ESPERI>IENTAL
Efferent
KEC’ROLOGY
61, 185-197 (1978)
Connections The RAYMOND
of the Nucleus TROIAN~
Basal
Forebrain
in the Cat:
Accumbens AND ALLAN
SIEGEL 1
The efferent connections of the nucleus accumbens in the cat were studied with radioautographic methods. Injections of [“Hlleucine were placed throughout the extent of this structure in adult cats. The results revealed the presence of a topographical organization of the projections from the nucleus accumbens to the brain stem. Fibers arising from the dorsomedial sector of the nucleus accumbens project through the medial aspect of the medial forebrain bundle to the rostra1 end of the ventral tegmental area. Fibers arising from the ventromedial sector of the nucleus accumbens project to slightly more caudal and lateral parts of the ventral tegmentum. In contrast, fibers which arise from lateral parts of the nucleus accumbens project through the lateral aspect of the medial forebrain bundle and medial tip of the internal capsule to terminate primarily within the pars reticulata of the substantia nigra and central tegmental field. The data also suggest that fibers from this part of the nucleus accumbens probably terminate within the dorsolateral aspect of the substantia innominata and adjacent parts of the pallidum. These findings indicate that the nucleus accumbens is linked to both the limbic system and the basal ganglia.
INTRODUCTION The nucleus accumbens has heen the subject of a number of recent (5, 9, 11, 16, 18), electrophysiological (3, S), and behavioral (4, 13) experiments. In spite of the findings from these investigations its integration with any of the subsystems of the forebrain has remained an enigma. Early anatomic descriptions of this structure focussed on its anatomical
1 This project was supported by National Institutes of Health Research Grant NS 07941-08, awarded by the National Institute of Neurological and Communicative Disorders and Stroke. The authors wish to acknowledge Mrs. Jean Holland for typing this manuscript.
185 0014-4886/78/0611
-OlSS$OZ.OO/O
Copyright 0 1978 by Academic Press,Inc. All rights of rcprodurtion in any form reserved.
186
TROIANO
AND
SIEGEL
proximity and possible association with neostriatum (7). Several recent studies supported this position. Swanson and Cowan (18) claimed that fibers from the nucleus accumbens are distributed to the entopeduncular nucleus and substantia nigra, thus leading them to conclude that the nucleus accumbens constitutes part of the basal ganglia. Heimer and Wilson (9) introduced the term “ventral striatum” to underline the similarities between the nucleus accumbens and neostriatum in regard to both untrastructure and input-output relationships. In contrast, several other authors linked the nucleus accumbens with the limbic system after having identified efferent connections of that structure with the lateral hypothalamus and midbrain tegmentum (11, 16). The presence of afferent fiber connections to the nucleus accumbens from the tegmentum (5) as well as from the fornix system (3) also seems to suggest its classification with the limbic system. Koikegami et al. (11) noted that the nucleus accumbens can be divided into several subnuclear groups. These include two dorsal and two ventral nuclei. Within the dorsal group the cell cluster situated proximal to the caudate nucleus is referred to as the nucleats accztnabens caudati and the cell group proximal to the lateral septal nucleus is called the nbfcleus accun+ bens septi. Ventral to these groups are two additional and larger cell clusters identified as the nucleus alccuwtbens principalis pars lateralis and pars medialis, respectively [see (11. p. 157), Figs. 1 and 21. Parceling of the nucleus accumbens into these subdivisions, when considered collectively with the opposing findings which attempt to link this structure to either the basal ganglia or limbic system, clearly suggests that it may constitute a transitional zone that is related to both of these systems. Such a hypothesis would predict that the nucleus accumbens represents a continuum of cells whose axons project topographically to structures associated with both the basal ganglia and limbic system. In the study described below a test of this hypothesis was made by examining the efferent connections of the nucleus accumbens by the radioautographic technique in the cat. METHODS Twenty-four adult cats of either sex were used in this study. Surgery was carried out under Nembutal anesthesia (40 mg/kg, i.p.) utilizing clean conditions. Injections of radioisotope were placed stereotaxically with the use of a l-p1 Hamilton microsyringe mounted to the stereotaxic carrier. Injections were made with commercially prepared tritiated leucine (L- [4,5-3H]leucine, sp act 30 to 50 Ci/mmol, New England Nuclear, Boston, Massachusetts). The isotope was concentrated to 10 &i/PI by evaporation and reconstitution in sterile distilled water. A quantity of
NUCLEL-S
ACCUMRENS
187
PROJECTIONS
