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Evidence for a GABA-containing projection from the entopeduncular nucleus to the lateral habenula in the rat
360
Brain Research, 145 I1978i 360-364 , :: Elsevier/North-Holland Biomedical Press
Evidence for a GABA.containing projection from the entopeduncular nucleus to the lateral habenula in the rat
J. I. NAGY, D. A. CARTER, J. LEHMANN and H. C. F1BIGER Division of Neurological Sciences, Department of Psychiatry, University of British Columbia, VancouveG B.C. V6T 1 W5 (Canada)
(Accepted November 3rd, 1977)
Major efferent projections of the striatum and globus pallidus (GP) are known to terminate in the substantia nigra2,S-1Z,lv,19.20,24,z.L the nucleus tegmenti pedunculo pontis (TPP), the subthalamic nucleus and certain thalamic nuclei 2,s,11,1%~. Recently, the internal division of the GP (GPi) in the primate has been shown to project to the lateral habenula is. In addition, the entopeduncular nucleus (EP), the homologue of GPi in subprimate species, has also been shown to project to the lateral habenula in the cat 21. There is evidence that some of the efferent projections of the striatum and G P contain gamma-aminobutyric acid (GABA) and its synthesizing enzyme, glutamic acid decarboxylase (GAD) 6'12"16"zo,2z. In a recent study, Gottesfeld et al. 7 observed that electrolytic lesions of the stria medullaris (SM), GP or GP plus EP significantly decreased G A D activity in the lateral habenula. The authors suggested that certain GABAcontaining neurons in the GP project to the lateral habenula, with a further contribution from the EP. The present investigation was designed to further elucidate the origin of GAD-containing afferents to the lateral habenula. Adult male Wistar rats (300-320 g) received stereotaxic injections of [aH]leucine (0.1-0.15 #1 of 6.5 #Ci/#l, sp.act. 50 Ci/mmole) into the EP (n == 5) or G P (n - 4) via a Hamilton microsyringe. The animals were killed 24 h later and 40/~m frontal sections of the brain were prepared for autoradiography as previously described ~. Another group of rats (n =~ 5) received unilateral stereotaxic electrolytic lesions (2 mA for 30 sec) of the EP through an electrode insulated except at the tip. These animals were killed by cervical fracture 10-14 days postoperatively and the brain was placed on a freezing microtome. Thick (300 ,urn) sections were taken through the habenula and from these sections the left and right habenula was dissected out with the aid of a dissecting microscope. The tissues were homogenized in 100 #1 of 50 mM Tris • acetate buffer p H 6.2, containing Triton X-100 (0.2 ~o v/v). G A D and choline acetyltransferase (CAT) activities were measured in homogenates as previously describedl,a,~,l"% Although injections of [ZH]leucine into the GP resulted in heavy accumulations of autoradiographic grains over the subthalamic nucleus and the substantia nigra 4, no labeling was observed in the habenula of any of the injected animals. In contrast, injections of [aH]leucine into EP resulted in distinct labeling of the lateral habenuta:
361
2,'
'r°~
.
.
.
.
.
:i,~
"
if
B
Fig. I. A: photomicrograph showing injection site of [3H]leucine into the entopeduncular nucleus, ,: 26. B: dark-field photomicrograph showing autoradiographic label in the lateral hahenula 24 h after an injection of [3H]leucine into the entopeduncular nucleus. Note heavy accumulation of grains in the ventrolateral aspect of the nucleus y, 125. C: photomicrograph of electrolytic lesion of the entopeduncular nucleus x 52.
