Peptidergic Neurons in the Basal Forebrain Magnocellular Complex of the Rhesus Monkey

Abstract

The basal forebrain magnocellular complex of primates is defined by the presence of large, hyperchromic, usually cholinergic neurons in the nucleus basalis of Meynert and nucleus of the diagonal band of Broca. Because there is growing evidence for noncholinergic neuronal elements in the basal forebrain complex, five neuropeptides and the enzyme choline acetyltransferase were studied immunocytochemically in this region of rhesus monkeys. Galaninlike immunoreactivity coexists with choline-acetyltransferase- like immunoreactivity in most large neurons and in some smaller neurons of the primate nucleus basalis and nucleus of the diagonal band. Four other peptides show immunoreactivity in more limited regions of the basal forebrain complex, usually in separate smaller, noncholinergic neurons. Numerous small, somatostatinlike-immunoreactive neurons occupy primarily anterior and intermediate segments of the nucleus basalis, especially laterally and ventrally. Somewhat fewer, small neuropeptide Y-like-immunoreactive somata are found in the same regions. Neurons that show neurotensinlike immunoreactivity are slightly larger than cells that contain immunoreactivity for somatostatin or neuropeptide Y, but these neurons also occur mainly in anterior and intermediate parts of the nucleus basalis. Overall, the usually small, leucine-enkephalin-like-immunoreactive neurons are infrequent in the basal forebrain complex and are most abundant in the rostra1 intermediate nucleus basalis. Thus, neurons that appear to contain somatostatin, neuropeptide Y, neurotensin, or enkephalin mingle with cholinergic/galaninergic neurons only in some subdivisions of the nucleus basalis/nucleus of the diagonal band, and their distributions suggest that some of these small neurons could be associated with structures that overlap with cholinergic neurons of the labyrinthine basal forebrain magnocellular complex. We also have found light microscopic evidence for innervation of basal forebrain cholinergic neurons by boutons that contain galanin-, somatostatin-, neuropeptide Y-, neurotensin-, or enkephalin-like immunoreactivity. The origins and functions of these putative synapses remain to be determined.

INTRODUCTION

Large neurons of the nucleus basalis of Meynert (nbM) and nucleus of the diagonal band of Broca (ndbB) are the major source of cholinergic innervation of the amygdala, hippocampus, and the neocortex in primate forebrain (Mesulam et al., ‘83, ‘86; Lehmann et al., ‘84; Walker et al., ‘85, ‘86; Struble et al., ‘86). In rhesus monkeys, these neurons form a continuum that extends from the dorsomedial septum into the rostra1 basal forebrain and then into the substantia innominata, where cells of the nbM border ventral and lateral aspects of inner and outer segments of the globus pallidus. With the advent of antibodies to the specific cholinergic marker choline acetyltransferase (ChAT), it has been firmly established that most large neurons in the basal forebrain magnocellular complex (BFMC) are cholinergic (Hedreen et al., ‘83; Mesulam et al., ‘83, ‘84; Satoh and Fibiger, ‘85a,b). Many large neurons of the BFMC also show immunoreactivity for galanin, a widely distributed 29-amino-acid neuropeptide (Rökaeus et al., ‘84; Melander et al., ‘85, ‘86c,d; Skofitsch and Jacobowitz, ‘85). Galanin coexists with ChAT in some neurons of the medial septum and ndbB in rats but is not found in neurons of the rat counterpart to the nbM, the nucleus basalis magnocellularis (Melander et al., ‘85). In the owl monkey, a nocturnal New World primate, galanin-containing cells are distributed more widely than in the rat, being found in the nbM as well as in the ndbB (Melander and Staines, ‘86). In the latter study, galanin was found to coexist with acetylcholinesterase in neurons of the BFMC, but the comparative localization of galanin and ChAT has not been established in primates.

