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Chapter 2 Cholinergic neurons of the rat and primate striatum are morphologically different
C . W . Arbuthnott and P.C. Emson (Eds.) Progress in Brain Research, Vol. 99 0 1993 Elsevier Science Publishers B . V . All rights reserved
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CHAPTER 2
Cholinergic neurons of the rat and primate striatum are morphologically different JCrBme Yelnik, GCrard Percheron, Chantal FranGois and Anouk Garnier Laboratoire de Neurornorphologie inforrnationnelle et de Neurologie expirimentale du rnouvemenl, INSERM U106,Hbpital de la Salp@tri&re,75013 Paris, France
Introduction
It is well-established that the striatum contains a population of cholinergic neurons which can be revealed by using either acetylcholinesterase (AChE) histochemistry or choline acetyltransferase (ChAT) immunohistochemistry (Levey et al., 1983; Eckenstein and Sofroniew, 1983; Mesulam et al., 1984; Woolf, 1991). Cholinergic neurons have been identified in the striatum of rats (Lynch et al., 1972; Butcher and Bilezikjian, 1975; Butcher and Hodge, 1976; Armstrong et al., 1983; Levey et al., 1983; Eckenstein and Sofroniew, 1983; Satoh et al., 1983; Bolam et al., 1984b; Phelps et al., 1985), cats (Kimura et al., 1981; Parent and O’ReillyFromentin, 1982), monkeys (Mesulam et al., 1984; Smith and Parent, 1984; Satoh and Fibiger, 1985; DiFiglia, 1987) and humans (Nagai et al., 1983). In order to disclose the dendritic and axonal morphology of cholinergic neurons, comparisons were made with previous classifications based on Golgiimpregnated material. These classifications were in fact elaborated in different animal species: rats (Lu and Brown, 1977; Danner and Pfister, 1979; Dimova et al., 1980; Chang et al., 1982), mice (Rafols et al., 1989), cats (Kemp and Powell, 1971), dogs (Tanaka, 1980; Leontovich, 1983), monkeys (Fox et al., 1971a,b; DiFiglia et al., 1976) and humans (Leontovich, 1954; Braak and Braak, 1982; Graveland et al., 1985). In addition, a variety of
different criteria such as cell body size, dendritic morphology, number of spines or axon length were used for identifying and classifying neurons. Finally, striatal cholinergic neurons were assumed to be similar in all animal species and their characterization became obscure. The goal of this paper is to compare the morphology of cholinergic neurons in primates and non-primates using data of our recent morphological taxonomy of the neurons of the primate striatum (Yelnik et al., 1991). The dendritic morphology of striatal neurons in primates
Neurons of the striatum of monkey and human were analyzed in adult brains impregnated following the Golgi method of Anderson (1954) or Davenport and Combs (1954). Dendritic arborizations were reconstructed from serial sections and digitized in three dimensions with the aid of a computer-aided technique (Yelnik et al., 1981). Their morphological features were described using topological, metrical and geometrical quantitative parameters. Topological parameters comprised the number of stems and tips, the dendritic formula, the branching index and the stature (Percheron, 1979, 1982). Metrical parameters comprised the mean length of dendritic segments (all segments, stems, internodes, twigs), the total dendritic length and the longest dendrite. Geometrical parameters consisted of the dimensions of
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the dendritic arborization (length, width, thickness) as measured with reference to its principal system of axes (Yelnik et al., 1983). Neuronal species, defined as sets of neurons having statistically similar morphological features, were isolated on the basis of multidimensional statistical tests. Four neuronal species were identified in primates (Yelnik et al., 1991). Spiny neurons (Fig. 1) represent the bulk (96%) of striatal neurons. They have different topological and metrical parameters in monkeys and humans: a dendritic formula of 5 - 35 (5 stems, 35 tips) in monkeys and 6 - 42 in humans, a mean length of dendritic segments of 93 pm in monkeys and 75 pm in humans. They have also a higher density of dendritic spines in monkeys (8 per 10 pm) thanin humans (2.6per 10 pm). Conversely, the geometry of dendritic arborizations (430 x 330 x 230 pm) is remarkably similar in both species. Leptodendritic neurons (2%) d o not differ in monkeys and humans and are statistically undistinguishable from pallidal (Yelnik et al., 1984) and nigral pars reticulata (Yelnik et al., 1987) neurons. They are mainly characterized by very long and sparsely ramified dendrites (Fig. 2B): the dendritic formula is 4 - 20, the dendritic segments are 196 pm long and the dendritic arborization is 1200 pm long (up to 1600 pm). Spidery neurons (1 YO)are immediately recognizable with their large, globular cell body and their very thick dendritic stems which branch profusely into varicose processes which curve back toward the soma (Fig. 2 0 . They have a dendritic formula of 12 - 129, a mean length of dendritic segments of 95 pm and a dendritic arborization of 600 pm long. Microneurons (1 070) have a dendritic formula of 6 - 64, a mean length of dendritic segments of 40 pm and a dendritic arborization of 240 pm long. These four neuronal species can be found, under different namings, in previous studies (table 2 in Yelnik et al., 1991). Differences bear mainly on the
classification of large neurons. Fox et al. (197 lb) illustrate leptodendritic neurons (their figs. 3, 16) and spidery neurons (their figs. 4, 8) but classified both types as “large aspiny neurons”. DiFiglia et al. (1976) classified leptodendritic neurons as “large version of spiny type 11” and spidery neurons as “aspiny type 11”. In the classification of Braak and Braak (1982), they appear as type I11 and type IV respectively. Characterization of cholinergic neurons in the primate striatum
The brain of one monkey (Macaca irus) was sectioned on a freezing microtome and processed according to the DFP-histochemical technique of Poirier et al. (1977) to reveal AChE-containing neurons. Sections were couterstained with neutral red or cresyl violet. Samples of both AChE-containing and unlabeled cell bodies were selected for quantitative analysis. The largest contour of each cell body was digitized on an XY digitizer and its area was measured using the formula of Pullen (1984). The biggest and smallest diameters were determined by program. A shape index (biggest/smallest diameter) was calculated. This procedure was also applied to the cell bodies of Golgi-impregnated neurons. By this way, AChE-containing cell bodies could be correlated quantitatively with the neuronal species defined previously on the basis of dendritic features. Only three types of cell bodies are identifiable on the basis of their size and shape in the primate striatum. Indeed, spiny neurons and microneurons have both small and round cell bodies and are not distinguishable on these somatic criteria. Spidery neurons have round cell bodies which are the largest cell bodies of the primate striatum. Leptodendritic neurons have an elongated cell body whose size is intermediate. Finally the three types of cell bodies are: small round, medium elongated and large round. This is
Fig. 1. Spiny neurons of the striatum of human ( A ) , monkeys (B)and rats (0.Each dendritic arborization was reconstructed from serial sections and drawn through the x 100 immersion objective. Note that the overall dimensions of arborizations are not significantly different in the three species. Conversely, there are differences in the number of dendritic segments and in their density of dendritic spines (see text). Golgi method.
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B
1-1
100pm
Fig. 2. Three of the four neuronal species of the monkey (Pupiopupio) striatum: spiny neuron ( A ) , leptodendritic neuron (B), spidery neuron (0.Golgi method. Same magnification for the three neurons. Note that the spidery neuron has the largest cell body (1406 pm2), the leptodendritic neuron is medium-sized (684 pm’) and the spiny neuron has a small round cell body (254 prn2).