362 TABLE 1 The activity of G A D and CA T in the habemda on the lesioned amt control side after unih:tcra/ electrolytic lesions of'the entopeduncular nucleus' Side
n
GAD
nt~t/es/tltg protein hour
Lesion Control
(~ (~
C,4 ]"
Per cent control
13.6 2.40": 63.6 21.4 ~ 2.34 100.0
* Significantly different from control P
mnoh, s/mg proteh~ hour
Per cent control
122 124
98.4 100.0
8. t 10.8
0.05~
Fig. 1A shows the dense concentration of autoradiographic grains over the [3H ]teucine injection site in the EP. In this. as in other animals, there was a small amount of diffusion either dorsally, along the needle tract, or ventromedially into the lateral hypothalamus. Fig. 1B is a dark field photomicrograph showing the heavy labeling of the ventrolaterat aspect of the lateral habenula in the same animal. A histology of a representative electrol3 tic lesion of the EP is gwen in Fig. 1C. It is apparent that the lesion destroyed most of the EP at this level and extended beyond its limits both dorsally and ventrally. The lesion did not encroach upon the GP. The effects of similar lesions on habenular CAT and G A D activities are g~ven in Table I. EP lesions produced a significant decrease in G A D to 64 0,i of the control value without significantly affecting CAT. The present results have demonstrated the existence of an entoped uncular-lateral habenula projection in the rat. Injections of [~H]leucine produced clearly defined autoradiographic labeling in the ventrotateral aspect of the lateral habenula. Projections of the EP were also identified in the ventrolateral aspect of the ventrolateral -ventroanterior thalamic complex, the parafascicular nucleus and the nucleus tegmenti pedunculopontis. These observations have been described in detail elsewhereL Furthermore. that the autoradiographic labeling in the lateral habenula was due to a projection from EP rather than a result of diffusion of [aH]leucine to other nuclei in the vicinity of the EP has been confirmed using horseradish peroxidase histochemistry ~,t:~. The fact that electrolytic lesions of the EP significantly reduced G A D activity in the h abenula suggests that this is a GABA-containing pathway. Moreover, the failure of the lesions to have an effect on CAT in the habenula points to the specificity of this effect. Gottesfeld et al. 7 reported that electrolytic lesions of GP resulted in a 22'~,; decrease in lateral habenular G A D activity, lesions of GP plus EP produced a 32",(~ decrease, and lesions m stria terminalis a 56% decrease. The present autoradiographic results suggest that the reduced G A D activity observed by Gottesfeld et al. 7 after GP lesions was the result of damage to fibers originating in EP and ascending in the vicinity of the GP. Support for this suggestmn is found in the fact that the EP lesions utilized in the present study produced a similar decrease (34 o/;) in habenular G A D activity as was found after the combined GP-EP lesions used by Gottesfeld et al. 7. ALternatively, the decrease found by Gottesfeld et al. 7 after GP lesions alone may have been due to a ventral encroachment of the lesion upon the substantia innominata ~a- The failure of
363 EP lesions to reduce habenular G A D by more than 36 o,O/ m a y have been due to incomplete destruction of the EP. Alternatively, other proposed afferents to the lateral or medial habenula such as the lateral hypothalamus, nucleus of the diagonal band, substantia innominata, lateral preoptic area, mesencephalic raphe, supra-commissural septum and ventral tegmental area 13 may also contain G A D and explain the higher loss of G A D after stria terminalis lesions 7. The remaining part of G A D in the habenula could be due to local neurons. Furthermore, inasmuch as the entopeduncular projection terminates mainly in the ventrolateral aspect of the lateral habenula (Fig. I B), it is possible that if only this part of the nucleus had been assayed, then a greater decrease in G A D may have been obtained. The main proportion of G A D in habenula is localized to its lateral part ~. Nauta 2a has commented on the potential importance of a projection from the EP to the lateral habenula inasnmch as it may represent a point of communication between the extrapyramidal and limbic systems. In this regard, it is significant that the lateral habenula is known to project to and receive afferents from both of these systems. For example with respect to afferents, the lateral habenula receives a major projection from the entopeduncular nucleus as well as from the septum and preoptic area 13,'~a. The efferent extrapyramidal-related projections of the lateral habenula include the substantia nigra pars compacta, and the ventromedial nucleus of the thalamus2, t4. The efferent limbic-related projections include preoptic area, the lateral hypothalamus and the central gray 14,'~2. The extent to which these two streams are integrated or remain segregated in the lateral habenula awaits further study. In any event, it is suggested that the EP to lateral habenular pathway is an inhibitory link in the extrapyramidal circuit, utilizing the neurotransmitter GABA, by which entopeduncular output may modulate not only nuclei of the extrapyramidal system, but also parts of the limbic system. Wang and Aghajanian ~ have recently shown that the projection from the lateral habenula to the dorsal raphe nucleus is inhibitory. This, together with the present observations, raises the possibility that the extrapyramidal system may modulate the activity of serotonergic neurons in midbrain raphe nuclei via the EP-lateral habenula projection. If other lateral habenular efferents prove to be inhibitory, then the presumed inhibitory GABA pathway from EP to lateral habenula may physiologically disinhibit components of both extrapyramidal and limbic function.