Intermixed among large cells of the primate BFMC is a population of smaller neurons (Beccari, ‘11; Jones et al., ‘76; Hedreen et al., ‘84), some of which are not cholinergic (Mesulam et al., ‘83). In parts of the basal forebrain complex of primates, there is immunocytochemical evidence for the existence of fibers and/or putative terminals that contain enkephalin (Haber and Elde, ‘82a,b; Candy et al., ‘85; Haber and Watson, ‘85), neuropeptide Y (Smith et al., ‘85), neurotensin (Mai et al., ‘87), proopiomelanocortin peptides (Khachaturian et al., ‘84; Candy et al., ‘85), somatostatin (Candy et al., ‘85; Bennett-Clarke and Joseph, ‘86), substance P, cholecystokinin octapeptide, vasoactive intestinal polypeptide, and oxytocin (Candy et al., ‘85). However, except for somatostatinergic neurons (Bennett-Clarke and Joseph, ‘86), the transmitter identity of noncholinergic neurons is not known, and there is no information about the topographic relationship of noncholinergic to cholinergic neurons.

In the present study, we mapped the distribution of cholinergic (ChAT-like immunoreactive) and peptidergic (galanin-, somatostatin-, neuropeptide Y-, enkephalin-, or neurotensinlike immunoreactive) neurons in the BFMC of the rhesus monkey. The results indicate that galanin coexists with ChAT in large neurons throughout the BFMC of this Old World primate. Nerve cells immunoreactive for somatostatin, neuropeptide Y, enkephalin, or neurotensin mingle with cholinergic neurons in limited areas of the basal forebrain complex, but these cells are relatively small and infrequent and appear not to be cholinergic. Each of the five peptides studied appears in some putative nerve terminals that abut cholinergic neurons.

MATERIALS AND METHODS

Three male rhesus monkeys (Macaca mulatta), approximately 10, 15, and 16 years of age, were used. Animal 84–28 (15 years of age) was perfused under deep sodium pentobarbital anesthesia with sodium-phosphate-buffered 4% paraformaldehyde (pH 7.3) followed by phosphate-buffered 5% sucrose. The brain was blocked, cryoprotected in phosphate-buffered 25% sucrose (4°C) for 24 hours, frozen on dry ice, and stored (−80°C). Animal 86–7 (16 years of age) was anesthetized with sodium pentobarbital and given an injection of 1000 μg colchicine (5 μg/μl of sterile buffered saline, 200 μl total volume) into the left lateral ventricle under stereotaxic guidance. After surgery, the animal was intubated with a cuffed endotracheal tube, and anesthesia was maintained with a mixture of halothane and oxygen by the use of intermittent positive-pressure ventilation. After 24 hours, the monkey was perfused with sodium-phosphate-buffered 4% paraformaldehyde (pH 7.3) and then with the same fixative with 5% sucrose. The brain was postfixed in this latter solution for 90 minutes, blocked, cryoprotected (4°C) for 22 hours, and frozen in isopentane (−30°C) prior to storage (−80°C). Animal 87–9 (10 years of age) was anesthetized with sodium pentobarbital and given an injection of 1000 μg colchicine (4 μg/μl sterile buffered saline, 250 μl total volume) into the fourth ventricle. The animal was then kept under anesthesia, as described for animal 86–7, for 34 hours. At this time, the monkey was perfused with cold, acetate-buffered paraformaldehyde (pH 6.5, 20 minutes) and then with cold, borate-buffered paraformaldehyde (pH 9.5, 20 minutes) (Berod et al., ‘81). The brain was blocked stereotaxically, cryoprotected in borate-buffered glycerol (pH 9.5), frozen on dry ice, and stored (−80°C) until analysis.

