact rats resulted in an identical impairment of weight gain relative to their normal controls (Fig. I, inset). M e H g rats exhibited a mixed spastic/dyskinetic syndrome which appeared between P N D 20 26. Clinical signs of neurological impairment were the same as those reported in previous studies [7 10], including hypertonicity of limb muscles, flexion deformities, myoclonic jerking of hindlimbs and generalized m o t o r convulsions. The same signs were observed in decorticate MeHg animals, with no differences in either age of onset or severity. These gross m o t o r abnormalities were not observed in NC decorticate animals. A distinctly different set of subtle abnormalities in postural reflexes and fine m o t o r control following neonatal decortication has been demonstrated in previous studies [5].
274
Histological examination of sections stained for CO revealed a striking difference between MeHg and control animals in the mesencephalon at the level of the magnocellular red nucleus (RMC). In NC and W M C animals, cell bodies of the R M C neurons were darkly stained and surrounded by neuropil of moderate staining intensity (Fig. 2A). In all MeHg animals (n = 18) a population of small, very intensely COpositive neurons was observed within RMC, extending into the interrubral mesencephalon to the midline (Fig. 2B). These neurons were not observed in any NC or W M C animals (n = 36). There was a moderate decrease in the staining intensity of the neuropil and fewer of the larger CO-positive cell bodies were observed. Nissl sections from the same region revealed no degenerative changes in R M C neurons (e.g.
Fig. 2. Cytochrome oxidase histochemistry in the mesencephalon at the level of the magnocellular red nucleus in (A) WMC, (B) MeHg and (C) decorticate MeHg animals. The midline of the mesencephalon is oriented along the right-hand side of each figure. Neurons indicated by the arrow in C are shown in D at higher magnification. Calibration bars in itm. Magnification is identical in A, B and C.
275
pyknotic nuclei or gliosis). In addition, the size and number of RMC neurons appeared to be equivalent in MeHg and control animals. The neurotoxicity of methylmercury in decorticate animals resulted in identical changes in CO histochemistry of the RMC and the interrubral mesencephalon (Fig. 2C). At higher magnification, the very intensely stained CO-positive cells were confirmed to be neurons by virtue of stained dendritic processes on many profiles (Fig. 2D, arrow). Cytochrome oxidase histochemistry demonstrated changes in the oxidative metabolic activity of neurons in the RMC and in the interrubral mesencephalon. In the RMC of MeHg animals there appeared to be a decrease in CO levels in large neurons with a concomitant increase in smaller neurons. The presence of a normal complement of RMC neurons in MeHg animals, as seen on Nissl sections, suggests that the changes in CO histochemistry were not simply the result of atrophy or degeneration. Since the RMC has been shown to project to the spinal cord in a number of animals including the rat [4, 6, 11, 13], these alterations in oxidative metabolic activity likely contribute to methylmercury-induced movement and postural disorders. The very intense CO-positive neurons in the interrubral mesencephalon are particularly interesting in that they do not correspond to any previously defined functional group. Their possible functional relationship to the RMC and role in the actiology of methylmercury-induced developmental disorders has yet to be determined. The neurotransmitter specificity of this anomalous neuronal population and the targets of its axonal projections are currently under investigation. Perhaps the most interesting finding in the present study is the fact that neonatal decortication alters neither the cytochrome oxidase histopathology nor the course of motor impairment during development. Thus, it is more likely that abnormal CO levels in rnesencephalic neurons result from a direct neurotoxic effect of methylmercury or from a disinhibition of other afferents to these neurons. This project was supported by a grant from the United Cerebral Palsy Research and Educational Foundation, Inc. J.R.O. is a Research Scholar of the Medical Research Council of Canada. R.H.D. is on leave of absence from The University of Lethbridge, Lethbridge, Alta., Canada. 1 Amin-Zaki, L., Majeed, M.A., Elhassani, S.B., Clarkson, T.W., Greenwood, M.R. and Doherty. R.A., Prenatal methylmercury poisoning, Am. J. Dis. Child., 133 (1979) 172 177. 2 Chang, L.W. and Annau, Z., Developmental neuropathology and behavioral teratology ofmcthylmercury. In J. Yanai (Ed.), Neurobehavioral Teratology, Elsevier, Amsterdam, 1984, pp. 405 432. 3 Darriet, D., Der, T. and Collins, R.C,, Distribution of cytochrome oxidase in rat brain: studies with diaminobenzidine histochemistry in vitro and [~4C]cyanide tissue labeling in vivo, J. Ccreb. Blood I~'low Metab., 6 (1986) 8 14. 4 Gwyn, D.G. and Flumerfelt, B.A., A comparison of the distribution of cortical and cerebellar afl'erents in the red nucleus of the rat, Brain Res., 69 (1974) 130 135. 5 Kolb, B.E. and Whishaw, I.Q., Decortication in rats in infancy or adulthood produced comparable functional losses on learned and species-typical behaviors, J. Comp. Physiol. Psychol., 95 (1981) 468 483. 6 Kuypers, H.G.J.M. and Lawrence, D.G., Cortical projections to the red nucleus and the brain stem in the rhesus monkey, Brain Res., 4 (1967) 151 188.