I Albers, R. W. and Brady, R. O., The distribution of glutamate decarboxylase in the nervous system of the rhesus monkey, J. biol. Chem., 234 (1959) 926-928. 2 Bunney, B. C. and Aghajanian, G. K., The precise localization of nigral afferents in the rat as determined by retrograde tracing technique, Brahl Research, 117 (1976) 423-435. 3 Chalmers, A., McGeer, E. G., Wickson, V. and McGeer, P. L., Distribution of glutamic acid decarboxylase in the brains of various mammalian species, Comp. gen. Pharmacol., I (1970) 385-390. 4 Carter, D. A. and FiNger, H. C., The projections of the entopeduncular nucleus and globus pal[idus in rat as demonstrated by autoradiography and horseradish peroxidase histochemistry, J. eomp. Neurol., 177 (1978) 113 124. 5 Fonnum, F., Radiochemical microassays for the determination of choline acetyltransferase and acetylcholinesterase activities, Biochem. J., 115 (1969) 465 472. 6 Fonnum, F., Grofovfi, I., Rinvik, E., Storm-Mathisen, J. and Walberg, F., Origin and distribution of glutamate decarboxylase in substantia nigra of the cat, Brahl Research, 71 (1974) 77-92.
364 7 Gottesfeld. Z., Massari, V. J., M uth, E. A. and Jacobowitz, D. M., Stria medullaris: a possible pathway containing GABA-ergic afferents to the lateral hahenula, Brain Research. 130 ~1977) 184-189. 8 Grofov~i. I.. The identification of striatal and pallidal neurons projecting to the substantia nigra. An experimental study by means of retrograde axonal transport of horseradish peroxidase, Brait~ Research. 91 (1975) 286-291. 9 Grofovfi.. I. and Rinvik. E., An experimental electron microscopic study on the striato nigral projection in the cat, Exp. Brain Res.. 11 (1970) 249-262. 10 Hajdu, F. R.. Hassler. R. and Bak. I. J.. Electron microscopic study of the substantia nigra and the strio nigral projection in the rat, Z. Zellforsch.. 146 (1973) 207-221 l 1 Hattori, T., FiNger, H. C. and McGeer. P. L.. Demonstration o f a pallido-nigral projection innervating dopaminergic neurons, J. comp. Neurol., 162 (1975) 487-504. 12 Hattori, T., McGeer, P. L., Fibiger, H. C. and McGeer, E. G., On the source of GABA-containing terminals in the substantia nigra. Electron microscopic autoradiographic and biochemical studies. Brain Research, 5411973) 103-t 14. 13 Herkenham, M. and Nauta. W. J H.. Afferent connections of the habenular nuclei m the rat. A horseradish peroxidase study, with a note on the fiber-of-passage problem, J. comp. NeuroL, 173 (1977D 123-145. 14 Herkenham, M. and Nauta. W. J H., Projections of the habenular nuclei in the rat. Anat. Rec.. 187 (1977~ 603. 15 McCaman. R. E. and Dehurst. S. A., Choline actyltransferase in individual neurons of Aplysia californica, J. Neurochem.. 17 (1970) 1421-1426. 16 Kataoka, K.. Bak, I. J., Hassler, R.. Kim, J. S. and Wagner, A.. L-Glutamate decarboxylase and choline acetyltransferase activity in the substantia nigra and the striatum after surgical interruption of the strionigral fibers of the baboon, Exp. Brain Res., 19 (19741 217-227. 17 Kemp, J. M.. The termination of strio-pallidal and strio-nigral fibers. Brain Researd~, t7 (t970) 125-128. 18 Kim, R. K., Nakano, A., Jayaraman, A. and Carpenter. M. B., Projections of the globus pallidus and adjacent structures: An autoradiographic study in the monkey, J, eomp. Nem'ol., 169 (1976) 263-290. 19 McGeer. P. L., Fibiger, H. C., Maler, L.. Hattori. T. and McGeer. E. G., Evidence tot descending paUido-nigral GABA-containmg neurons, Advanc. Neurol.. 5 (1974) 153-159. 20 McGeer, E. G., Fibiger. H. C., McGeer. P. L. and Brooke, S., Temporal changes in amine synthesizing enzymes of rat extrapyramidal structures after hemitransections of 6-hydroxydopamine administration, Brain Research, 52 (1973) 289-300. 