0.2 ~1 was injected into different parts of the nucleus accumbens and adjacent regions over a period of 40 min. A micromanipulator positioned directly on top of the syringe allowed for mliform advancement of the plunger which produced slow and controlled delivery of the isotope. Survival times varied from 1 to 2 days following placement of the injection. Cats were then killed with an overdose of I2’emhutal and perfused transcardially with solutions of 0.9? saline and 10% formalin. Brains were removed, stored in 10% formalin for several weeks, and then embedded in paraffin. They were serially sectioned at 8 to 10 pm, mounted on glass slides, and deparaffinized. Sections were coated with Kodak NTB or NTB-2 emulsion and exposed for approximately 2 to 3 weeks. Autoradiographs were developed in D-19, fixed in Ektaflo, and stained lightly with cresylechtviolet. RESULTS Injections were placed throughout the extent of the nucleus accumbens and included each of the cell divisions described by Koikegami et nl. ( 11) . The principal focus of each injection is summarized in Table 1 and illustrated in Fig. 1. Projectiorls
~YOIII
the
Dorsal
and Medial
Asficcts
of the Nucleus
rlccurrl-
hr~. In two cases injections were placed into the dorsomedial aspect of the nucleus accumbens. These injection sites correspond to the region designated as the rllrcl~~s accunlhem septi by Koikegami et al. (11). In case 1 the injection site included the dorsomedial aspect of the nucleus and the cells adjacent to the vertical limb of the diagonal band. In case 2 the injection \vas placed more rostrally and included cells in both the dorsomedial accumbens and adjoining infralimbic area. I,abeled axons were traced in a ventral and caudal direction through the preoptic area and into the medial forebrain bundle. Caudally descending labeled axons situated in the perifornical region could be followed through the hypothalamus to the rostra1 end of the ventral tegmental area. Labeled axon terminals were noted only in the medial aspect of this region at a level just behind the mammillary bodies (Fig. 2). Other fibers from the injection sites were traced through the stria medullaris to the medial half of the medioclorsal thalamic nucleus. However, hasetl on other studies described in the rat (141, it is likely that this component of the projection is more closely related to cell groups adjacent to the vertical limb of the diagonal band than to cells situated exclusively in the nucleus accumbensproper. Yrojcrtiorls
f~onr tltc Ve~ztroulrdial
A.s/vct
of the ATuclr~rs
Accul~~Dcns.
In five cases injections were placed in the ventrometlial aspect of the nucleus accmmben-the area identified by Koikegami ct al. ( 11) as the n//c-lrus c~ccu~~~Zuv~s priwipa1i.s pars rrwdialis. In three of the cases the
188
TROIANO
AND
TABLE Principal Case
1 2 3 4 5 6 7 8 9 10 11 12 13 14 1.5 16 17 18
Injection site within nucleus accumbens
Dorsomedial Dorsomedial Ventromedial Ventromedial Ventromedial Ventromedial Ventromedial Ventrolateral Ventrolateral Ventrolateral Ventrolateral Ventrolateral Ventrolateral Ventrolateral Dorsolateral Dorsolateral Dorsolateral Dorsolateral
foci of Labeled Ventral
tegmental area
Axon
SIEGEL
1 Terminals
in the Brain
Substantia Compact zone
++ ++ ++ i-f ++ ++ fS ++ 0 ? ++ 0 0 0 0 0 0 0
0 0 0 0 0 0 0 0 ? I ? 0 0 0 0 0 0 0
a + to + + +-increasing quantities of labeled axon labeled axon terminals; ?-uncertainty due to difficulty labeled axon terminals and fibers of passage.
nigra
Stem% Central
tegmental field
Reticular zone 0 0 0 0 0 0 0 0 + I ? + ? + + + ? ? terminals; O-no in discriminating
0 0 0 0 0 0 0 ++ + ? + +++ +++ + +-t-+ +-I-+ +++ +++ evidence of between
labeled cells were limited to the medial half of the nucleus accumbens principalis and in two other animals the injection sites also labeled a few cells in the adjacent olfactory tubercle. The projections to the brain stem from each of these injections were similar but differed somewhat from the pathway arising from the nucleus accumbens septi described above. Labeled axons were traced in a caudal direction from their origin in the nucleus accumbens principalis into the medial forebrain bundle at a position slightly lateral to the position occupied by fibers associated with the nucleus accumbens septi. Labeled axons could be further traced through the entire extent of the hypothalamus and ventral tegmentum to the level of the red nucleus. This fiber bundle appeared to terminate exclusively within the ventral tegmental area but in a position slightly caudal and lateral to the fibers arising from the nucleus accumbens septi. It did not appear from the injections involving either the dorsomedial or ventromedial aspects of the nucleus accumbens that any labeled fibers terminated directly within the substantia innominata. In fact, the labeled
NUCLEI5
ACCUMHENS
PKOJKTIONS
189
FIG. 1. Schematic representation of all injection sites of [“Hlleucine described in text. Diagrams from rostra1 (A) and caudal (B) nucleus accumbens. Abbreviations in this and the following figures : A-nucleus accumbens ; AC-anterior commissure ; C-caudate nucleus ; CP-cerebral peduncle ; CTF-central tegmental field ; ICinternal capsule ; IP-interpeduncular nucleus ; LH-lateral hypothalamus ; Mmammillary bodies ; MD-mediodorsal thalamic nucleus ; OT-olfactory tubercle ; PG-pontine gray ; PO-preoptic area ; K-red nucleus ; S-septal area ; VY-ventral tegmental area.