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in agreement with the distribution of cell bodies as it appears in Nissl-stained sections (fig. 1 in Pasik et al., 1979; fig. 11 in Yelnik et al., 1991). In material processed for revelation of AChE, positive cell bodies were large and round. They were larger than all non-labeled counterstained neurons including both medium elongated and small round cell bodies (Yelnik et al., 1991). As AChE histochemistry has been demonstrated to closely correspond to ChAT immunoreactivity in the striatum of macaques (Mesulam et al., 1984), it can be concluded that AChEpositive neurons are the cholinergic neurons of the striatum of monkeys. As they have the largest cell bodies of the striatum, it can be concluded that they correspond to spidery neurons. This conclusion, suggested by Smith and Parent (1984), is in agreement with the electron microscope analysis of DiFiglia (1987). Cholinergic neurons, or spidery neurons, of the primate striatum are thus characterized by a voluminous and globular cell body which is the largest one of all other striatal neurons in primates. They correspond to the larger achromatic cells of cytoarchitectonic preparations. They have a large number (12) of very thick (up to 10 pm) dendritic stems. These stems branch rapidly into numerous branches which become thinner and thinner at each bifurcation. Distal branches are very numerous (129) and most of them are thin and varicose. They often curve back toward the soma. As a whole, the dendritic arborization of spidery neurons is highly recognizable on its high density and the curved and varicose aspect of its distal processes (Fig. 2 0 .
The dendritic morphology of striatal neurons in rats Golgi-impregnated brains of adult rats were also examined for this study. Rats were perfused through the aorta with 1% paraformaldehyde and 1% glutaraldehyde in 0.1 M phosphate buffer. Brains were processed by the Golgi method according to the modification of Van der Loos (1956). Neurons were drawn through the camera lucida ( x 100 immersion objective) and analyzed qualitatively. Some of
them, which could be reconstructed from serial sections, were analyzed quantitatively according to our computer-aided method. The overall distribution of striatal neurons in the rat is the same as in primates, i.e., comprising about 96% spiny neurons and a few neurons having a larger cell body than spiny neurons. Spiny neurons of the rat striatum (Fig. 1C) have lower topological and metrical parameters than spiny neurons of monkey and human. The longest dendrite in rats was 220 pm (Fig. lC), 200- 250 pm (Kitai et al., 1979), 280 pm (Wilson and Groves, 1980), 200 - 220 pm (Dimova et al., 1980), 220 pm (Bishop et al., 1982), which is smaller than the 275 pm measured in primates (Yelnik et al., 1991). The mean length of dendritic segments (67 pm in Fig. 1C) is also smaller than in monkeys (93 pm) and humans (75 pm), and so is the dendritic formula: 6 - 30 for the rat, 5 - 35 for the monkey and 6-42 for the human. Conversely, the dimensions of dendritic arborizations were 360 x 230 x 180 pm in Fig. 1C and 260 - 530 x 240-425 x 260- 350 in Preston et al. (1980), which is similar t o the 430 x 330 x 230 pm that we measured in primates (Yelnik et al., 1991). Spine distribution (5 per 10 pm in Fig. 1C) was intermediate. Two different types of large neurons were identifiable in our material. Neurons of the first type (Fig. 3B) had a fusiform cell body and 3 - 4 dendritic trees with long, smooth and sparsely branched dendrites (9 - 11 dendritic tips). They correspond to the large type I of Chang et al. (1982) and Chang and Kitai (1982) and resemble theleptodendritic neurons of primates. However, their dendrites were definitely thinner and shorter (400 pm for the longest one) than their primate counterpart (compare Figs. 2Band 3B). This observation is consistent with previous measurements in the rat: up to 150 pm (Takagi et al., 1984), 250 pm (Bolam et al., 1984a), 350 pm (Rafols et al., 1989), 400 pm (Chang et al., 1982) and 450 pm (Bishop et al., 1982). As the longest dendrite was 1270 pm in primates (Yelnik et al., 1991), this could suggest that large neurons of rats constitute a particular type. However, as they have a low dendritic formula and relatively long dendrites, they fit with the criteria of Ramon-Moliner
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A
B
C
4-1
100pm
Fig. 3. Three neuronal types of the rat striatum. Golgi method. Same magnification as in Fig. 2. A . Spiny neuron with a small cell body (135 pm2). B . Two large neurons (497 and 392 pm’) with elongated cell bodies and sparsely ramified dendrites. Note that the cell bodieq are smaller and the dendrites far shorter than for leptodendritic neurons of primates (Fig. 2B). C . Two large neurons (471 and 430 pm’) with round cell bodies and more numerous, varicose and tortuous dendrites. Note that these neurons strongly differ from the spidery neurons of primates shown in Fig. 2C.
(1969) and are likely to represent the leptodendritic neurons of the rat striatum. The second type of large neurons that we observed (Fig. 3C) had a round or polygonal cell body with 4 - 6 dendritic trees, As a whole the dendritic arbori-
zation had 21 - 28 dendritic tips. The mean length of segments was 40 - 60 pm, the longest dendrite was 200 - 250 pm, the total dendritic length 1400 2500 pm. This type is similar to the large type I1 of Chang et al. (1982) and Chang and Kitai (1982). Ac-
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cording to Bolam et al. (1984a), this type would be similar to the type IV of Braak and Braak (1982) in humans and the aspiny type I1 of DiFiglia et al. (1976) in monkeys, i.e., our spidery neurons. In our opinion, however, this comparison does not hold true because of significant differences, both qualitative (compare Figs. 2 C and 3C) and quantitative: 21 -28 tips vs. 129 in primates, a total dendritic length of 1400 - 2500 Fm vs. 23400 pm in primates. This strongly suggests that there are no spidery neurons in the rat striatum, which raises the question of the morphology of the rat cholinergic neurons.
Characterization of cholinergic neurons in the rat striatum The first studies of cholinergic neurons in nonprimate species were based on AChE histochemistry (Lynch et al., 1972; Butcher and Bilezikjian, 1975; Butcher and Hodge, 1976; Henderson, 1981; Armstrong et al., 1983) and concluded that two populations of neurons, one with medium-sized cell bodies, the other with large fusiform or multipolar cell bodies, were AChE-positive. Later, however, Eckenstein and Sofroniew (1983), Levey et al. (1983) and Phelps et al. (1985) using immunohistochemical detection of ChAT concluded that the apparent heterogeneity of ChAT-positive cell bodies might be explained by different orientations of elongated cell bodies and that cholinergic neurons constituted a single population of large neurons. Moreover, Bolam et al. (1984b) demonstrated, by combining Golgi impregnation, electron microscopy, AChE histochemistry and ChAT immunohistochemistry, that only one out of three populations of AChEpositive neurons (Bolam et al., 1984a) were immunoreactive to ChAT. For Bolam et al. (1984b), cholinergic neurons have a large triangular or multipolar cell body, “long infrequently branching dendrites”, and correlate with the giant neurons of Kemp and Powell (1971), the type V of Dimova et al. (1980), the giant neurons of Danner and Pfister (1981) and the type I large of Chang et al. (1982). These different types correspond to the leptodendritic neuron of Fig. 3B.
Bolam et al. (1984b) specify that cholinergic neurons are not “the same as the large neurons with a ‘spidery’ appearance and ‘swirling’ dendrites”, i.e., their type 2 which comprises the type I1 large of Chang et al. (1982) and their large version type 3 AChE-positive (Bolam et al., 1984a). Phelps et al. (1985) described ChAT-immunoreactive neurons as having large oval or multipolar somata and “three to four primary dendrites that branch and extend long distance”. Such a description is also that of a leptodendritic neuron. Kubota et al. (1987) described ChAT-immunoreactive neurons as having a large, oval, spindle, triangular or multipolar cell body, 3-5 dendritic trees emerging from the poles of the soma, and long and sparsely branched dendrites. They correlated these cholinergic neurons with the giant neurons of Kemp and Powell (1971) and the type I large of Chang et al. (1982), thus, also with leptodendritic neurons.