Blocks from one or both hemispheres were cut (40 μm) on a freezing-sliding microtome for immunocytochemical analysis by using antibodies against ChAT (rat/mouse monoclonal AB-8, at a concentration of 1:250, antibody courtesy of Dr. Bruce H. Wainer; see Levey et al., ‘83), galanin (rabbit polyclonal, 1: 1000, Peninsula Laboratories Inc., Belmont, California), somatostatin (rabbit polyclonal, 1:1000, Immunonuclear Corp., Stillwater, Minnesota), neuropeptide Y (rabbit polyclonal, 1:2000, Cambridge Research Biochemicals, Ltd., Cambridge, United Kingdom), leucine-enkephalin (leu-enkephalin; rabbit polyclonal, 1:1,000, Immunonuclear Corp.), or neurotensin (rabbit polyclonal, 1:2,000, Immunonuclear Corp.). For single immunolabeling, sections were processed with the peroxidase-antiperoxidase method of Sternberger (‘79) with diaminobenzidine as chromogen. This peroxidase-antiperoxidase method also was used in double-immunolabeling studies with diaminobenzidine (DAB) to stain one antigen golden brown and with benzidine dihydrochloride (BDHC) to stain a separate antigen granular blue in the same tissue section (Levey et al., ‘86). Control sections were incubated with antigen-blocked primary antisera or with nonimmune sera; these sections were negative. It should be noted that, although antibodies used in this study were directed against specific domains of the respective antigens, variability in immunocytochemical staining could result from the presence of antigenically similar domains on other molecules or from species differences in the molecular makeup of antigens.

Labeled neurons in selected sections were mapped with an X-Y plotter (DiLog Instruments, Tallahassee, Florida) attached to a Leitz microscope and photographed with a Zeiss photomicroscope. In animals 84–28 and 86–7, the distribution of peptidergic neurons was determined throughout most of the basal forebrain complex in relation to cholinergic neurons in nearby single-immunostained sections. Some of these sections were counterstained lightly with cresyl violet. In animal 86–7, selected sections through the rostral basal forebrain complex, where most peptidergic somata were seen, were double immunostained for both cholinergic and peptidergic markers. In animal 87–9, sections at regular intervals throughout the basal forebrain complex were double immunostained in this manner.

Choline-acetyltransferase- and galaninlike-immunoreactive neurons were counted in sections that represented different levels of the basal forebrain complex; only neurons with evident nuclei were included in the tally. Neurons were counted manually by systematic superimposition of an ocular grid over all parts of the basal forebrain complex within each section. In rostral and intermediate subdivisions of the basal forebrain complex of the two colchicine-treated monkeys, neuronal size was measured with a calibrated ocular scale; no correction was made for shrinkage during tissue processing because comparisons of cell size were made within tissues that were processed in essentially the same way.

RESULTS

Somata.

Choline acetyltransferase.

Choline-acetyltransferaselike-immunoreactive neurons are located throughout the basal forebrain complex (Fig. 1). These neurons are most numerous in the combined nbM and ndbB just rostral to and at the level of the decussation of the anterior commissure in coronal sections. Caudal to the commissural decussation, the number of cholinergic cells decreases gradually in the intermediate and then posterior nbM (Ch4i and Ch4p of Mesulam and colleagues, ‘83); ChAT-like-immunoreactive neurons in posterior nbM are confined to the increasingly restricted space between the optic tract and putamen, and in the laminae of the globus pallidus. Coronally sectioned cholinergic somata that were measured in anterior (Ch4a) and intermediate segments of the nbM vary somewhat in size and shape, with a mean maximum width of approximately 15 μm and a mean maximum length of approximately 28 μm. Occasionally, small ChAT-like-immunoreactive neurons may be encountered (Fig. 2).

Figure 1.

The distribution of neurons immunostained for ChAT (left column) and galanin (GAL, right column)-like immunoreactivity at four rostrocaudal levels of the BFMC of a rhesus monkey pretreated with colchicine. The two sections shown at each level are adjacent; each dot represents a single neuron from one tissue section.

Figure 2.

ChAT-like-immunoreactive small neuron (arrow) in the nbM. Nissl counterstain. Bar = 25 μm.

Galanin.