fibers of passage seemed to be contained entirely within the substance of the preoptic area. The possible presence of terminals in the preoptic region could not be excluded. Projections from the Vc~~t~olateral As/m-t of tlze AFltclrirs .-fcciflrlhols. In seven cats injections were placed into the ventrolateral part of the nucleus accumbens-the region proximal to the anterior commissure and identified as the alcclezts acczil1lben.r )mincipalis pavs lateralis by Koikegami ct al. (11). In two additional cases the injections appeared to extend beneath the ventral border of the nucleus accumbens and labeled cells associated with the olfactory tubercle and the rostra1 limit of the substantia innominata. The projections from each of these injections seemed to he substantially similar. The primary projection from this part of the nucleus accumbens. which is typical for other cases with similar injection sites, is shown in Fig. 3. From the injection site labeled axons were observed passing in a caudal direction through the substantia innominata in a position proximal to the globus pallidus to join the medial tip of the internal capsule and lateral part of the medial forebain bundle. It is entirely l)ossible, and indeed, probable that some of these labeled fillers terminate in the dorsolateral part of the substantia innominata-the region identified by
190
TROIANO
AND
SIEGEL
Heimer and Wilson (9) as the ventral pallidurn. However, as many labeled fibers of passage are present in this region as well, a definitive statement concerning whether or not any of this label was present over terminals cannot be made at this time. The larger percentage of axons could be followed passing caudally within the internal capsule and medial forebrain bundle to the level of the red nucleus, at which point they entered and terminated within the reticular division of the substantia nigra and overlying central tegmental area. The terminal distribution sites of these fibers were located more caudally and laterally within the midbrain than the projections arising from more medial parts of the nucleus accumbens described in Fig. 3. Projections from the Ctousolateral Part of the Nucletas Acctmbens. In four cases injections involved the dorsolateral aspect of the nucleus accumbens-the region identified as the nwleus accunzbens caudati by Koikegami ef al. (11). The projections from this part of the nucleus accumbens to the brain stem were similar to the pathway arising from the ventrolateral division of the nucleus accumbens (Fig. 4). The fibers from the dorsolateral part of the nucleus accumbens pass through the substantia innominata and proximal part of the globus pallidus prior to entering the medial part of the internal capsule and lateral aspect of the medial forebrain bundle. Again, it is quite likely that some of these fibers in fact
FIG. 2. Outline of an injection of [“Hlleucine placed into the dorsomedial the nucleus accumbens and distribution of label plotted from rostra1 (A) (D) levels (case 1).
aspect of to caudal
NUCLEUS
ACCUMRENS
PROJECTIONS
191
FIG. 3. Outline of au injection of [“H]leucine placed into the ventromedial aspect of the nucleus accumbens and distribution of label plotted from rostra1 (A) to caudal (E) levels (case 3). terminate within these regions. Because many descending fibers of passage are also clearly present here it is difficult, if not impossible, to conclude unequivocally that any of the labeled grains are present over axon terminals.
Nevertheless,
tributed
to the
central
tegmental
Although lateral
not
also
pathway more
arising caudally
into
group
the
anterior
of
in
the
with
which
In
projections
the
the
central
terminal
elsewhere
nigra. dorso-
the
the
disof the
of the
through
appeared
stem-at
were part
substantia
supply
general!
arising
the
passed to
brain
fibers lateral
injections
accmllbens
the
the
of
cases
direction
of in
division all
of fibers
dorsolateral
laterally
any
the
for
to be present.
from
terminated
reticular
dorsomedial
appeared and
juncture--than
a
accunihens. Control 1lljrctiou.r. placetl in neighborillg placed
and
majority
they
observed
a small in
overwhelming
where
field
field
substance
stem,
uniformly
accumbens,
tegmental
the
brain
to
central
gra)
fields
of the
he situated
midbrain-pontine
in the
nucleus
In a series of aninlals injections of [“Hlleucine were regions of the forebrain. In one case an injection part
of
the
lateral
septal
nucleus
resulted
in
a
192
TROIANO
AND
SIEGEL
massive distribution of labeled fibers to the preoptic area and anterior lateral hypothalamus via the medial forebrain bundle. This classical projection was described with degeneration techniques in the cat (15) and most recently with radioautography in the rat (14). In other cases [described in the following paper (20) ] injections involved the olfactory tubercle and adjacent portions of the substantia innominata. The projections from all but one of these sites displayed different projection patterns than those associated with the nucleus accumbens. Fibers from the olfactory tubercle region were distributed primarily to the thalamic nucleus, and anterior amygdala, cerebral cortex, mediodorsal hypothalamus; in cases in which the labeled cells were located exclusively within the substantia innominata, fibers were also traced through the medial forebrain bundle into the rostra1 part of the ventral tegmental area and adjacent substantia nigra. DISCUSSION The results of the present investigation reveal the presence of a topographical organization linking the nucleus accumbens with the tegmentum
C
FIG. 4. Outline of an injection of [“H]leucine placed of the nucleus accumbens and distribution of label plotted (E) levels (case 14).
into the ventrolateral from rostra1 (A)
aspect to caudal
NUCLECS
ACCUMBENS
l’ROJECTIOhY5
FIG. 5. Outline of an injection of [“Hjleucine placed into the dorsolateral the nucleus accumbens and distribution of label l)lotted from rostra1 (A) (F) levels (case 16).