Conclusions Finally it appears from our results and from the data of the literature that: (1) cholinergic neurons of the striatum have the largest cell bodies of this cerebral region in both primate and non-primate species; (2) those striatal neurons having the largest cell body in the striatum are leptodendritic neurons in nonprimate species whereas they are spidery neurons in monkeys and man; (3) cholinergic neurons of nonprimate species are leptodendritic neurons whereas they are spidery neurons in primates; and (4) spidery neurons do not exist in non-primate species. The morphological difference between striatal cholinergic neurons of the rat and primate is considerable and raises several questions. One concerns the differentiation of the neuronal species of the striatum through phylogenesis. A first important point is that the dimensions of the dendritic arborizations of spiny neurons do not change from rats to humans. This implies that the grain of the dendritic lattice does not change (Yelnik et al., 1991), which is likely to be a fundamental feature of the striatal organization. Conversely, the present results demonstrate that leptodendritic neurons are subject to a
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considerable increase of dendritic length from rats to primates. Such an increase was also observed for the leptodendritic neurons of the globus pallidus (700 pm in rats, 1200 pm in primates; Yelnik et al., 1984) and substantia nigra pars reticulata (900 pm in rats, 1100 pm in macaques, 1600 pm in humans; Yelnik et al., 1987) but it does not represent a general rule of phylogenesis. For example, subthalamic neurons, as well as spiny neurons, remain unchanged from rats and cats to monkey and human (Yelnik and Percheron, 1979; Hammond and Yelnik, 1983). In rats, cholinergic neurons of the striatum are of the same morphological type as the cholinergic neurons of the nucleus basalis of Meynert and the peripallidal laminae (Ingham et al., 1985)and even of other brain regions (Woolf, 1991). Thus, in rats, there is a correlation between morphology and chemical content. In primates, conversely, cholinergic neurons of the striatum are definitely different from the cholinergic neurons of the basal forebrain. They have highly specific morphological features which are not observed in other regions of the nervous system. Pasik et al. (1991) have recently raised the question of the axonal nature of the cholinergic neurons of the striatum. As cholinergic neurons in rats are leptodendritic neurons and as leptodendritic neurons in primates (their large spiny 11) are projection neurons, they deduce that cholinergic neurons of rats should be projection neurons. However, the interneuronal nature of cholinergic neurons has been demonstrated by fluorescent tracer experiments (Woolf and Butcher, 1981). In addition, leptodendritic neurons projecting to the substantia nigra have been shown in only the ventral part of the rat striatum (Bolam et al., 1981), which suggests that the remaining leptodendritic neurons of the rat striatum, i.e., the cholinergic neurons, could be interneurons. An abundant literature has been devoted to the functional role of cholinergic neurons but it has never taken into account interspecific morphological differences. In Woolf's (1991) review for example, cholinergic neurons are considered as a global system contrasting with the modularity of sensory systems. While this is likely to be true for the
cholinergic neurons of the rat striatum, it does not seem to be applicable to spidery neurons, the cholinergic neurons of the primate striatum, which have relatively small but very dense arborizations. As they represent only 1% of striatal neurons, they constitute a dendritic lattice which is characterized by zones of high dendritic density alternating with zones which are almost free of cholinergic processes (see fig. 12 in Yelnik et al., 1991). This implies that groups of spiny neurons could be strongly controlled by the activity of cholinergic neurons while others could not. Therefore, the anatomical organization of the cholinergic system of the striatum is submitted to a considerable transformation from rats to humans. An evolution from a globally organized system to a fine-grain modular system is certainly a highly significant feature of the primate striatum, which should be carefully considered in the analysis of the function of the striatum.
References Anderson, A.D. (1954) Alcoholic thionin counterstaining following Golgi dichromate-silver impregnation. Stain Techno/., 29: 179- 184. Armstrong, D.M., Saper, C.B., Levey, A.I., Wainer, B.H. and Terry, R.D. (1983) Distribution of cholinergic neurons in rat brain: demonstrated by the immunocytochemical localization of choline acetyltransferase. J . Comp. Neurol., 216: 53 - 68. Bishop, G.A., Chang, H.T. and Kitai, S.T. (1982) Morphological and physiological properties of neostriatal neurons: an intracellular horseradish peroxidase study in the rat. Neuroscience, 7: 179- 191. Bolam, J.P., Somogyi, P., Totterdell, S. and Smith, A.D. (1981) A second type of striatonigral neuron: a comparison between retrogradely labelled and Golgi-stained neurons at the light and electron microscopic levels. Neuroscience, 6 : 2141 - 21 57. Bolam, J.P., Ingham, C.A. and Smith, A.D. (1984a) The section-Golgi-impregnation procedure, 3. Combination of Golgi-impregnation with enzyme histochemistry and electron microscopy to characterize acetylcholinesterase-containing neurons in the rat striatum. Neuroscience, 12: 687 - 709. Bolam, J.P., Wainer, B.H. and Smith, A.D. (1984b) Characterization of cholinergic neurons in the rat neostriatum. A combination of choline acetyltransferase immunocytochemistry, Golgi impregnation and electron microscopy. Neuroscience. 12: 71 1 - 718. Braak, H. and Braak, E. (1982) Neuronal types in the striatum of man. Cell Tissue Res., 227: 319-342. Butcher, L.L. and Bilezikjian, L. (1975) Acetylcholinesterase-
33 containing neurons in the neostriatum and substantia nigra revealed after punctate intracerebral injection of diisopropylfluorophosphate. Eur. J . Phurrnacol., 34: 115 - 125. Butcher, L.L. and Hodge, G.K. (1976)Postnatal development of acetylcholinesterase in the caudate-putamen nucleus and substantia nigra of rats. Bruin Rex, 106:223 - 240. Chang, H.T. and Kitai, S.T. (1982)Large neostriatal neurons in the rat: an electron microscopic study of gold-toned Golgistained cells. Bruin Res. Bull., 8:631 -643. Chang, H.T., Wilson, C.J. andKitai, S.T. (1982)A Golgi study of the rat neostriatal neurons: light microscopic analysis. J. Cornp. Neurol., 208: 107- 126. Danner, H. and Pfister, C. (1979)Untersuchungen zur Struktur der Neostriatum des Ratte. J. Hirnforsch., 20: 285 -301. Danner, H. and Pfister, C. (1981)4 Spine-lose Neurontypen im Neostriatum der Ratte. 1.Hirnforsch., 22: 465 - 477. Davenport, H.A. and Combs, C.M. (1954)Golgi’s dichromatesivler method. 3. Chromating fluids. Stain Techno/., 29: 165 - 173. DiFiglia, M. (1987)Synaptic organization of cholinergic neurons in themonkey neostriatum. J. Cornp. Neurol., 255:245 - 258. DiFiglia, M., Pasik, P . and Pasik, T. (1976)A Golgi study of neuronal types in the neostriatum of monkeys. Brain Res., 114:245 - 256. Dimova, R., Vuillet, J. and Seite, R. (1980)Study of the rat neostriatum using a combined Golgi electron microscope technique and serial sections. Neuroscience, 5: 1581 - 1596. Eckenstein, F. and Sofroniew, M.V. (1983)Identification of central cholinergic neurons containing both choline acetyltransferase and acetylcholinesterase and of central neurons containing only acetylcholinesterase. J. Neurosci., 3:2286 - 2291. Fox, C.A., Andrade, A.N., Hillman, D.E. and Schwyn, R.C. (1971a)The spiny neurons in the primate striatum: a Golgi and electron microscopic study. J. Hirnforsch., 13: 181 - 201. Fox, C.A., Andrade, A.N., Schwyn, R.C. and Rafols, J.A. (1971b) The aspiny neurons and the glia in the primate striatum: a Golgi and electron microscopic study. J. Hirnforsch., 13: 341 - 362. Graveland, G.A., Williams, R.S. and DiFiglia, M. (1985) A Golgi study of the human neostriatum: neurons and afferent fibers. J. Cornp. Neurol., 34: 317 - 333. Hammond, C. and Yelnik, J. (1983)Intracellular labelling of rat subthalamic neurones with horseradish peroxidase: computer analysis of dendrites and characterization of axon arborization. Neuroscience, 8: 781 - 790. Henderson, 2. (1981) Ultrastructure and acetylcholinesterase content of neurons forming connections between the striatum and substantianigraof rat. J. Cornp. Neurol., 197: 185 - 196. Ingham, C.A., Bolam, J.P., Wainer, B.H. and Smith, A.D. (1985)A correlated light and electron microscopic study of identified cholinergic basal forebrain neurons that project to the cortex in the rat. J. Comp. Neurol., 239: 176- 192. Kemp, J.M. and Powell, T.P.S. (1971)The structure of the caudate nucleus of the cat: light and electron microscopy. Phil.