21 Nauta. W. J. H.. Evidence o f a pallido habenular pathway in the cat, J. comp. Nemol.. 156 (1974) 9-28. 22 Nauta, W. J. H.. Hippocampal projections and related neural pathways to the midbrain in the cat. Brain, 81 (1958) 319-341. 23 Nauta, W. J. H. and Haymaker. W., Hypothalamic Nuclei and Fiber Connections. W. Haymaker, E. Anderson and W. J. H. Nauta (Eds.), Charles C. Thomas, Springfield, Ill., 1969, pp. 136-209. 24 Nauta, W. J. H. and Mehler, W. R.. Projections of the lentiform nucleus in the monkey, Brain Research, 1 I 1966) 3~42. 25 Niimi, K.. Ikeda, T.. Kawamura. S. and lnoshita, H., Efferent projections of the head of the caudate in the cat, Brain Research. 21 ~1970) 327-344. 26 Wang, R. Y. and Aghajanian. G. K., Physiological evidence lbr habenula as major link between forebrain and midbrain, Science. 197 (1977) 89-91.
Brain Research, 145 I1978i 360-364 , :: Elsevier/North-Holland Biomedical Press
Evidence for a GABA.containing projection from the entopeduncular nucleus to the lateral habenula in the rat
J. I. NAGY, D. A. CARTER, J. LEHMANN and H. C. F1BIGER Division of Neurological Sciences, Department of Psychiatry, University of British Columbia, VancouveG B.C. V6T 1 W5 (Canada)
(Accepted November 3rd, 1977)
Major efferent projections of the striatum and globus pallidus (GP) are known to terminate in the substantia nigra2,S-1Z,lv,19.20,24,z.L the nucleus tegmenti pedunculo pontis (TPP), the subthalamic nucleus and certain thalamic nuclei 2,s,11,1%~. Recently, the internal division of the GP (GPi) in the primate has been shown to project to the lateral habenula is. In addition, the entopeduncular nucleus (EP), the homologue of GPi in subprimate species, has also been shown to project to the lateral habenula in the cat 21. There is evidence that some of the efferent projections of the striatum and G P contain gamma-aminobutyric acid (GABA) and its synthesizing enzyme, glutamic acid decarboxylase (GAD) 6'12"16"zo,2z. In a recent study, Gottesfeld et al. 7 observed that electrolytic lesions of the stria medullaris (SM), GP or GP plus EP significantly decreased G A D activity in the lateral habenula. The authors suggested that certain GABAcontaining neurons in the GP project to the lateral habenula, with a further contribution from the EP. The present investigation was designed to further elucidate the origin of GAD-containing afferents to the lateral habenula. Adult male Wistar rats (300-320 g) received stereotaxic injections of [aH]leucine (0.1-0.15 #1 of 6.5 #Ci/#l, sp.act. 50 Ci/mmole) into the EP (n == 5) or G P (n - 4) via a Hamilton microsyringe. The animals were killed 24 h later and 40/~m frontal sections of the brain were prepared for autoradiography as previously described ~. Another group of rats (n =~ 5) received unilateral stereotaxic electrolytic lesions (2 mA for 30 sec) of the EP through an electrode insulated except at the tip. These animals were killed by cervical fracture 10-14 days postoperatively and the brain was placed on a freezing microtome. Thick (300 ,urn) sections were taken through the habenula and from these sections the left and right habenula was dissected out with the aid of a dissecting microscope. The tissues were homogenized in 100 #1 of 50 mM Tris • acetate buffer p H 6.2, containing Triton X-100 (0.2 ~o v/v). G A D and choline acetyltransferase (CAT) activities were measured in homogenates as previously describedl,a,~,l"% Although injections of [ZH]leucine into the GP resulted in heavy accumulations of autoradiographic grains over the subthalamic nucleus and the substantia nigra 4, no labeling was observed in the habenula of any of the injected animals. In contrast, injections of [aH]leucine into EP resulted in distinct labeling of the lateral habenuta:
361
2,'
'r°~
.