The distribution of galaninlike-immunoreactive neurons overlaps extensively with that of cholinergic neurons throughout the basal forebrain complex (Fig. 1). The number of galaninlike-immunoreactive cells in our material was always somewhat less than the number of ChAT-like-immunoreactive cells in adjacent sections, although this disparity may have been the result of differences in immunocytochemical staining characteristics of antigens. (Galaninlike immunoreactivity was relatively light and rarely extended very far into dendrites as compared to ChAT-like immunoreactivity). Galaninlike-immunoreactive cells measured in the anterior and intermediate nbM are essentially the same mean size as cholinergic neurons in adjacent sections (14 μm × 28 μm). Thus, most galaninlike-immunoreactive neurons are medium to large, but small neurons could be found. Double immunostaining of selected sections showed that most cholinergic neurons of the basal forebrain complex also show immunoreactivity for galanin (Fig. 3). Staining intensity for both antigens was most robust in the rostral basal forebrain complex and weakest in the posterior nbM. Some neurons also were seen that were immunoreactive only for ChAT or galanin, but the lack of double staining may have been due to limitations in the immunocytochemical method.

Figure 3.

Neurons in the nbM that are double immunostained for ChAT (granular blue BDHC) and galanin (homogeneous brown DAB)-like immunoreactivity. Bar = 25 μm.

Somatostatin.

In the rostralmost basal forebrain complex, a few small somatostatinlike-immunoreactive cells are located in the dorsal and lateral aspects of the septum as well as among neurons of the ndbB. Most such neurons are peripheral to the BFMC proper. Just rostral to, and at the level of, the decussation of the anterior commissure, small (mean size = 10 μm × 16 μm; Fig. 4A) somatostatinergic neurons within the limits of the nbM are located primarily in the lateral part (Ch4al) (Fig. 5). At rostral intermediate levels (rostral Ch4i), somatostatinergic neurons continue to occupy the basal forebrain complex (Fig. 6), but, more caudally, the number of cells diminishes, and few somatostatinergic cells are detectable in the posterior nbM (Ch4p). In internal and external laminae of the globus pallidus, it is difficult to determine whether small-to-medium-size peptidergic neurons belong to the nucleus basalis proper or to the basal ganglia. However, some somatostatinlike-immunoreactive neurons could be found, usually in the external pallidal lamina.

Figure 4.

Noncholinergic peptidergic somata in the nbM. Cholinergic neurons are stained with granular blue BDHC. Peptidergic neurons are stained with brown DAB. A: Somatostatinlike-immunoreactive neuron (arrow). B: Neuropeptide Y-like-immunoreactive neuron (arrow) and a possible terminal on a proximal dendrite of a cholinergic neuron (arrowhead). C: Neurotensinlike-immunoreactive neuron (arrow). D: Leu-enkephalin-like-immunoreactive neuron (arrow) and putative terminal on a proximal dendrite of a cholinergic neuron (arrowhead). Bar = 25 μm.

Figure 5.

Distributions of four types of noncholinergic peptide-containing neurons in a series of adjacent sections through the BFMC at the level of the decussation of the anterior commissure. The tissue was double immunostained to show neurons that are immunoreactive for ChAT (open circles) and either somatostatin-, neuropeptide Y-, neurotensin-, or leu-enkephalin-like immunoreactivity (solid dots). Each open circle represents one to three cholinergic neurons, and each solid dot represents one peptidergic neuron within a single section. Neurons clearly belonging to structures outside of the basal forebrain (e.g., to the preoptic area, amygdala, or striatum) are not indicated.

Figure 6.

Distributions of four types of noncholinergic peptide-containing neurons in a series of adjacent sections through the intermediate level of the nbM. The tissue has been double immunostained to show neurons that are stained for ChAT (open circles). and either somatostatin-, neuropeptide Y-, neurotensin-, or leu-enkephalin-like immunoreactivity (solid dots). Each open circle represents one to three cholinergic neurons and each solid dot represents one peptidergic neuron within a single section. Neurons clearly belonging to structures outside the BFMC (e.g., hypothalamus, amygdala, or striatum) are not indicated.

Neuropeptide Y.