276 7 0 ' K u s k y , J.R., Synaptic degeneration in rat visual cortex after neonatal administration of methylmercury, Exp. Neurol., 89 (1985) 32~47. 8 O'Kusky, J.R., Boyes, B.E. and McGeer, E.G., Methylmercury-induced movement and postural disorders in developing rat: regional analysis of brain catecholamines and indoleamines, Brain Res., 439 (1988) 138 146. 9 O'Kusky, J.R. and McGeer, E.G., Methylmercury poisoning of the developing nervous system in the rat: decreased activity of glutamic acid decarboxylase in cerebral cortex and neostriatum, Dev. Brain Res., 21 (1985) 299--306. I0 O'Kusky, JR., Radke, J.M. and Vincent, S.R., Methylmercury-induced movement and postural disorders in developing rat: loss of somatostatin-immunoreactive interneurons in the striatum, Dev. Brain Res., in press. I 1 Pompeiano, O. and Brodal, A., Experimental demonstration of a somatotopical origin of rubrospinal fibres in the cat, J. Comp. Neurol., 108 (1957) 225 251. 12 Popovic, V. and Popovic, P., Hypothermia in Biology and in Medicine, Grune & Stratton, New York, 1974, 305 pp. 13 Reid, J.M., Gwyn, D.G. and Flumerfelt, B.A., A cytoarchitectonic and golgi study of the red nucleus in the rat, J. Comp. Neurol., 162 (1975) 337 349. 14 Wong-Riley, M.T.T., Changes in the visual system of monocularly sutured or enucleated cats demonstrable with cytochrome oxidase histochemistry, Brain Res., 171 (1979) I 1 28. 15 Wong-Riley, M. and Carroll, E.W., Effect of impulse blockade on cytochrome oxidase activity in monkey visual system, Nature (Lond.), 307 (1984) 262 264. 16 Wong-Riley, M.T.T. and Welt, C., Histochemical changes in cytochrome oxidase of cortical barrels after vibrissal removal in neonatal and adult mice, Proc. Natl. Acad, Sci. U.S.A., 77 (1980) 2333-2337.
27 I
Elsevier Scientific Publishers Ireland Ltd.
NSL 05407
Increased cytochrome oxidase activity of mesencephalic neurons in developing rats displaying methylmercury-induced movement and postural disorders Richard H. Dyck 2 and John R. O'Kusky I IDepartment ~/" Pathology, Division o["Medical Microbiology and:the Kinsmen Laboratory O! Neurological Research, University qf British Columbia, Vancouver, B.C. (Canada) (Received 30 December 1987; Revised version received 16 March 1988; Accepted 18 March 1988)
Key words." Methylmercury; Neurotoxicit~; Development; Red nucleus; Cytochrome oxidase: Cerebral palsy Subcutaneous administration of the neurotoxin methylmercuric chloride to developing rats produced movement and postural disorders during the 4th postnatal week. Cytochrome oxidase histochemistry revealed an increase in the oxidative metabolic activity of small neurons within the magnocellular red nucleus (RMC) and the interrubral mesencephalon. A concurrent suppression of cytochrome oxidase activity in the large neurons and neuropil of RMC was apparent relative to controls. Decortication on postnatal day 3 did not alter the course of motor impairment or the cytochrome oxidase histopathology, suggesting that the role of neocortex in the pathogenesis of methylmercury-induced movement and postural disorders is minimal.