103
aq)ect of to caudal
in the cat. The oganization of this system is depicted in Fig. 6 and its origin within the nucleus accumbens appears to resernhle the face of a clock. In brief, fibers arising from the dorsomedial aspect of the nucleus accunlbens (i.e., nucleus accumhens septi) are distihuted through the medial part of the medial forebrain bundle to the rostra1 end of the ventral tegtnental area. Fibers arising from the ventromedial aspect of the nucleus accumbens (i.e., nucleus accunlbens principalis pars medialis) pass through nlore lateral parts of the medial forebrain bundle and terminate in nlore caudal and lateral portions of the ventral tegmentum. Fillers arising front the lateral aspect of the nucleus accunlhens (i.e., nucleus accunlbens principalis pars lateralis and nucleus accunlhens caudati) project through the far lateral part of the medial forebrain bundle and medial tip of the internal capsule and terminate principally in the reticular division of the suhstantia nigra and more caudal and lateral levels of the central tegniental field. An additional sinall group of fibers also apl)ears to innervate the adjacent central gray substance. Furthermore, it also seenis likely that sonic fillers enroute to the nledial forebrain I~undle and adjoining internal
194
TROIANO
AND
SIEGEL
capsule, and which arise from lateral parts of the nucleus accumbens, synapse within the dorsolateral part of the substantia innominata and adjacent globus pallidus as well. Thus, the overall findings indicate that the nucleus accumbens and midbrain tegmentum share reciprocal connections that are topographically organized in both directions (5). Although the functional significance of this descending pathway to the tegmentum remains unknown, it is certainly possible that some of these fibers, at least, serve as an anatomic substrate underlying feedback regulation of basal forebrain dopamine levels. The present data raise a number of related experimental problems. First,
FIG. 6. Summary diagram illustrating topographical organization of descending fibers from the nucleus accumbens to the brain stem in the cat. Circles-fibers associated with the dorsomedial accumbens; squares-fibers associated with the ventromedial accumbens ; diamonds-fibers associated with the ventrolateral and dorsolateral accumbens.
Frc. nucleus medial lateral labeled site in grains
7. a-Photomicrograph of an injection site in the ventromedial aspect of the accumbens in case 3. X 13; h--presence of labeled axons of passage within the tip of the internal capsule in case 13 after an injection placed into the ventroaccumbens, X 230 ; c--dark-field photomicrograph indicating the presence of axon terminals kvithin the central tegmental field ipsilateral to the injection case 13, x 230; d--dark-field phototnicrogrnpll indicating abwnce of labeled in contralater:d central tegmeutnl ficltl in case 13, X 230.
196
TROIANO
AND
SIEGEL
it would be of interest to determine the nature of the transmitter associated with this pathway and its electrophysiological actions upon neurons in the tegmentum. Studies by Lorens et al. (13) have already suggested that serotonin may play a role in this system. It would be of further significance to compare the electrophysiologic effects of stimulation of the medial and lateral parts of the nucleus accumbens on tegmental neurons. Previous studies in our l.aboratory (6) and elsewhere (IO, 17) showed that lateral hypothalamic stimulation can inhibit or excite tegmental neurons. Therefore, it would also be important to compare the effects of medial forebrain bundle activation on mesencephalic neurons when this fiber system is driven from the nucleus accumbens in contrast to when it is driven from the lateral hypothalamus. Finally, a possible resolution to the issue of whether the nucleus accumbens is more closely associated with the limbic system or with the basal ganglia is evident from the present data. Because the medial aspect of the nucleus accumbens projects to the ventral tegmental area whereas the lateral region projects to the reticular division of the substantia nigra and central tegmental field, it is apparent that this nucleus contributes to the organization of both these major functional systems. The critical problem to be solved from this finding is to determine whether the nucleus accumhens is simply an anatomic zone of a topographically organized transition between two functionally discrete systems or whether the nucleus accumbens may represent a focal point for the integration of signals which regulate components of both the basal ganglia and limbic system. The ventral tegmental area sends projections to a variety of limbic structures such as the anterior cingulate gyrus (l), septal area (12, 19), amygdala (21) , and olfactory tubercle (21) , whereas fibers from the substantia nigra innervate motor nuclei of the thalamus [i.e., ventrolateral (2). Therefore, and ventral anterior nuclei (2) 1, and the neostriatum activation of the nucleus accumbens could then result in the integration of discharge patterns in portions of both the basal ganglia and limbic system. The significance of the possible relationship between these two systems may potentially be better understood within the context of the total “behavioral response” traditionally associated with the limbic system. Such responses include feeding, aggression, and sexual behavior and obviously require for their fullest expression an intimate relationship between the limbic system and structures such as the various components of the basal ganglia which modulate the sensorimotor cortex. REFERENCES 1. BECKSTEAD, R. M. 1976. Convergent thalamic and mesencephalic projections the anterior medial cortex in the rat. J. Cotnp. Ncurol. 166: 403-416.