Neurons containing neuropeptide Y are distributed similarly to somatostatinergic cells, although neuropeptide Y neurons are less numerous. A few neuropeptide Y-like-immunoreactive cells are located in the rostral septum, usually lateral to cholinergic cells of the vertical limb of the ndbB. Ventrally and caudally in the basal forebrain complex, somata that contain neuropeptide Y (Fig. 4B) are most abundant in the anterolateral nbM (Ch4al) and in the rostral intermediate nbM (Figs. 5, 6). These neurons are usually smaller (9 μm × 15 μm) than ChAT- or galaninlike immunoreactive cells in the nbM. Proceeding caudally, the number of neuropeptide Y-like-immunoreactive cells diminishes in the caudal part of the intermediate nbM (caudal Ch4i) and in the posterior nbM (Ch4p). A few neuropeptide Y-like-immunoreactive neurons could be seen in the external pallidal lamina.

Neurotensin.

Some neurotensinlike-immunoreactive neurons mix with cholinergic cells of the vertical limb of the ndbB, but most neurotensin cells in septum are dorsal or lateral to the ndbB. Ventrally and caudally, neurotensin- like-immunoreactive neurons are most numerous in the anterior nbM and tend to be distributed more evenly in the nbM than somatostatin- and neuropeptide Y-like-immunoreactive neurons (Figs. 5, 6). Although many neurotensinlike-immunoreactive cells in the intermediate nbM are ventral and/or lateral to cholinergic neurons, neurotensin neurons in the anterior and intermediate nbM mix with cholinergic somata in double-immunostained sections (Fig. 4C). Few neurotensinlike-immunoreactive cells were detected in the most posterior nbM (Ch4p), and none was seen in pallidal laminae. Neurotensinlike-immunoreactive neurons are somewhat larger (10 μm × 19 μm) than neuropeptide Y-, somatostatin-, and leucine-enkephalin-like-immunoreactive somata in the nbM.

Leucine-enkephalin.

Leucine-enkephalin-like-immunoreactive neurons are infrequent in the basal forebrain complex. Cells are scattered sparsely throughout the septum, occasionally intermixing with cholinergic neurons of the ndbB. At the level of the decussation of the anterior commissure, numerous leu-enkephalin-like-immunoreactive neurons are seen in the regions of the bed nucleus of the stria terminalis and the preoptic area, but only a few small neurons (9 μm × 16 μm; Fig. 4D) actually invade the basal forebrain complex proper (Fig. 5). In the rostral intermediate nbM (rostral Ch4i), leu-enkephalin-like-immunoreactive cells mingle with the more ventral population of cholinergic neurons of the nbM (Fig. 6). In the external pallidal lamina, some enkephalinlike-immunoreactive neurons could be found, and occasionally such cells coexist with cholinergic neurons in the posterior nbM (Ch4p).

Fibers and terminals

Immunoreactivity for galanin-, somatostatin-, neuropeptide Y-, neurotensin-, or leu-enkephalin-like peptides can be found in fibers that course through the substantia innominata and in axonal enlargements that abut cholinergic neurons of the nbM. Examples of putative synaptic terminals, in the form of boutons apposed to cholinergic neurons, can be seen in Figures 4B,D and 7. The origin of these elements cannot be discerned in these immunocytochemical preparations. Synaptic contacts were relatively infrequent on immunostained portions of cholinergic neurons. Because large somata and their proximal dendrites receive few synapses in the primate nbM (Walker et al., ‘83), it is likely that the majority of peptidergic synapses occur on distal, usually unstained, dendrites of cholinergic neurons.

Figure 7.

Double-immunostained section showing a cholinergic neuron (granular blue BDHC) in the nbM that is studded with putative nerve terminals (arrowhead) containing neuropeptide Y-like immunoreactivity (brown DAB). Neurons in the basal forebrain complex that show such a high density of axosomatic contacts are rare; however, infrequent axonal enlargements containing galanin-, somatostatin-, neurotensin-, or leu-enkephalin-like immunoreactivity also can be seen abutting cholinergic neurons. Bar = 25 μm.