.
.
.
.
:i,~
"
if
B
Fig. I. A: photomicrograph showing injection site of [3H]leucine into the entopeduncular nucleus, ,: 26. B: dark-field photomicrograph showing autoradiographic label in the lateral hahenula 24 h after an injection of [3H]leucine into the entopeduncular nucleus. Note heavy accumulation of grains in the ventrolateral aspect of the nucleus y, 125. C: photomicrograph of electrolytic lesion of the entopeduncular nucleus x 52.
362 TABLE 1 The activity of G A D and CA T in the habemda on the lesioned amt control side after unih:tcra/ electrolytic lesions of'the entopeduncular nucleus' Side
n
GAD
nt~t/es/tltg protein hour
Lesion Control
(~ (~
C,4 ]"
Per cent control
13.6 2.40": 63.6 21.4 ~ 2.34 100.0
* Significantly different from control P
mnoh, s/mg proteh~ hour
Per cent control
122 124
98.4 100.0
8. t 10.8
0.05~
Fig. 1A shows the dense concentration of autoradiographic grains over the [3H ]teucine injection site in the EP. In this. as in other animals, there was a small amount of diffusion either dorsally, along the needle tract, or ventromedially into the lateral hypothalamus. Fig. 1B is a dark field photomicrograph showing the heavy labeling of the ventrolaterat aspect of the lateral habenula in the same animal. A histology of a representative electrol3 tic lesion of the EP is gwen in Fig. 1C. It is apparent that the lesion destroyed most of the EP at this level and extended beyond its limits both dorsally and ventrally. The lesion did not encroach upon the GP. The effects of similar lesions on habenular CAT and G A D activities are g~ven in Table I. EP lesions produced a significant decrease in G A D to 64 0,i of the control value without significantly affecting CAT. The present results have demonstrated the existence of an entoped uncular-lateral habenula projection in the rat. Injections of [~H]leucine produced clearly defined autoradiographic labeling in the ventrotateral aspect of the lateral habenula. Projections of the EP were also identified in the ventrolateral aspect of the ventrolateral -ventroanterior thalamic complex, the parafascicular nucleus and the nucleus tegmenti pedunculopontis. These observations have been described in detail elsewhereL Furthermore. that the autoradiographic labeling in the lateral habenula was due to a projection from EP rather than a result of diffusion of [aH]leucine to other nuclei in the vicinity of the EP has been confirmed using horseradish peroxidase histochemistry ~,t:~. The fact that electrolytic lesions of the EP significantly reduced G A D activity in the h abenula suggests that this is a GABA-containing pathway. Moreover, the failure of the lesions to have an effect on CAT in the habenula points to the specificity of this effect. Gottesfeld et al. 7 reported that electrolytic lesions of GP resulted in a 22'~,; decrease in lateral habenular G A D activity, lesions of GP plus EP produced a 32",(~ decrease, and lesions m stria terminalis a 56% decrease. The present autoradiographic results suggest that the reduced G A D activity observed by Gottesfeld et al. 7 after GP lesions was the result of damage to fibers originating in EP and ascending in the vicinity of the GP. Support for this suggestmn is found in the fact that the EP lesions utilized in the present study produced a similar decrease (34 o/;) in habenular G A D activity as was found after the combined GP-EP lesions used by Gottesfeld et al. 7. ALternatively, the decrease found by Gottesfeld et al. 7 after GP lesions alone may have been due to a ventral encroachment of the lesion upon the substantia innominata ~a- The failure of