to
NITCLEI;S
AC’CL’MHl:NS
PROJECTIONS
107
2. CARPENTER, M. B., AND E’. PETER. 1972. h’igrostriatal and nigrothalamir fibers in the rhesus monkey. .I. (‘OIJI~. h’cwol. 144 : 93-116. 3. DE FRANCE, J. F.. AND H. YOSFIIHARA. 1975. Limbic input to the nucleus accumbens septi. Brain Kcs. 90: 159-163. 4. DILL, R. E., AND E. COSTA. 1977. Behavioral dissociation of the enkephalinergic systems of nucleus accumbens and nucleus caudatus. Nrlirnplrarirrncoiogy 16 : 323-326. 5. Do~r~srcrc, V. B., R. RI. BECKSTEAD, AND W. J. NANTA. 1976. Some ascending and descending projections of the substantia nigra and ventral tegmental area in the rat. Nczlvusri. .46&r. 2: 61. 6. EDIXGER, H. M., S. F. KRAMER, AND A. SIEXL. 1977. Effect of hypothalamic stimulation on mesencephalic neurons. Exp. Ncrtrol. 54 : 91-103. 7. Fox, C. A. 1940. Certain basal telencephalic centers in the cat. J. Cowp. hTczwol. 72: l-62. 8. GARDNER, C. R., AND S. W. PHILLIPS. 1976. Heterogeneity of inhibitory mechanisms in the nucleus accumbens and preoptic area of the rat. Br. J. Plramacol. 58: 267-268. 9. HEIMER, L., AND R. D. Wrtso~. 1975. The subcortical projections of the allocortex: Similarities in the new-01 associations of the hippocampus, the piriform cortex, and the neocortex. Pages 177-193 in M. Santini, Ed., Co& Cwtwrrial S~~rrposiw~ : Prrsprctivrs i~r ivrlrr~,hiolo~/~l. Raven Press, New York. 10. HL-ANG, Y. H., AIYD J. P. FLYNN. 1975. Unit activities in the hypothalamus and midbrain during stimulation of hypothalamic attack sites. Rwin Kcs. 93: 419440. 11. KOIKEGAMI, H., Y. HIRATA, AND J. OGREIIA. 1967. Studies on the paralimbic brain structures. I. Definition and delimitation of the paralimbic brain structures and some experiments on the nucleus accumbens. Folio Pqrhiat. Nrzrrol. Jap. 21: 151-180. 12. LINDVALL, 0. 1975. Mesencephalic dopaminergic afferents to the lateral septal nucleus in the rat. Brain Kcs. 87 : 89-95. 13. LOREITS, L. A., J. P. SOREKSEN, AND J. A. HARVEY. 1970. Lesions in the nuclei accumbens septi of the rat. J. Colrrp. Pkysiol. Ps~~cld. 73 : 284-290. 14. MEIBACH, R. C., AND A. SIEGEL. 1977. The efferent connections of the septal area in the rat: An analysis utilizing retrograde and anterograde transport methods. Bruin Krs. 119 : l-20. 1.5. NADTA, W. J. H. 1958. Hippocampal projections and related neural pathways to the midbrain in the cat. Bvni~ Kcs. 81 : 319-340. 16. POWELL, E, W., AND R. B. LEMAN. 1976. Connections of the nucleus accumbens, Brairr Rrs. 105: 389-403. 17. SUTIN, J., R. L. MCBRIDE, R. H. THALMANN, AND E. L. VAN ATTA. 1975. Organization of sotne brainstem and limbic connections of the hypothalamus. Plzarwncol. Biochrm. Bclrae~ 3: Suppl. 1, 49-59. 18. SWANSON, L. W., AND W. M. COWAN. 1975. A note on the connections and development of the nucleus accumbens. B&IL Rrs. 92: 324-330. 19. T’HOIAXO, R., AND A. SIEGEL. 1975. The ascending and descending connections of the hypothalamus in the cat. Exp. Ncurol. 49: 161-173. 20. TROIANO, R., AND A. SIEGEL. 1978. Efferent connections of the basal forebrain in the cat: The substantia innominata. E.zp. Nmrol. 61 : 198-213. 21. UNGIIRSTEDT, V. 1971. Stereotaxic mapping of the monoamine pathways in the rat brain. .-Zrttr )-‘h~si~~f. Smrtrl. 82 : Suppl. 367, l-48.
Efferent
KEC’ROLOGY
61, 185-197 (1978)
Connections The RAYMOND
of the Nucleus TROIAN~
Basal
Forebrain
in the Cat:
Accumbens AND ALLAN
SIEGEL 1
The efferent connections of the nucleus accumbens in the cat were studied with radioautographic methods. Injections of [“Hlleucine were placed throughout the extent of this structure in adult cats. The results revealed the presence of a topographical organization of the projections from the nucleus accumbens to the brain stem. Fibers arising from the dorsomedial sector of the nucleus accumbens project through the medial aspect of the medial forebrain bundle to the rostra1 end of the ventral tegmental area. Fibers arising from the ventromedial sector of the nucleus accumbens project to slightly more caudal and lateral parts of the ventral tegmentum. In contrast, fibers which arise from lateral parts of the nucleus accumbens project through the lateral aspect of the medial forebrain bundle and medial tip of the internal capsule to terminate primarily within the pars reticulata of the substantia nigra and central tegmental field. The data also suggest that fibers from this part of the nucleus accumbens probably terminate within the dorsolateral aspect of the substantia innominata and adjacent parts of the pallidum. These findings indicate that the nucleus accumbens is linked to both the limbic system and the basal ganglia.