Trans. R . SOC.Lond. (Biol.), 262: 383 - 401. Kimura,H.,McGeer,P,L.,Peng, J.H.andMcGeer,E.G.(1981) The central cholinergic system studied by choline acetyltransferase immunohistochemistry in the cat. J. Cornp. Neurol., 200: 151-201. Kitai, S.T., Preston, R.J., Bishop,G.A. andKocsis, J.D. (1979) Striatal projection neurons: morphological and electrophysiological studies. In: L.J. Pokier, T.L. Sourkes and P.J. Bedard (Eds.), Advances in Neurology, Vol. 24, Raven Press, New York, pp. 45-51. Kubota, Y., Inagaki, S., Shimada, S., Kito, S., Eckenstein, F. and Tohyama, M. (1987) Neostriatal cholinergic neurons receive direct synaptic inputs from dopaminergic axons. Bruin Res., 413: 179- 184. Leontovich, T.A. (1954)Fine structure of subcortical ganglia (in Russian). Z.Nevropat. Psikh., 54: 168 - 178. Leontovich, T.A. (1983)Spatial arrangement of tissue elements of nucleus caudatus in dog (in Russian). Neurophysiology, IS: 474 - 484. Levey, A.I., Wainer, B.H., Mufson, E. J . and Mesulam, M.-M. (1983) Co-localization of acetylcholinesterase and choline acetyltransferase in the rat cerebrum. Neuroscience, 9:9 - 22. Lu, E.J. and Brown, W.J. (1977) The developing caudate nucleus in the euthyroid and hypothyroid rat. J . Cornp. Neuro/., 171: 261 -284. Lynch, G.S., Lucas, P.A. and Deadwyler, S.A. (1972) The demonstration of acetylcholinesterase containing neurones within the caudate nucleus of the rat. Brain Res., 45: 617-621. Mesulam, M.-M., Mufson, E.J., Levey, A.1. and Wainer, B.H. (1984)Atlas of cholinergic neurons in the forebrain and upper brainstem of the macaque based on monoclonal choline acetyltransferase immunohistochemistry and acetylcholinesterase histochemistry. Neuroscience, 12:669 - 686. Nagai, T., Pearson, T., Peng, F., McGeer, E.G. and McGeer, P.L. (1983)Immunohistochemical staining of the human forebrain with monoclonal antibody to human choline acetyltransferase. Brain Rex, 265: 300 - 306. Parent, A. and O’Reilly-Fromentin, J. (1982)Distribution and morphological characteristics of acetylcholinesterase-containing neurons in the basal forebrain of the cat. Brain Res. Bull., 8: 183 - 196. Pasik, P., Pasik, T. and DiFiglia, M. (1979)The internal organization of the neostriatum of mammals. In: 1. Divac and R.G.E. Oberg (Eds.), The Neostriaturn, Pergamon Press, New York, pp. 5 - 36. Pasik, P., Pasik, T. and Holstein, G.R. (1991)The ultrastructural chemoanatomy of the basal ganglia: 1984-1989.I. The neostriatum. In: G. Bernardi et al. (Eds.), The Basal Ganglia III, Plenum, New York, pp. 187- 197. Percheron, G. (1979)Quantitative analysis of dendritic branching. 1. Simple formulae for the quantitative analysis of dendritic branching. Neurosci. Lett., 14: 287 - 293. Percheron, G. (1982)Principles and methods of the graph theo-
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retical analysis of natural binary arborescences. J. Theor. Biol., 99: 509 - 552. Phelps, P.E., Houser, C.R. and Vaughn, J.E. (1985) Immunocytochemical localization of choline acetyltransferase within the rat neostriatum: a correlated light and electron microscopic study of cholinergic neurons and synapses. J . Comp. Neurol., 238: 286- 307. Poirier, L.J., Parent, A , , Marchand, R. and Butcher, L.L. (1977) Morphological characteristics of the acetylcholinesterase-containing neurons in the CNS of DFPtreated monkeys. Part 1 . Extrapyramidal and related structures. J. Neurol. Sci., 31: 181 - 198. Preston, R.J., Bishop, G.A. and Kitai, S.T. (1980) Medium spiny neuron projection from the rat neostriatum: an intracelMar horseradish peroxidase study. Brain Res., 183: 253 - 263. Pullen, A.H. (1984) A structured program in Basic for cell morphometry: its application to the spinal motoneurone. J. Neurosci. Methods, 12: 1 5 5 - 178. Rafols, J.A., Cheng, H.W. and McNeill, T.H. (1989) Golgi study of the mouse striatum: age-related dendritic changes in different neuronal populations. J. Comp. Neurol., 279: 2 12 - 227. Ramon-Moliner, E. (1969) The leptodendritic neuron: its distribution and significance. Ann. N. Y. Acad. Sci., 167: 65 - 70. Satoh, K. and Fibiger, H.C. (1985) Distribution of central cholinergic neurons in the baboon (Pupio pupio). I. General morphology. J . Comp. Neurol., 236: 197-214. Satoh, K., Staines, W.A., Atmadja, S. and Fibiger, H.C. (1983) Ultrastructural observations of the cholinergic neuron in the rat striatum as identified by acetylcholinesterase pharmacohistochemistry. Neuroscience, 10: 1121 - 1136. Smith, Y. and Parent, A. (1984) Distribution of acetylcholinesterase-containing neurons in the basal forebrain and upper brainstem of the squirrel monkey (Suimirisciureus). Bruin Res. Bull., 12: 95 - 104. Takagi,H.,Somogyi,P.andSmith,A.D. (1984)Aspinyneurons and their local axons in the neostriatum of the rat: a correlated light and electron microscopic study of Golgi-impregnated material. J. Neurocytol., 13: 239 - 265.
Tanaka Jr., D. (1980) Development of spiny and aspiny neurons in the caudate nucleus of the dog during the first postnatal month. J. Comp. Neurol., 192: 241-263. Van der Loos, H. (1956) Une combinaison de deux vieilles methodes histologiques pour le systeme nerveux central. Mschr. Psychiatr. Neurol., 132: 330- 334. Wilson, C.J. andGroves, P.M. (1980) Fine structure and synaptic connections of the common spiny neuron of the rat neostriatum: a study employing intracellular injection of horseradish peroxidase. J. Comp. Neurol., 194: 599-616. Woolf, N. J . (1991) Cholinergic systems in mammalian brain and spinal cord. Prog. Neurobiol., 37: 475 - 524. Woolf, N.J. and Butcher, L.L. (1981)Cholinergic neurons in the caudate-putamen complex proper are intrinsically organized: a combined Evans Blue and acetylcholinesterase analysis. Brain Res. Bull., 7: 487 - 507. Yelnik, J . and Percheron, G. (1979) Subthalamic neurons in primates: a quantitative and comparative analysis. Neuroscience, 4: 1117-1143. Yelnik, J., Percheron, G . , Perbos, J. and Francois, C. (1981) A computer-aided method for the quantitative analysis of dendritic arborizations reconstructed from serial sections. J . Neurosci. Methods, 4: 347 - 364. Yelnik, J., Percheron, G., Francois, C. and Burnod, Y . (1983) Principal component analysis: a suitable method for the threedimensional study of the shape, dimensions and orientation of dendritic arborizations. J. Neurosci. Methods, 9: 115 - 125. Yelnik, J ., Percheron, G . and Francois, C. (1984) A Golgi analysis of the primate globus pallidus. 11. Quantitative morphology and spatial orientation of dendritic arborizations. J. Comp. Neurol., 221: 200-213. Yelnik, J . , Francois, C., Percheron, G . and Heyner, S. (1987) Golgi study of the primate substantia nigra I. Quantitative morphology and typology of nigral neurons. J. Comp. Neurol., 265: 455 - 472. Yelnik, J., Francois, C., Percheron, G. and Tande, D. (1991) Morphological taxonomy of the neurons of the primate striaturn. J. Comp. Neurol., 313: 273 - 294.