The neurotoxicity of methylmercury in humans during prenatal and early postnatal development can result in neurological disorders resembling cerebral palsy [1, 2]. Chronic postnatal administration of methylmercury to developing rats has been shown to produce a similar syndrome, including spasticity, ataxia and myoclonic jerking, which appears during the animal's fourth postnatal week [7 10]. Using this animal model, ultrastructural examination of the brain at the onset of neurological impairment has revealed a relatively selective degeneration of aspinous and sparsely-spinous stellate neurons in the cerebral cortex [7]. In the neocortex and caudate-putamen, neurochemical assays have shown a significant reduction in the specific activity of glutamate decarboxylase [9, 10], the enzyme which synthesizes 7aminobutyric acid (GABA). These studies indicate that inhibitory GABAergic interneurons in neocortex are highly susceptible to methylmercury toxicity. A consequent
Correspondence: R.H. Dyck, Kinsmen Laboratory of Neurological Research, 2255 Wesbrook Mall, Vancouver, B.C. Canada, V6T IW5. 0304-3940/88/$ 03.50 O 1988 Elsevier Scientific Publishers Ireland Ltd.
272 disinhibition of cortical efferent neurons may contribute to the observed motor impairment by increasing activity along corticospinal or cortico-bulbospinal pathways. Previous studies have demonstrated that changes in activity within discrete neuronal populations can be detected using an histochemical technique for the presence of the mitochondrial enzyme cytochrome oxidase (CO) [3, 14-16]. This hi stochemical method was used in the present study to investigate abnormal neuronal activity in brainstem nuclei of methylmercury-treated rats at the onset of neurological impairment. If increased activity of brainstem neurons is caused primarily by an increase in corticofugal activity, then neonatal decortication should reduce or eliminate methylmercury-induced changes. This hypothesis was tested by comparing CO histochemistry following methylmercury treatment of both normal and decorticate neonatal rats. Each of 6 litters of Sprague-Dawley rats were culled to 3 triplets of weight- and gender-matched pups on postnatal day (PND) 3. Individual pups within each triplet were randomly assigned as methylmercury-treated (MeHg), weight-matched control (WMC), or normal control (NC). The matched triplets in 3 litters received complete aspirative neocortical lesions on PND 3. At the time of surgery, pups were placed in a cotton-lined box and hypothermic anaesthesia was induced by cooling for 4-8 min at - 10°C. The utility and effectiveness of using hypothermia as an anaesthetic is reviewed in Popovic and Popovic [12]. Animals were monitored until a toe-pinch no longer elicited limb withdrawal or a crossed extensor reflex. Using this technique animals were completely immobilized for 6-8 min, while the surgical procedure required less than 4 min. Decortication was performed using the method of Kolb and Whishaw [5]. The frontal and parietal bones were removed with iris scissors leaving only a narrow strip of bone in the midline. The dura overlying the exposed cortex was excised and the neocortex was aspirated using a glass pipette under visual guidance with the aid of a surgical microscope. Frontal and posterior cortex was removed from the midline laterally to the rhinal fissure. The wound was sutured and the rats were allowed to warm under a heat lamp before being returned to the home cage. Beginning on PND 5, MeHg animals received daily subcutaneous injections of 0.01 M methylmercuric chloride in physiological saline (5 mg Hg/kg). Control animals received injections of an equivalent volume of physiological saline. WMC animals were periodically isolated in an incubator at 37°C to maintain body weight within _+5% of MeHg rats. At the onset o f neurological impairment (PND 20-26), an individual MeHg rat and its matched controls were injected i.p. with a lethal dose of sodium pentobarbital and perfused through the ascending aorta with ice-cold fixative containing 4% paraformaldehyde and 4% sucrose in 0.1 M phosphate buffer (pH 7.4) at a pressure of 120 mmHg for 15 rain. The brains were removed immediately and postfixed for 45 min, at which time they were transferred to 0.1 M phosphate buffer containing 4% sucrose (PBS) for 30 min. The 3 brains from each triplet were blocked and mounted on a brass freezing platform and quickly frozen with powdered dry ice. Coronal sections (40 pm) from all 3 brains were cut simultaneously through the brainstem and collected in cold PBS. Sections were processed for the presence of cytochrome oxi-
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Postnatal Day Fig. 1. Postnatal changes in body weight for methylmercury-treated (MeHg) and normal control (NC)
animals, including those animals receivingaspirative neocortical lesions (Decort.) on PND 3. Inset: abnormal weight gain in decorticate and cortex-intact animals, expressed as a percentage of corresponding norreal controls.