INTRODUCTION The nucleus accumbens has heen the subject of a number of recent (5, 9, 11, 16, 18), electrophysiological (3, S), and behavioral (4, 13) experiments. In spite of the findings from these investigations its integration with any of the subsystems of the forebrain has remained an enigma. Early anatomic descriptions of this structure focussed on its anatomical
1 This project was supported by National Institutes of Health Research Grant NS 07941-08, awarded by the National Institute of Neurological and Communicative Disorders and Stroke. The authors wish to acknowledge Mrs. Jean Holland for typing this manuscript.
185 0014-4886/78/0611
-OlSS$OZ.OO/O
Copyright 0 1978 by Academic Press,Inc. All rights of rcprodurtion in any form reserved.
186
TROIANO
AND
SIEGEL
proximity and possible association with neostriatum (7). Several recent studies supported this position. Swanson and Cowan (18) claimed that fibers from the nucleus accumbens are distributed to the entopeduncular nucleus and substantia nigra, thus leading them to conclude that the nucleus accumbens constitutes part of the basal ganglia. Heimer and Wilson (9) introduced the term “ventral striatum” to underline the similarities between the nucleus accumbens and neostriatum in regard to both untrastructure and input-output relationships. In contrast, several other authors linked the nucleus accumbens with the limbic system after having identified efferent connections of that structure with the lateral hypothalamus and midbrain tegmentum (11, 16). The presence of afferent fiber connections to the nucleus accumbens from the tegmentum (5) as well as from the fornix system (3) also seems to suggest its classification with the limbic system. Koikegami et al. (11) noted that the nucleus accumbens can be divided into several subnuclear groups. These include two dorsal and two ventral nuclei. Within the dorsal group the cell cluster situated proximal to the caudate nucleus is referred to as the nucleats accztnabens caudati and the cell group proximal to the lateral septal nucleus is called the nbfcleus accun+ bens septi. Ventral to these groups are two additional and larger cell clusters identified as the nucleus alccuwtbens principalis pars lateralis and pars medialis, respectively [see (11. p. 157), Figs. 1 and 21. Parceling of the nucleus accumbens into these subdivisions, when considered collectively with the opposing findings which attempt to link this structure to either the basal ganglia or limbic system, clearly suggests that it may constitute a transitional zone that is related to both of these systems. Such a hypothesis would predict that the nucleus accumbens represents a continuum of cells whose axons project topographically to structures associated with both the basal ganglia and limbic system. In the study described below a test of this hypothesis was made by examining the efferent connections of the nucleus accumbens by the radioautographic technique in the cat. METHODS Twenty-four adult cats of either sex were used in this study. Surgery was carried out under Nembutal anesthesia (40 mg/kg, i.p.) utilizing clean conditions. Injections of radioisotope were placed stereotaxically with the use of a l-p1 Hamilton microsyringe mounted to the stereotaxic carrier. Injections were made with commercially prepared tritiated leucine (L- [4,5-3H]leucine, sp act 30 to 50 Ci/mmol, New England Nuclear, Boston, Massachusetts). The isotope was concentrated to 10 &i/PI by evaporation and reconstitution in sterile distilled water. A quantity of
NUCLEL-S
ACCUMRENS
187
PROJECTIONS
0.2 ~1 was injected into different parts of the nucleus accumbens and adjacent regions over a period of 40 min. A micromanipulator positioned directly on top of the syringe allowed for mliform advancement of the plunger which produced slow and controlled delivery of the isotope. Survival times varied from 1 to 2 days following placement of the injection. Cats were then killed with an overdose of I2’emhutal and perfused transcardially with solutions of 0.9? saline and 10% formalin. Brains were removed, stored in 10% formalin for several weeks, and then embedded in paraffin. They were serially sectioned at 8 to 10 pm, mounted on glass slides, and deparaffinized. Sections were coated with Kodak NTB or NTB-2 emulsion and exposed for approximately 2 to 3 weeks. Autoradiographs were developed in D-19, fixed in Ektaflo, and stained lightly with cresylechtviolet. RESULTS Injections were placed throughout the extent of the nucleus accumbens and included each of the cell divisions described by Koikegami et nl. ( 11) . The principal focus of each injection is summarized in Table 1 and illustrated in Fig. 1. Projectiorls
~YOIII
the
Dorsal
and Medial
Asficcts
of the Nucleus
rlccurrl-
hr~. In two cases injections were placed into the dorsomedial aspect of the nucleus accumbens. These injection sites correspond to the region designated as the rllrcl~~s accunlhem septi by Koikegami et al. (11). In case 1 the injection site included the dorsomedial aspect of the nucleus and the cells adjacent to the vertical limb of the diagonal band. In case 2 the injection \vas placed more rostrally and included cells in both the dorsomedial accumbens and adjoining infralimbic area. I,abeled axons were traced in a ventral and caudal direction through the preoptic area and into the medial forebrain bundle. Caudally descending labeled axons situated in the perifornical region could be followed through the hypothalamus to the rostra1 end of the ventral tegmental area. Labeled axon terminals were noted only in the medial aspect of this region at a level just behind the mammillary bodies (Fig. 2). Other fibers from the injection sites were traced through the stria medullaris to the medial half of the medioclorsal thalamic nucleus. However, hasetl on other studies described in the rat (141, it is likely that this component of the projection is more closely related to cell groups adjacent to the vertical limb of the diagonal band than to cells situated exclusively in the nucleus accumbensproper. Yrojcrtiorls
f~onr tltc Ve~ztroulrdial
A.s/vct
of the ATuclr~rs
Accul~~Dcns.