DISCUSSION

From a cytological point of view, the basal forebrain complex of the primate is not a homogeneous structure. Although noncholinergic neurons intermingle with cholinergic neurons in all subdivisions of the complex, our data show that neurons that contain somatostatin-, neuropeptide Y-, leu-enkephalin-, or neurotensinlike immunoreactivity are concentrated most heavily in anterior and intermediate parts of the nbM. Occasionally, these smaller peptidergic neurons are found in the vertical limb of the ndbB, and relatively few, if any, reside in the posterior nbM or among cells of the nbM that extend into internal and external medullary laminae of the globus pallidus. There are many noncholinergic neurons in the region of the vertical limb of the ndbB, particularly in the dorsal septum, yet few of these cells are immunoreactive for somatostatin, neuropeptide Y, leu-enkephalin, or neurotensin. Although small neurons occasionally may be immunoreactive for ChAT (Fig. 2), no cholinergic neurons were shown conclusively to contain somatostatin-, neuropeptide Y-, leu-enkephalin-, or neurotensinlike immunoreactivity. The predominant distribution of small peptidergic neurons in the anterior nbM and rostra1 intermediate nbM suggests that some of these cells could actually belong to neighboring nuclei that invade the substantia innominata to some degree, particularly the lateral hypothalamus, preoptic area, and amygdala. For example, portions of central and medial amygdala of the rat penetrate the sublenticular substantia innominata (de Olmos et al., ‘85).

The existence of galanin in most, if not all, medium-to-large cholinergic neurons of the BFMC of rhesus monkeys confirms a previous study that shows galanin throughout this system in the owl monkey (Melander and Staines, ‘86), and indicates that this peptide could act as a cotransmitter with acetylcholine. A recent study (Wenk and Rökaeus, ‘88) has shown that sectioning the fimbria-fornix in rats lowers levels of immunoreactivity for galanin and ChAT to a similar degree in hippocampus. Furthermore, lesions of the fimbria or septum in rats diminish the number of hippocampal binding sites for galanin, suggesting that these sites may reside on cholinergic processes that project from the medial septum/ndbB to the hippocampus (Fisone et al., ‘87). Most high-affinity binding sites for galanin in rat hippocampus are in the ventral part, and galanin inhibits the evoked discharge of acetylcholine in ventral, but not in dorsal, hippocampus (Fisone et al., ‘87).

Galanin colocalizes with a variety of putative transmitters in many other parts of the central nervous system of the rat (Melander et al., ‘85, ‘86b; Kohler et al., ‘86). Specific receptor binding sites for galanin have been demonstrated in the rodent central nervous system (MeIander et al., ‘86a; Skofitsch et al., ‘86; Fisone et al., ‘87; Servin et al., ‘87), and a regulatory role for galanin in the hypothalamic-pituitary axis is suggested by the influence of galanin on the release of growth hormone (Bauer et al., ‘86; Ottlecz et al., ‘86; Murakami et al., ‘87), prolactin (Koshiyama et al., ‘87), and dopamine (Nordström et al., ‘87). Intrahypothalamic injection of galanin also stimulates feeding (Kyrkouli et al., ‘86).

The four small peptidergic cell types that we describe could be only a sample of the total population of noncholinergic somata in the basal forebrain complex. For example, we have preliminary data that show neurons containing substance P and vasoactive intestinal polypeptide in the substantia innominata. Although y-aminobutyric acid (GABA)-containing neurons have been demonstrated in the basal forebrain complex of rats (Brashear et al., ‘861, to date, we have not been able to stain unequivocally neurons in the basal forebrain complex with antibodies to the synthetic enzyme glutamic acid decarboxylase (GAD) or GABA in primates. However, recently, we have demonstrated neurons containing mRNA coding for GAD in the nbM of rhesus monkeys by in situ hybridization techniques (L.C. Walker, D.L. Price, and W.S. Young III, personal observations). Zaborszky and colleagues (‘86) found GAD-immunoreactive terminals on basal forebrain cholinergic neurons of the rat: we have seen light microscopic evidence for GABAergic innervation of these neurons in monkeys (L.C. Walker and C.A. Kitt, personal observations).