dase using the histochemical method of Wong-Riley [14]. Sections were incubated at 3T~C for 2 h in a PBS solution containing 0.03% cytochrome c (Sigma C-2506) and 0.05% 3,3'-diaminobenzidine (Sigma D-5637), washed in 3 changes of PBS for 15 rain each and then mounted on chrome alum-coated slides. The mounted sections were dehydrated briefly in absolute ethanol, followed by 5 rain in xylene and then coverslipped with Permount. Representative sections were stained for Nissl substance using 0.1% thionin in acetate buffer (pH 3.7). The first detectable clinical sign of methylmercury toxicity was an impaired rate of weight gain in M e H g relative to NC animals beginning around P N D 15 (Fig. I). Neonatal decortication alone resulted in an impaired rate of weight gain (Fig. 1); however, methy[mercury administration to decorticate or cortex-intact rats resulted in an identical impairment of weight gain relative to their normal controls (Fig. I, inset). M e H g rats exhibited a mixed spastic/dyskinetic syndrome which appeared between P N D 20 26. Clinical signs of neurological impairment were the same as those reported in previous studies [7 10], including hypertonicity of limb muscles, flexion deformities, myoclonic jerking of hindlimbs and generalized m o t o r convulsions. The same signs were observed in decorticate MeHg animals, with no differences in either age of onset or severity. These gross m o t o r abnormalities were not observed in NC decorticate animals. A distinctly different set of subtle abnormalities in postural reflexes and fine m o t o r control following neonatal decortication has been demonstrated in previous studies [5].
274
Histological examination of sections stained for CO revealed a striking difference between MeHg and control animals in the mesencephalon at the level of the magnocellular red nucleus (RMC). In NC and W M C animals, cell bodies of the R M C neurons were darkly stained and surrounded by neuropil of moderate staining intensity (Fig. 2A). In all MeHg animals (n = 18) a population of small, very intensely COpositive neurons was observed within RMC, extending into the interrubral mesencephalon to the midline (Fig. 2B). These neurons were not observed in any NC or W M C animals (n = 36). There was a moderate decrease in the staining intensity of the neuropil and fewer of the larger CO-positive cell bodies were observed. Nissl sections from the same region revealed no degenerative changes in R M C neurons (e.g.
Fig. 2. Cytochrome oxidase histochemistry in the mesencephalon at the level of the magnocellular red nucleus in (A) WMC, (B) MeHg and (C) decorticate MeHg animals. The midline of the mesencephalon is oriented along the right-hand side of each figure. Neurons indicated by the arrow in C are shown in D at higher magnification. Calibration bars in itm. Magnification is identical in A, B and C.