363 EP lesions to reduce habenular G A D by more than 36 o,O/ m a y have been due to incomplete destruction of the EP. Alternatively, other proposed afferents to the lateral or medial habenula such as the lateral hypothalamus, nucleus of the diagonal band, substantia innominata, lateral preoptic area, mesencephalic raphe, supra-commissural septum and ventral tegmental area 13 may also contain G A D and explain the higher loss of G A D after stria terminalis lesions 7. The remaining part of G A D in the habenula could be due to local neurons. Furthermore, inasmuch as the entopeduncular projection terminates mainly in the ventrolateral aspect of the lateral habenula (Fig. I B), it is possible that if only this part of the nucleus had been assayed, then a greater decrease in G A D may have been obtained. The main proportion of G A D in habenula is localized to its lateral part ~. Nauta 2a has commented on the potential importance of a projection from the EP to the lateral habenula inasnmch as it may represent a point of communication between the extrapyramidal and limbic systems. In this regard, it is significant that the lateral habenula is known to project to and receive afferents from both of these systems. For example with respect to afferents, the lateral habenula receives a major projection from the entopeduncular nucleus as well as from the septum and preoptic area 13,'~a. The efferent extrapyramidal-related projections of the lateral habenula include the substantia nigra pars compacta, and the ventromedial nucleus of the thalamus2, t4. The efferent limbic-related projections include preoptic area, the lateral hypothalamus and the central gray 14,'~2. The extent to which these two streams are integrated or remain segregated in the lateral habenula awaits further study. In any event, it is suggested that the EP to lateral habenular pathway is an inhibitory link in the extrapyramidal circuit, utilizing the neurotransmitter GABA, by which entopeduncular output may modulate not only nuclei of the extrapyramidal system, but also parts of the limbic system. Wang and Aghajanian ~ have recently shown that the projection from the lateral habenula to the dorsal raphe nucleus is inhibitory. This, together with the present observations, raises the possibility that the extrapyramidal system may modulate the activity of serotonergic neurons in midbrain raphe nuclei via the EP-lateral habenula projection. If other lateral habenular efferents prove to be inhibitory, then the presumed inhibitory GABA pathway from EP to lateral habenula may physiologically disinhibit components of both extrapyramidal and limbic function.
I Albers, R. W. and Brady, R. O., The distribution of glutamate decarboxylase in the nervous system of the rhesus monkey, J. biol. Chem., 234 (1959) 926-928. 2 Bunney, B. C. and Aghajanian, G. K., The precise localization of nigral afferents in the rat as determined by retrograde tracing technique, Brahl Research, 117 (1976) 423-435. 3 Chalmers, A., McGeer, E. G., Wickson, V. and McGeer, P. L., Distribution of glutamic acid decarboxylase in the brains of various mammalian species, Comp. gen. Pharmacol., I (1970) 385-390. 4 Carter, D. A. and FiNger, H. C., The projections of the entopeduncular nucleus and globus pal[idus in rat as demonstrated by autoradiography and horseradish peroxidase histochemistry, J. eomp. Neurol., 177 (1978) 113 124. 5 Fonnum, F., Radiochemical microassays for the determination of choline acetyltransferase and acetylcholinesterase activities, Biochem. J., 115 (1969) 465 472. 6 Fonnum, F., Grofovfi, I., Rinvik, E., Storm-Mathisen, J. and Walberg, F., Origin and distribution of glutamate decarboxylase in substantia nigra of the cat, Brahl Research, 71 (1974) 77-92.