In five cases injections were placed in the ventrometlial aspect of the nucleus accmmben-the area identified by Koikegami ct al. ( 11) as the n//c-lrus c~ccu~~~Zuv~s priwipa1i.s pars rrwdialis. In three of the cases the
188
TROIANO
AND
TABLE Principal Case
1 2 3 4 5 6 7 8 9 10 11 12 13 14 1.5 16 17 18
Injection site within nucleus accumbens
Dorsomedial Dorsomedial Ventromedial Ventromedial Ventromedial Ventromedial Ventromedial Ventrolateral Ventrolateral Ventrolateral Ventrolateral Ventrolateral Ventrolateral Ventrolateral Dorsolateral Dorsolateral Dorsolateral Dorsolateral
foci of Labeled Ventral
tegmental area
Axon
SIEGEL
1 Terminals
in the Brain
Substantia Compact zone
++ ++ ++ i-f ++ ++ fS ++ 0 ? ++ 0 0 0 0 0 0 0
0 0 0 0 0 0 0 0 ? I ? 0 0 0 0 0 0 0
a + to + + +-increasing quantities of labeled axon labeled axon terminals; ?-uncertainty due to difficulty labeled axon terminals and fibers of passage.
nigra
Stem% Central
tegmental field
Reticular zone 0 0 0 0 0 0 0 0 + I ? + ? + + + ? ? terminals; O-no in discriminating
0 0 0 0 0 0 0 ++ + ? + +++ +++ + +-t-+ +-I-+ +++ +++ evidence of between
labeled cells were limited to the medial half of the nucleus accumbens principalis and in two other animals the injection sites also labeled a few cells in the adjacent olfactory tubercle. The projections to the brain stem from each of these injections were similar but differed somewhat from the pathway arising from the nucleus accumbens septi described above. Labeled axons were traced in a caudal direction from their origin in the nucleus accumbens principalis into the medial forebrain bundle at a position slightly lateral to the position occupied by fibers associated with the nucleus accumbens septi. Labeled axons could be further traced through the entire extent of the hypothalamus and ventral tegmentum to the level of the red nucleus. This fiber bundle appeared to terminate exclusively within the ventral tegmental area but in a position slightly caudal and lateral to the fibers arising from the nucleus accumbens septi. It did not appear from the injections involving either the dorsomedial or ventromedial aspects of the nucleus accumbens that any labeled fibers terminated directly within the substantia innominata. In fact, the labeled
NUCLEI5
ACCUMHENS
PKOJKTIONS
189
FIG. 1. Schematic representation of all injection sites of [“Hlleucine described in text. Diagrams from rostra1 (A) and caudal (B) nucleus accumbens. Abbreviations in this and the following figures : A-nucleus accumbens ; AC-anterior commissure ; C-caudate nucleus ; CP-cerebral peduncle ; CTF-central tegmental field ; ICinternal capsule ; IP-interpeduncular nucleus ; LH-lateral hypothalamus ; Mmammillary bodies ; MD-mediodorsal thalamic nucleus ; OT-olfactory tubercle ; PG-pontine gray ; PO-preoptic area ; K-red nucleus ; S-septal area ; VY-ventral tegmental area.
fibers of passage seemed to be contained entirely within the substance of the preoptic area. The possible presence of terminals in the preoptic region could not be excluded. Projections from the Vc~~t~olateral As/m-t of tlze AFltclrirs .-fcciflrlhols. In seven cats injections were placed into the ventrolateral part of the nucleus accumbens-the region proximal to the anterior commissure and identified as the alcclezts acczil1lben.r )mincipalis pavs lateralis by Koikegami ct al. (11). In two additional cases the injections appeared to extend beneath the ventral border of the nucleus accumbens and labeled cells associated with the olfactory tubercle and the rostra1 limit of the substantia innominata. The projections from each of these injections seemed to he substantially similar. The primary projection from this part of the nucleus accumbens. which is typical for other cases with similar injection sites, is shown in Fig. 3. From the injection site labeled axons were observed passing in a caudal direction through the substantia innominata in a position proximal to the globus pallidus to join the medial tip of the internal capsule and lateral part of the medial forebain bundle. It is entirely l)ossible, and indeed, probable that some of these labeled fillers terminate in the dorsolateral part of the substantia innominata-the region identified by
190
TROIANO
AND
SIEGEL
Heimer and Wilson (9) as the ventral pallidurn. However, as many labeled fibers of passage are present in this region as well, a definitive statement concerning whether or not any of this label was present over terminals cannot be made at this time. The larger percentage of axons could be followed passing caudally within the internal capsule and medial forebrain bundle to the level of the red nucleus, at which point they entered and terminated within the reticular division of the substantia nigra and overlying central tegmental area. The terminal distribution sites of these fibers were located more caudally and laterally within the midbrain than the projections arising from more medial parts of the nucleus accumbens described in Fig. 3. Projections from the Ctousolateral Part of the Nucletas Acctmbens. In four cases injections involved the dorsolateral aspect of the nucleus accumbens-the region identified as the nwleus accunzbens caudati by Koikegami ef al. (11). The projections from this part of the nucleus accumbens to the brain stem were similar to the pathway arising from the ventrolateral division of the nucleus accumbens (Fig. 4). The fibers from the dorsolateral part of the nucleus accumbens pass through the substantia innominata and proximal part of the globus pallidus prior to entering the medial part of the internal capsule and lateral aspect of the medial forebrain bundle. Again, it is quite likely that some of these fibers in fact
FIG. 2. Outline of an injection of [“Hlleucine placed into the dorsomedial the nucleus accumbens and distribution of label plotted from rostra1 (A) (D) levels (case 1).