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CHAPTER 2
Cholinergic neurons of the rat and primate striatum are morphologically different JCrBme Yelnik, GCrard Percheron, Chantal FranGois and Anouk Garnier Laboratoire de Neurornorphologie inforrnationnelle et de Neurologie expirimentale du rnouvemenl, INSERM U106,Hbpital de la Salp@tri&re,75013 Paris, France
Introduction
It is well-established that the striatum contains a population of cholinergic neurons which can be revealed by using either acetylcholinesterase (AChE) histochemistry or choline acetyltransferase (ChAT) immunohistochemistry (Levey et al., 1983; Eckenstein and Sofroniew, 1983; Mesulam et al., 1984; Woolf, 1991). Cholinergic neurons have been identified in the striatum of rats (Lynch et al., 1972; Butcher and Bilezikjian, 1975; Butcher and Hodge, 1976; Armstrong et al., 1983; Levey et al., 1983; Eckenstein and Sofroniew, 1983; Satoh et al., 1983; Bolam et al., 1984b; Phelps et al., 1985), cats (Kimura et al., 1981; Parent and O’ReillyFromentin, 1982), monkeys (Mesulam et al., 1984; Smith and Parent, 1984; Satoh and Fibiger, 1985; DiFiglia, 1987) and humans (Nagai et al., 1983). In order to disclose the dendritic and axonal morphology of cholinergic neurons, comparisons were made with previous classifications based on Golgiimpregnated material. These classifications were in fact elaborated in different animal species: rats (Lu and Brown, 1977; Danner and Pfister, 1979; Dimova et al., 1980; Chang et al., 1982), mice (Rafols et al., 1989), cats (Kemp and Powell, 1971), dogs (Tanaka, 1980; Leontovich, 1983), monkeys (Fox et al., 1971a,b; DiFiglia et al., 1976) and humans (Leontovich, 1954; Braak and Braak, 1982; Graveland et al., 1985). In addition, a variety of
different criteria such as cell body size, dendritic morphology, number of spines or axon length were used for identifying and classifying neurons. Finally, striatal cholinergic neurons were assumed to be similar in all animal species and their characterization became obscure. The goal of this paper is to compare the morphology of cholinergic neurons in primates and non-primates using data of our recent morphological taxonomy of the neurons of the primate striatum (Yelnik et al., 1991). The dendritic morphology of striatal neurons in primates
Neurons of the striatum of monkey and human were analyzed in adult brains impregnated following the Golgi method of Anderson (1954) or Davenport and Combs (1954). Dendritic arborizations were reconstructed from serial sections and digitized in three dimensions with the aid of a computer-aided technique (Yelnik et al., 1981). Their morphological features were described using topological, metrical and geometrical quantitative parameters. Topological parameters comprised the number of stems and tips, the dendritic formula, the branching index and the stature (Percheron, 1979, 1982). Metrical parameters comprised the mean length of dendritic segments (all segments, stems, internodes, twigs), the total dendritic length and the longest dendrite. Geometrical parameters consisted of the dimensions of
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the dendritic arborization (length, width, thickness) as measured with reference to its principal system of axes (Yelnik et al., 1983). Neuronal species, defined as sets of neurons having statistically similar morphological features, were isolated on the basis of multidimensional statistical tests. Four neuronal species were identified in primates (Yelnik et al., 1991). Spiny neurons (Fig. 1) represent the bulk (96%) of striatal neurons. They have different topological and metrical parameters in monkeys and humans: a dendritic formula of 5 - 35 (5 stems, 35 tips) in monkeys and 6 - 42 in humans, a mean length of dendritic segments of 93 pm in monkeys and 75 pm in humans. They have also a higher density of dendritic spines in monkeys (8 per 10 pm) thanin humans (2.6per 10 pm). Conversely, the geometry of dendritic arborizations (430 x 330 x 230 pm) is remarkably similar in both species. Leptodendritic neurons (2%) d o not differ in monkeys and humans and are statistically undistinguishable from pallidal (Yelnik et al., 1984) and nigral pars reticulata (Yelnik et al., 1987) neurons. They are mainly characterized by very long and sparsely ramified dendrites (Fig. 2B): the dendritic formula is 4 - 20, the dendritic segments are 196 pm long and the dendritic arborization is 1200 pm long (up to 1600 pm). Spidery neurons (1 YO)are immediately recognizable with their large, globular cell body and their very thick dendritic stems which branch profusely into varicose processes which curve back toward the soma (Fig. 2 0 . They have a dendritic formula of 12 - 129, a mean length of dendritic segments of 95 pm and a dendritic arborization of 600 pm long. Microneurons (1 070) have a dendritic formula of 6 - 64, a mean length of dendritic segments of 40 pm and a dendritic arborization of 240 pm long. These four neuronal species can be found, under different namings, in previous studies (table 2 in Yelnik et al., 1991). Differences bear mainly on the
classification of large neurons. Fox et al. (197 lb) illustrate leptodendritic neurons (their figs. 3, 16) and spidery neurons (their figs. 4, 8) but classified both types as “large aspiny neurons”. DiFiglia et al. (1976) classified leptodendritic neurons as “large version of spiny type 11” and spidery neurons as “aspiny type 11”. In the classification of Braak and Braak (1982), they appear as type I11 and type IV respectively. Characterization of cholinergic neurons in the primate striatum
The brain of one monkey (Macaca irus) was sectioned on a freezing microtome and processed according to the DFP-histochemical technique of Poirier et al. (1977) to reveal AChE-containing neurons. Sections were couterstained with neutral red or cresyl violet. Samples of both AChE-containing and unlabeled cell bodies were selected for quantitative analysis. The largest contour of each cell body was digitized on an XY digitizer and its area was measured using the formula of Pullen (1984). The biggest and smallest diameters were determined by program. A shape index (biggest/smallest diameter) was calculated. This procedure was also applied to the cell bodies of Golgi-impregnated neurons. By this way, AChE-containing cell bodies could be correlated quantitatively with the neuronal species defined previously on the basis of dendritic features. Only three types of cell bodies are identifiable on the basis of their size and shape in the primate striatum. Indeed, spiny neurons and microneurons have both small and round cell bodies and are not distinguishable on these somatic criteria. Spidery neurons have round cell bodies which are the largest cell bodies of the primate striatum. Leptodendritic neurons have an elongated cell body whose size is intermediate. Finally the three types of cell bodies are: small round, medium elongated and large round. This is
Fig. 1. Spiny neurons of the striatum of human ( A ) , monkeys (B)and rats (0.Each dendritic arborization was reconstructed from serial sections and drawn through the x 100 immersion objective. Note that the overall dimensions of arborizations are not significantly different in the three species. Conversely, there are differences in the number of dendritic segments and in their density of dendritic spines (see text). Golgi method.
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B
1-1
100pm
Fig. 2. Three of the four neuronal species of the monkey (Pupiopupio) striatum: spiny neuron ( A ) , leptodendritic neuron (B), spidery neuron (0.Golgi method. Same magnification for the three neurons. Note that the spidery neuron has the largest cell body (1406 pm2), the leptodendritic neuron is medium-sized (684 pm’) and the spiny neuron has a small round cell body (254 prn2).
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in agreement with the distribution of cell bodies as it appears in Nissl-stained sections (fig. 1 in Pasik et al., 1979; fig. 11 in Yelnik et al., 1991). In material processed for revelation of AChE, positive cell bodies were large and round. They were larger than all non-labeled counterstained neurons including both medium elongated and small round cell bodies (Yelnik et al., 1991). As AChE histochemistry has been demonstrated to closely correspond to ChAT immunoreactivity in the striatum of macaques (Mesulam et al., 1984), it can be concluded that AChEpositive neurons are the cholinergic neurons of the striatum of monkeys. As they have the largest cell bodies of the striatum, it can be concluded that they correspond to spidery neurons. This conclusion, suggested by Smith and Parent (1984), is in agreement with the electron microscope analysis of DiFiglia (1987). Cholinergic neurons, or spidery neurons, of the primate striatum are thus characterized by a voluminous and globular cell body which is the largest one of all other striatal neurons in primates. They correspond to the larger achromatic cells of cytoarchitectonic preparations. They have a large number (12) of very thick (up to 10 pm) dendritic stems. These stems branch rapidly into numerous branches which become thinner and thinner at each bifurcation. Distal branches are very numerous (129) and most of them are thin and varicose. They often curve back toward the soma. As a whole, the dendritic arborization of spidery neurons is highly recognizable on its high density and the curved and varicose aspect of its distal processes (Fig. 2 0 .