364 7 Gottesfeld. Z., Massari, V. J., M uth, E. A. and Jacobowitz, D. M., Stria medullaris: a possible pathway containing GABA-ergic afferents to the lateral hahenula, Brain Research. 130 ~1977) 184-189. 8 Grofov~i. I.. The identification of striatal and pallidal neurons projecting to the substantia nigra. An experimental study by means of retrograde axonal transport of horseradish peroxidase, Brait~ Research. 91 (1975) 286-291. 9 Grofovfi.. I. and Rinvik. E., An experimental electron microscopic study on the striato nigral projection in the cat, Exp. Brain Res.. 11 (1970) 249-262. 10 Hajdu, F. R.. Hassler. R. and Bak. I. J.. Electron microscopic study of the substantia nigra and the strio nigral projection in the rat, Z. Zellforsch.. 146 (1973) 207-221 l 1 Hattori, T., FiNger, H. C. and McGeer. P. L.. Demonstration o f a pallido-nigral projection innervating dopaminergic neurons, J. comp. Neurol., 162 (1975) 487-504. 12 Hattori, T., McGeer, P. L., Fibiger, H. C. and McGeer, E. G., On the source of GABA-containing terminals in the substantia nigra. Electron microscopic autoradiographic and biochemical studies. Brain Research, 5411973) 103-t 14. 13 Herkenham, M. and Nauta. W. J H.. Afferent connections of the habenular nuclei m the rat. A horseradish peroxidase study, with a note on the fiber-of-passage problem, J. comp. NeuroL, 173 (1977D 123-145. 14 Herkenham, M. and Nauta. W. J H., Projections of the habenular nuclei in the rat. Anat. Rec.. 187 (1977~ 603. 15 McCaman. R. E. and Dehurst. S. A., Choline actyltransferase in individual neurons of Aplysia californica, J. Neurochem.. 17 (1970) 1421-1426. 16 Kataoka, K.. Bak, I. J., Hassler, R.. Kim, J. S. and Wagner, A.. L-Glutamate decarboxylase and choline acetyltransferase activity in the substantia nigra and the striatum after surgical interruption of the strionigral fibers of the baboon, Exp. Brain Res., 19 (19741 217-227. 17 Kemp, J. M.. The termination of strio-pallidal and strio-nigral fibers. Brain Researd~, t7 (t970) 125-128. 18 Kim, R. K., Nakano, A., Jayaraman, A. and Carpenter. M. B., Projections of the globus pallidus and adjacent structures: An autoradiographic study in the monkey, J, eomp. Nem'ol., 169 (1976) 263-290. 19 McGeer. P. L., Fibiger, H. C., Maler, L.. Hattori. T. and McGeer. E. G., Evidence tot descending paUido-nigral GABA-containmg neurons, Advanc. Neurol.. 5 (1974) 153-159. 20 McGeer, E. G., Fibiger. H. C., McGeer. P. L. and Brooke, S., Temporal changes in amine synthesizing enzymes of rat extrapyramidal structures after hemitransections of 6-hydroxydopamine administration, Brain Research, 52 (1973) 289-300. 21 Nauta. W. J. H.. Evidence o f a pallido habenular pathway in the cat, J. comp. Nemol.. 156 (1974) 9-28. 22 Nauta, W. J. H.. Hippocampal projections and related neural pathways to the midbrain in the cat. Brain, 81 (1958) 319-341. 23 Nauta, W. J. H. and Haymaker. W., Hypothalamic Nuclei and Fiber Connections. W. Haymaker, E. Anderson and W. J. H. Nauta (Eds.), Charles C. Thomas, Springfield, Ill., 1969, pp. 136-209. 24 Nauta, W. J. H. and Mehler, W. R.. Projections of the lentiform nucleus in the monkey, Brain Research, 1 I 1966) 3~42. 25 Niimi, K.. Ikeda, T.. Kawamura. S. and lnoshita, H., Efferent projections of the head of the caudate in the cat, Brain Research. 21 ~1970) 327-344. 26 Wang, R. Y. and Aghajanian. G. K., Physiological evidence lbr habenula as major link between forebrain and midbrain, Science. 197 (1977) 89-91.