The possible coexistence of peptides with one another in small neurons of the basal forebrain complex also remains an issue for future investigation. Differences in the relative size, number, and distribution of neurons that contain neurotensin or leu-enkephalin suggest that regular coexistence of these substances with one another, with somatostatin, or with neuropeptide Y is unlikely. However, the size and distribution (but not the density) of cells that contain somatostatin or neuropeptide Y are similar in the BFMC, and these peptides have been shown to coexist within neurons of the cerebral cortex and hippocampus (Chan-Palay, ‘87; Kohler et al., ‘87). Our preliminary double-immunostaining experiments using antibodies directed against somatostatin and neuropeptide Y did not reveal single neurons in the BFMC that stained for both peptides.

The role of small neurons in the function of the BFMC is not yet known. Studies of retrogradely labeled neurons following injections of tracers into cerebral cortex of primates have shown that cortical innervation is accomplished primarily by large neurons (Kievit and Kuypers, ‘75; Jones et al., ‘76; Tigges et al., ‘82, ‘83; Mesulam et al., ‘83; Walker et al., ‘83, ‘85). Preliminary studies in which retrograde tracing was combined with immunocytochemistry confirm that few if any small peptidergic neurons project to distant telencephalic targets in monkey (V.E. Koliatsos, personal observation). As expected from the colocalization of galanin and ChAT, neurons that contain galanin project to these regions.

The presence of peptidergic axonal enlargements on cholinergic neurons of the BFMC suggests that small peptidergic cells and, perhaps, larger neurons that contain galanin and acetylcholine could act as local-circuit neurons to regulate cholinergic activity. For example, enkephalin is known to influence the function of basal forebrain cholinergic neurons in rats (Wenk, ‘84). However, it is possible that many peptidergic inputs arise from outside the BFMC. Numerous regions of brain may project to neurons of the BFMC in primates (Jones et al., ‘76; Russchen et al., ‘85; Irle and Markowitsch, ‘86); unfortunately, the neurochemical identity of most of these afferents is poorly understood. If small peptidergic neurons act as local-circuit neurons that regulate the function of nearby large cells, the relative scarcity of small cells in some parts of the basal forebrain complex suggests functional differences among the various subdivisions. In recent years, the BFMC has been defined by the cholinergic nature of large neurons that characterize the nbM and ndbB (Mesulam et al., ‘83). Our data in primates emphasize that acetylcholine may not be the only neuroactive substance in these large neurons because most, if not all, of them also appear to contain galanin. In addition, numerous smaller, mostly noncholinergic neurons, many of which contain neuropeptides, mingle with larger neurons, mainly in the anterior and intermediate parts of the basal forebrain complex. The existence of these elements underscores the anatomical complexity and possible functional heterogeneity of the BFMC in primates.

Abbreviations

BDHC

benzidine dihydrochloride

BFMC

basal forebrain magnocellular complex

BLA

nucleus basalis lateralis amygdalae

CA

commissura anterior

Cd

nucleus caudatus

ChAT

choline acetyltransferase

Ch4a

anterior nucleus basalis of Meynert

Ch4al

anterolateral nucleus basalis of Meynert

Ch4i

intermediate nucleus basalis of Meynert

Ch4p

posterior nucleus basalis of Meynert

CI

capsula interna

DAB

diaminobenzidine

F

fornix

GABA

γ-aminobutyric acid

GAD

glutamic acid decarboxylase

GAL

galanin

GP

globus pallidus

GPe

globus pallidus pars externa

GPi

globus pallidus pars interna

GU

gyrus uncinatus

Lenk

leucine-enkephalin

nbM

nucleus basalis of Meynert

ndbB

nucleus of the diagonal band of Broca

NPY

neuropeptide Y

N STh

nucleus subthalamicus

NT

neurotensin

Put

putamen

S

septum

SN

substantia nigra

SOM

somatostatin

Thal

thalamus

TO

tractus opticus

V Lat

ventriculus lateralis

III

ventriculus tertius