The dendritic morphology of striatal neurons in rats Golgi-impregnated brains of adult rats were also examined for this study. Rats were perfused through the aorta with 1% paraformaldehyde and 1% glutaraldehyde in 0.1 M phosphate buffer. Brains were processed by the Golgi method according to the modification of Van der Loos (1956). Neurons were drawn through the camera lucida ( x 100 immersion objective) and analyzed qualitatively. Some of
them, which could be reconstructed from serial sections, were analyzed quantitatively according to our computer-aided method. The overall distribution of striatal neurons in the rat is the same as in primates, i.e., comprising about 96% spiny neurons and a few neurons having a larger cell body than spiny neurons. Spiny neurons of the rat striatum (Fig. 1C) have lower topological and metrical parameters than spiny neurons of monkey and human. The longest dendrite in rats was 220 pm (Fig. lC), 200- 250 pm (Kitai et al., 1979), 280 pm (Wilson and Groves, 1980), 200 - 220 pm (Dimova et al., 1980), 220 pm (Bishop et al., 1982), which is smaller than the 275 pm measured in primates (Yelnik et al., 1991). The mean length of dendritic segments (67 pm in Fig. 1C) is also smaller than in monkeys (93 pm) and humans (75 pm), and so is the dendritic formula: 6 - 30 for the rat, 5 - 35 for the monkey and 6-42 for the human. Conversely, the dimensions of dendritic arborizations were 360 x 230 x 180 pm in Fig. 1C and 260 - 530 x 240-425 x 260- 350 in Preston et al. (1980), which is similar t o the 430 x 330 x 230 pm that we measured in primates (Yelnik et al., 1991). Spine distribution (5 per 10 pm in Fig. 1C) was intermediate. Two different types of large neurons were identifiable in our material. Neurons of the first type (Fig. 3B) had a fusiform cell body and 3 - 4 dendritic trees with long, smooth and sparsely branched dendrites (9 - 11 dendritic tips). They correspond to the large type I of Chang et al. (1982) and Chang and Kitai (1982) and resemble theleptodendritic neurons of primates. However, their dendrites were definitely thinner and shorter (400 pm for the longest one) than their primate counterpart (compare Figs. 2Band 3B). This observation is consistent with previous measurements in the rat: up to 150 pm (Takagi et al., 1984), 250 pm (Bolam et al., 1984a), 350 pm (Rafols et al., 1989), 400 pm (Chang et al., 1982) and 450 pm (Bishop et al., 1982). As the longest dendrite was 1270 pm in primates (Yelnik et al., 1991), this could suggest that large neurons of rats constitute a particular type. However, as they have a low dendritic formula and relatively long dendrites, they fit with the criteria of Ramon-Moliner
30
A
B
C
4-1
100pm
Fig. 3. Three neuronal types of the rat striatum. Golgi method. Same magnification as in Fig. 2. A . Spiny neuron with a small cell body (135 pm2). B . Two large neurons (497 and 392 pm’) with elongated cell bodies and sparsely ramified dendrites. Note that the cell bodieq are smaller and the dendrites far shorter than for leptodendritic neurons of primates (Fig. 2B). C . Two large neurons (471 and 430 pm’) with round cell bodies and more numerous, varicose and tortuous dendrites. Note that these neurons strongly differ from the spidery neurons of primates shown in Fig. 2C.
(1969) and are likely to represent the leptodendritic neurons of the rat striatum. The second type of large neurons that we observed (Fig. 3C) had a round or polygonal cell body with 4 - 6 dendritic trees, As a whole the dendritic arbori-
zation had 21 - 28 dendritic tips. The mean length of segments was 40 - 60 pm, the longest dendrite was 200 - 250 pm, the total dendritic length 1400 2500 pm. This type is similar to the large type I1 of Chang et al. (1982) and Chang and Kitai (1982). Ac-
31
cording to Bolam et al. (1984a), this type would be similar to the type IV of Braak and Braak (1982) in humans and the aspiny type I1 of DiFiglia et al. (1976) in monkeys, i.e., our spidery neurons. In our opinion, however, this comparison does not hold true because of significant differences, both qualitative (compare Figs. 2 C and 3C) and quantitative: 21 -28 tips vs. 129 in primates, a total dendritic length of 1400 - 2500 Fm vs. 23400 pm in primates. This strongly suggests that there are no spidery neurons in the rat striatum, which raises the question of the morphology of the rat cholinergic neurons.
Characterization of cholinergic neurons in the rat striatum The first studies of cholinergic neurons in nonprimate species were based on AChE histochemistry (Lynch et al., 1972; Butcher and Bilezikjian, 1975; Butcher and Hodge, 1976; Henderson, 1981; Armstrong et al., 1983) and concluded that two populations of neurons, one with medium-sized cell bodies, the other with large fusiform or multipolar cell bodies, were AChE-positive. Later, however, Eckenstein and Sofroniew (1983), Levey et al. (1983) and Phelps et al. (1985) using immunohistochemical detection of ChAT concluded that the apparent heterogeneity of ChAT-positive cell bodies might be explained by different orientations of elongated cell bodies and that cholinergic neurons constituted a single population of large neurons. Moreover, Bolam et al. (1984b) demonstrated, by combining Golgi impregnation, electron microscopy, AChE histochemistry and ChAT immunohistochemistry, that only one out of three populations of AChEpositive neurons (Bolam et al., 1984a) were immunoreactive to ChAT. For Bolam et al. (1984b), cholinergic neurons have a large triangular or multipolar cell body, “long infrequently branching dendrites”, and correlate with the giant neurons of Kemp and Powell (1971), the type V of Dimova et al. (1980), the giant neurons of Danner and Pfister (1981) and the type I large of Chang et al. (1982). These different types correspond to the leptodendritic neuron of Fig. 3B.
Bolam et al. (1984b) specify that cholinergic neurons are not “the same as the large neurons with a ‘spidery’ appearance and ‘swirling’ dendrites”, i.e., their type 2 which comprises the type I1 large of Chang et al. (1982) and their large version type 3 AChE-positive (Bolam et al., 1984a). Phelps et al. (1985) described ChAT-immunoreactive neurons as having large oval or multipolar somata and “three to four primary dendrites that branch and extend long distance”. Such a description is also that of a leptodendritic neuron. Kubota et al. (1987) described ChAT-immunoreactive neurons as having a large, oval, spindle, triangular or multipolar cell body, 3-5 dendritic trees emerging from the poles of the soma, and long and sparsely branched dendrites. They correlated these cholinergic neurons with the giant neurons of Kemp and Powell (1971) and the type I large of Chang et al. (1982), thus, also with leptodendritic neurons.