275
pyknotic nuclei or gliosis). In addition, the size and number of RMC neurons appeared to be equivalent in MeHg and control animals. The neurotoxicity of methylmercury in decorticate animals resulted in identical changes in CO histochemistry of the RMC and the interrubral mesencephalon (Fig. 2C). At higher magnification, the very intensely stained CO-positive cells were confirmed to be neurons by virtue of stained dendritic processes on many profiles (Fig. 2D, arrow). Cytochrome oxidase histochemistry demonstrated changes in the oxidative metabolic activity of neurons in the RMC and in the interrubral mesencephalon. In the RMC of MeHg animals there appeared to be a decrease in CO levels in large neurons with a concomitant increase in smaller neurons. The presence of a normal complement of RMC neurons in MeHg animals, as seen on Nissl sections, suggests that the changes in CO histochemistry were not simply the result of atrophy or degeneration. Since the RMC has been shown to project to the spinal cord in a number of animals including the rat [4, 6, 11, 13], these alterations in oxidative metabolic activity likely contribute to methylmercury-induced movement and postural disorders. The very intense CO-positive neurons in the interrubral mesencephalon are particularly interesting in that they do not correspond to any previously defined functional group. Their possible functional relationship to the RMC and role in the actiology of methylmercury-induced developmental disorders has yet to be determined. The neurotransmitter specificity of this anomalous neuronal population and the targets of its axonal projections are currently under investigation. Perhaps the most interesting finding in the present study is the fact that neonatal decortication alters neither the cytochrome oxidase histopathology nor the course of motor impairment during development. Thus, it is more likely that abnormal CO levels in rnesencephalic neurons result from a direct neurotoxic effect of methylmercury or from a disinhibition of other afferents to these neurons. This project was supported by a grant from the United Cerebral Palsy Research and Educational Foundation, Inc. J.R.O. is a Research Scholar of the Medical Research Council of Canada. R.H.D. is on leave of absence from The University of Lethbridge, Lethbridge, Alta., Canada. 1 Amin-Zaki, L., Majeed, M.A., Elhassani, S.B., Clarkson, T.W., Greenwood, M.R. and Doherty. R.A., Prenatal methylmercury poisoning, Am. J. Dis. Child., 133 (1979) 172 177. 2 Chang, L.W. and Annau, Z., Developmental neuropathology and behavioral teratology ofmcthylmercury. In J. Yanai (Ed.), Neurobehavioral Teratology, Elsevier, Amsterdam, 1984, pp. 405 432. 3 Darriet, D., Der, T. and Collins, R.C,, Distribution of cytochrome oxidase in rat brain: studies with diaminobenzidine histochemistry in vitro and [~4C]cyanide tissue labeling in vivo, J. Ccreb. Blood I~'low Metab., 6 (1986) 8 14. 4 Gwyn, D.G. and Flumerfelt, B.A., A comparison of the distribution of cortical and cerebellar afl'erents in the red nucleus of the rat, Brain Res., 69 (1974) 130 135. 5 Kolb, B.E. and Whishaw, I.Q., Decortication in rats in infancy or adulthood produced comparable functional losses on learned and species-typical behaviors, J. Comp. Physiol. Psychol., 95 (1981) 468 483. 6 Kuypers, H.G.J.M. and Lawrence, D.G., Cortical projections to the red nucleus and the brain stem in the rhesus monkey, Brain Res., 4 (1967) 151 188.
276 7 0 ' K u s k y , J.R., Synaptic degeneration in rat visual cortex after neonatal administration of methylmercury, Exp. Neurol., 89 (1985) 32~47. 8 O'Kusky, J.R., Boyes, B.E. and McGeer, E.G., Methylmercury-induced movement and postural disorders in developing rat: regional analysis of brain catecholamines and indoleamines, Brain Res., 439 (1988) 138 146. 9 O'Kusky, J.R. and McGeer, E.G., Methylmercury poisoning of the developing nervous system in the rat: decreased activity of glutamic acid decarboxylase in cerebral cortex and neostriatum, Dev. Brain Res., 21 (1985) 299--306. I0 O'Kusky, JR., Radke, J.M. and Vincent, S.R., Methylmercury-induced movement and postural disorders in developing rat: loss of somatostatin-immunoreactive interneurons in the striatum, Dev. Brain Res., in press. I 1 Pompeiano, O. and Brodal, A., Experimental demonstration of a somatotopical origin of rubrospinal fibres in the cat, J. Comp. Neurol., 108 (1957) 225 251. 12 Popovic, V. and Popovic, P., Hypothermia in Biology and in Medicine, Grune & Stratton, New York, 1974, 305 pp. 13 Reid, J.M., Gwyn, D.G. and Flumerfelt, B.A., A cytoarchitectonic and golgi study of the red nucleus in the rat, J. Comp. Neurol., 162 (1975) 337 349. 14 Wong-Riley, M.T.T., Changes in the visual system of monocularly sutured or enucleated cats demonstrable with cytochrome oxidase histochemistry, Brain Res., 171 (1979) I 1 28. 15 Wong-Riley, M. and Carroll, E.W., Effect of impulse blockade on cytochrome oxidase activity in monkey visual system, Nature (Lond.), 307 (1984) 262 264. 16 Wong-Riley, M.T.T. and Welt, C., Histochemical changes in cytochrome oxidase of cortical barrels after vibrissal removal in neonatal and adult mice, Proc. Natl. Acad, Sci. U.S.A., 77 (1980) 2333-2337.