Conclusions Finally it appears from our results and from the data of the literature that: (1) cholinergic neurons of the striatum have the largest cell bodies of this cerebral region in both primate and non-primate species; (2) those striatal neurons having the largest cell body in the striatum are leptodendritic neurons in nonprimate species whereas they are spidery neurons in monkeys and man; (3) cholinergic neurons of nonprimate species are leptodendritic neurons whereas they are spidery neurons in primates; and (4) spidery neurons do not exist in non-primate species. The morphological difference between striatal cholinergic neurons of the rat and primate is considerable and raises several questions. One concerns the differentiation of the neuronal species of the striatum through phylogenesis. A first important point is that the dimensions of the dendritic arborizations of spiny neurons do not change from rats to humans. This implies that the grain of the dendritic lattice does not change (Yelnik et al., 1991), which is likely to be a fundamental feature of the striatal organization. Conversely, the present results demonstrate that leptodendritic neurons are subject to a
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considerable increase of dendritic length from rats to primates. Such an increase was also observed for the leptodendritic neurons of the globus pallidus (700 pm in rats, 1200 pm in primates; Yelnik et al., 1984) and substantia nigra pars reticulata (900 pm in rats, 1100 pm in macaques, 1600 pm in humans; Yelnik et al., 1987) but it does not represent a general rule of phylogenesis. For example, subthalamic neurons, as well as spiny neurons, remain unchanged from rats and cats to monkey and human (Yelnik and Percheron, 1979; Hammond and Yelnik, 1983). In rats, cholinergic neurons of the striatum are of the same morphological type as the cholinergic neurons of the nucleus basalis of Meynert and the peripallidal laminae (Ingham et al., 1985)and even of other brain regions (Woolf, 1991). Thus, in rats, there is a correlation between morphology and chemical content. In primates, conversely, cholinergic neurons of the striatum are definitely different from the cholinergic neurons of the basal forebrain. They have highly specific morphological features which are not observed in other regions of the nervous system. Pasik et al. (1991) have recently raised the question of the axonal nature of the cholinergic neurons of the striatum. As cholinergic neurons in rats are leptodendritic neurons and as leptodendritic neurons in primates (their large spiny 11) are projection neurons, they deduce that cholinergic neurons of rats should be projection neurons. However, the interneuronal nature of cholinergic neurons has been demonstrated by fluorescent tracer experiments (Woolf and Butcher, 1981). In addition, leptodendritic neurons projecting to the substantia nigra have been shown in only the ventral part of the rat striatum (Bolam et al., 1981), which suggests that the remaining leptodendritic neurons of the rat striatum, i.e., the cholinergic neurons, could be interneurons. An abundant literature has been devoted to the functional role of cholinergic neurons but it has never taken into account interspecific morphological differences. In Woolf's (1991) review for example, cholinergic neurons are considered as a global system contrasting with the modularity of sensory systems. While this is likely to be true for the
cholinergic neurons of the rat striatum, it does not seem to be applicable to spidery neurons, the cholinergic neurons of the primate striatum, which have relatively small but very dense arborizations. As they represent only 1% of striatal neurons, they constitute a dendritic lattice which is characterized by zones of high dendritic density alternating with zones which are almost free of cholinergic processes (see fig. 12 in Yelnik et al., 1991). This implies that groups of spiny neurons could be strongly controlled by the activity of cholinergic neurons while others could not. Therefore, the anatomical organization of the cholinergic system of the striatum is submitted to a considerable transformation from rats to humans. An evolution from a globally organized system to a fine-grain modular system is certainly a highly significant feature of the primate striatum, which should be carefully considered in the analysis of the function of the striatum.
References Anderson, A.D. (1954) Alcoholic thionin counterstaining following Golgi dichromate-silver impregnation. Stain Techno/., 29: 179- 184. Armstrong, D.M., Saper, C.B., Levey, A.I., Wainer, B.H. and Terry, R.D. (1983) Distribution of cholinergic neurons in rat brain: demonstrated by the immunocytochemical localization of choline acetyltransferase. J . Comp. Neurol., 216: 53 - 68. Bishop, G.A., Chang, H.T. and Kitai, S.T. (1982) Morphological and physiological properties of neostriatal neurons: an intracellular horseradish peroxidase study in the rat. Neuroscience, 7: 179- 191. Bolam, J.P., Somogyi, P., Totterdell, S. and Smith, A.D. (1981) A second type of striatonigral neuron: a comparison between retrogradely labelled and Golgi-stained neurons at the light and electron microscopic levels. Neuroscience, 6 : 2141 - 21 57. Bolam, J.P., Ingham, C.A. and Smith, A.D. (1984a) The section-Golgi-impregnation procedure, 3. Combination of Golgi-impregnation with enzyme histochemistry and electron microscopy to characterize acetylcholinesterase-containing neurons in the rat striatum. Neuroscience, 12: 687 - 709. Bolam, J.P., Wainer, B.H. and Smith, A.D. (1984b) Characterization of cholinergic neurons in the rat neostriatum. A combination of choline acetyltransferase immunocytochemistry, Golgi impregnation and electron microscopy. Neuroscience. 12: 71 1 - 718. Braak, H. and Braak, E. (1982) Neuronal types in the striatum of man. Cell Tissue Res., 227: 319-342. Butcher, L.L. and Bilezikjian, L. (1975) Acetylcholinesterase-
33 containing neurons in the neostriatum and substantia nigra revealed after punctate intracerebral injection of diisopropylfluorophosphate. Eur. J . Phurrnacol., 34: 115 - 125. Butcher, L.L. and Hodge, G.K. (1976)Postnatal development of acetylcholinesterase in the caudate-putamen nucleus and substantia nigra of rats. Bruin Rex, 106:223 - 240. Chang, H.T. and Kitai, S.T. (1982)Large neostriatal neurons in the rat: an electron microscopic study of gold-toned Golgistained cells. Bruin Res. Bull., 8:631 -643. Chang, H.T., Wilson, C.J. andKitai, S.T. (1982)A Golgi study of the rat neostriatal neurons: light microscopic analysis. J. Cornp. Neurol., 208: 107- 126. Danner, H. and Pfister, C. (1979)Untersuchungen zur Struktur der Neostriatum des Ratte. J. Hirnforsch., 20: 285 -301. Danner, H. and Pfister, C. (1981)4 Spine-lose Neurontypen im Neostriatum der Ratte. 1.Hirnforsch., 22: 465 - 477. Davenport, H.A. and Combs, C.M. (1954)Golgi’s dichromatesivler method. 3. Chromating fluids. Stain Techno/., 29: 165 - 173. DiFiglia, M. (1987)Synaptic organization of cholinergic neurons in themonkey neostriatum. J. Cornp. Neurol., 255:245 - 258. DiFiglia, M., Pasik, P . and Pasik, T. (1976)A Golgi study of neuronal types in the neostriatum of monkeys. Brain Res., 114:245 - 256. Dimova, R., Vuillet, J. and Seite, R. (1980)Study of the rat neostriatum using a combined Golgi electron microscope technique and serial sections. Neuroscience, 5: 1581 - 1596. Eckenstein, F. and Sofroniew, M.V. (1983)Identification of central cholinergic neurons containing both choline acetyltransferase and acetylcholinesterase and of central neurons containing only acetylcholinesterase. J. Neurosci., 3:2286 - 2291. Fox, C.A., Andrade, A.N., Hillman, D.E. and Schwyn, R.C. (1971a)The spiny neurons in the primate striatum: a Golgi and electron microscopic study. J. Hirnforsch., 13: 181 - 201. Fox, C.A., Andrade, A.N., Schwyn, R.C. and Rafols, J.A. (1971b) The aspiny neurons and the glia in the primate striatum: a Golgi and electron microscopic study. J. Hirnforsch., 13: 341 - 362. Graveland, G.A., Williams, R.S. and DiFiglia, M. (1985) A Golgi study of the human neostriatum: neurons and afferent fibers. J. Cornp. Neurol., 34: 317 - 333. Hammond, C. and Yelnik, J. (1983)Intracellular labelling of rat subthalamic neurones with horseradish peroxidase: computer analysis of dendrites and characterization of axon arborization. Neuroscience, 8: 781 - 790. Henderson, 2. (1981) Ultrastructure and acetylcholinesterase content of neurons forming connections between the striatum and substantianigraof rat. J. Cornp. Neurol., 197: 185 - 196. Ingham, C.A., Bolam, J.P., Wainer, B.H. and Smith, A.D. (1985)A correlated light and electron microscopic study of identified cholinergic basal forebrain neurons that project to the cortex in the rat. J. Comp. Neurol., 239: 176- 192. Kemp, J.M. and Powell, T.P.S. (1971)The structure of the caudate nucleus of the cat: light and electron microscopy. Phil.
Trans. R . SOC.Lond. (Biol.), 262: 383 - 401. Kimura,H.,McGeer,P,L.,Peng, J.H.andMcGeer,E.G.(1981) The central cholinergic system studied by choline acetyltransferase immunohistochemistry in the cat. J. Cornp. Neurol., 200: 151-201. Kitai, S.T., Preston, R.J., Bishop,G.A. andKocsis, J.D. (1979) Striatal projection neurons: morphological and electrophysiological studies. In: L.J. Pokier, T.L. Sourkes and P.J. Bedard (Eds.), Advances in Neurology, Vol. 24, Raven Press, New York, pp. 45-51. Kubota, Y., Inagaki, S., Shimada, S., Kito, S., Eckenstein, F. and Tohyama, M. (1987) Neostriatal cholinergic neurons receive direct synaptic inputs from dopaminergic axons. Bruin Res., 413: 179- 184. Leontovich, T.A. (1954)Fine structure of subcortical ganglia (in Russian). Z.Nevropat. Psikh., 54: 168 - 178. Leontovich, T.A. (1983)Spatial arrangement of tissue elements of nucleus caudatus in dog (in Russian). Neurophysiology, IS: 474 - 484. Levey, A.I., Wainer, B.H., Mufson, E. J . and Mesulam, M.-M. (1983) Co-localization of acetylcholinesterase and choline acetyltransferase in the rat cerebrum. Neuroscience, 9:9 - 22. Lu, E.J. and Brown, W.J. (1977) The developing caudate nucleus in the euthyroid and hypothyroid rat. J . Cornp. Neuro/., 171: 261 -284. Lynch, G.S., Lucas, P.A. and Deadwyler, S.A. (1972) The demonstration of acetylcholinesterase containing neurones within the caudate nucleus of the rat. Brain Res., 45: 617-621. Mesulam, M.-M., Mufson, E.J., Levey, A.1. and Wainer, B.H. (1984)Atlas of cholinergic neurons in the forebrain and upper brainstem of the macaque based on monoclonal choline acetyltransferase immunohistochemistry and acetylcholinesterase histochemistry. Neuroscience, 12:669 - 686. Nagai, T., Pearson, T., Peng, F., McGeer, E.G. and McGeer, P.L. (1983)Immunohistochemical staining of the human forebrain with monoclonal antibody to human choline acetyltransferase. Brain Rex, 265: 300 - 306. Parent, A. and O’Reilly-Fromentin, J. (1982)Distribution and morphological characteristics of acetylcholinesterase-containing neurons in the basal forebrain of the cat. Brain Res. Bull., 8: 183 - 196. Pasik, P., Pasik, T. and DiFiglia, M. (1979)The internal organization of the neostriatum of mammals. In: 1. Divac and R.G.E. Oberg (Eds.), The Neostriaturn, Pergamon Press, New York, pp. 5 - 36. Pasik, P., Pasik, T. and Holstein, G.R. (1991)The ultrastructural chemoanatomy of the basal ganglia: 1984-1989.I. The neostriatum. In: G. Bernardi et al. (Eds.), The Basal Ganglia III, Plenum, New York, pp. 187- 197. Percheron, G. (1979)Quantitative analysis of dendritic branching. 1. Simple formulae for the quantitative analysis of dendritic branching. Neurosci. Lett., 14: 287 - 293. Percheron, G. (1982)Principles and methods of the graph theo-
34
retical analysis of natural binary arborescences. J. Theor. Biol., 99: 509 - 552. Phelps, P.E., Houser, C.R. and Vaughn, J.E. (1985) Immunocytochemical localization of choline acetyltransferase within the rat neostriatum: a correlated light and electron microscopic study of cholinergic neurons and synapses. J . Comp. Neurol., 238: 286- 307. Poirier, L.J., Parent, A , , Marchand, R. and Butcher, L.L. (1977) Morphological characteristics of the acetylcholinesterase-containing neurons in the CNS of DFPtreated monkeys. Part 1 . Extrapyramidal and related structures. J. Neurol. Sci., 31: 181 - 198. Preston, R.J., Bishop, G.A. and Kitai, S.T. (1980) Medium spiny neuron projection from the rat neostriatum: an intracelMar horseradish peroxidase study. Brain Res., 183: 253 - 263. Pullen, A.H. (1984) A structured program in Basic for cell morphometry: its application to the spinal motoneurone. J. Neurosci. Methods, 12: 1 5 5 - 178. Rafols, J.A., Cheng, H.W. and McNeill, T.H. (1989) Golgi study of the mouse striatum: age-related dendritic changes in different neuronal populations. J. Comp. Neurol., 279: 2 12 - 227. Ramon-Moliner, E. (1969) The leptodendritic neuron: its distribution and significance. Ann. N. Y. Acad. Sci., 167: 65 - 70. Satoh, K. and Fibiger, H.C. (1985) Distribution of central cholinergic neurons in the baboon (Pupio pupio). I. General morphology. J . Comp. Neurol., 236: 197-214. Satoh, K., Staines, W.A., Atmadja, S. and Fibiger, H.C. (1983) Ultrastructural observations of the cholinergic neuron in the rat striatum as identified by acetylcholinesterase pharmacohistochemistry. Neuroscience, 10: 1121 - 1136. Smith, Y. and Parent, A. (1984) Distribution of acetylcholinesterase-containing neurons in the basal forebrain and upper brainstem of the squirrel monkey (Suimirisciureus). Bruin Res. Bull., 12: 95 - 104. Takagi,H.,Somogyi,P.andSmith,A.D. (1984)Aspinyneurons and their local axons in the neostriatum of the rat: a correlated light and electron microscopic study of Golgi-impregnated material. J. Neurocytol., 13: 239 - 265.
Tanaka Jr., D. (1980) Development of spiny and aspiny neurons in the caudate nucleus of the dog during the first postnatal month. J. Comp. Neurol., 192: 241-263. Van der Loos, H. (1956) Une combinaison de deux vieilles methodes histologiques pour le systeme nerveux central. Mschr. Psychiatr. Neurol., 132: 330- 334. Wilson, C.J. andGroves, P.M. (1980) Fine structure and synaptic connections of the common spiny neuron of the rat neostriatum: a study employing intracellular injection of horseradish peroxidase. J. Comp. Neurol., 194: 599-616. Woolf, N. J . (1991) Cholinergic systems in mammalian brain and spinal cord. Prog. Neurobiol., 37: 475 - 524. Woolf, N.J. and Butcher, L.L. (1981)Cholinergic neurons in the caudate-putamen complex proper are intrinsically organized: a combined Evans Blue and acetylcholinesterase analysis. Brain Res. Bull., 7: 487 - 507. Yelnik, J . and Percheron, G. (1979) Subthalamic neurons in primates: a quantitative and comparative analysis. Neuroscience, 4: 1117-1143. Yelnik, J., Percheron, G . , Perbos, J. and Francois, C. (1981) A computer-aided method for the quantitative analysis of dendritic arborizations reconstructed from serial sections. J . Neurosci. Methods, 4: 347 - 364. Yelnik, J., Percheron, G., Francois, C. and Burnod, Y . (1983) Principal component analysis: a suitable method for the threedimensional study of the shape, dimensions and orientation of dendritic arborizations. J. Neurosci. Methods, 9: 115 - 125. Yelnik, J ., Percheron, G . and Francois, C. (1984) A Golgi analysis of the primate globus pallidus. 11. Quantitative morphology and spatial orientation of dendritic arborizations. J. Comp. Neurol., 221: 200-213. Yelnik, J . , Francois, C., Percheron, G . and Heyner, S. (1987) Golgi study of the primate substantia nigra I. Quantitative morphology and typology of nigral neurons. J. Comp. Neurol., 265: 455 - 472. Yelnik, J., Francois, C., Percheron, G. and Tande, D. (1991) Morphological taxonomy of the neurons of the primate striaturn. J. Comp. Neurol., 313: 273 - 294.