aspect of to caudal
NUCLEUS
ACCUMRENS
PROJECTIONS
191
FIG. 3. Outline of au injection of [“H]leucine placed into the ventromedial aspect of the nucleus accumbens and distribution of label plotted from rostra1 (A) to caudal (E) levels (case 3). terminate within these regions. Because many descending fibers of passage are also clearly present here it is difficult, if not impossible, to conclude unequivocally that any of the labeled grains are present over axon terminals.
Nevertheless,
tributed
to the
central
tegmental
Although lateral
not
also
pathway more
arising caudally
into
group
the
anterior
of
in
the
with
which
In
projections
the
the
central
terminal
elsewhere
nigra. dorso-
the
the
disof the
of the
through
appeared
stem-at
were part
substantia
supply
general!
arising
the
passed to
brain
fibers lateral
injections
accmllbens
the
the
of
cases
direction
of in
division all
of fibers
dorsolateral
laterally
any
the
for
to be present.
from
terminated
reticular
dorsomedial
appeared and
juncture--than
a
accunihens. Control 1lljrctiou.r. placetl in neighborillg placed
and
majority
they
observed
a small in
overwhelming
where
field
field
substance
stem,
uniformly
accumbens,
tegmental
the
brain
to
central
gra)
fields
of the
he situated
midbrain-pontine
in the
nucleus
In a series of aninlals injections of [“Hlleucine were regions of the forebrain. In one case an injection part
of
the
lateral
septal
nucleus
resulted
in
a
192
TROIANO
AND
SIEGEL
massive distribution of labeled fibers to the preoptic area and anterior lateral hypothalamus via the medial forebrain bundle. This classical projection was described with degeneration techniques in the cat (15) and most recently with radioautography in the rat (14). In other cases [described in the following paper (20) ] injections involved the olfactory tubercle and adjacent portions of the substantia innominata. The projections from all but one of these sites displayed different projection patterns than those associated with the nucleus accumbens. Fibers from the olfactory tubercle region were distributed primarily to the thalamic nucleus, and anterior amygdala, cerebral cortex, mediodorsal hypothalamus; in cases in which the labeled cells were located exclusively within the substantia innominata, fibers were also traced through the medial forebrain bundle into the rostra1 part of the ventral tegmental area and adjacent substantia nigra. DISCUSSION The results of the present investigation reveal the presence of a topographical organization linking the nucleus accumbens with the tegmentum
C
FIG. 4. Outline of an injection of [“H]leucine placed of the nucleus accumbens and distribution of label plotted (E) levels (case 14).
into the ventrolateral from rostra1 (A)
aspect to caudal
NUCLECS
ACCUMBENS
l’ROJECTIOhY5
FIG. 5. Outline of an injection of [“Hjleucine placed into the dorsolateral the nucleus accumbens and distribution of label l)lotted from rostra1 (A) (F) levels (case 16).
103
aq)ect of to caudal
in the cat. The oganization of this system is depicted in Fig. 6 and its origin within the nucleus accumbens appears to resernhle the face of a clock. In brief, fibers arising from the dorsomedial aspect of the nucleus accunlbens (i.e., nucleus accumhens septi) are distihuted through the medial part of the medial forebrain bundle to the rostra1 end of the ventral tegtnental area. Fibers arising from the ventromedial aspect of the nucleus accumbens (i.e., nucleus accunlbens principalis pars medialis) pass through nlore lateral parts of the medial forebrain bundle and terminate in nlore caudal and lateral portions of the ventral tegmentum. Fillers arising front the lateral aspect of the nucleus accunlhens (i.e., nucleus accunlbens principalis pars lateralis and nucleus accunlhens caudati) project through the far lateral part of the medial forebrain bundle and medial tip of the internal capsule and terminate principally in the reticular division of the suhstantia nigra and more caudal and lateral levels of the central tegniental field. An additional sinall group of fibers also apl)ears to innervate the adjacent central gray substance. Furthermore, it also seenis likely that sonic fillers enroute to the nledial forebrain I~undle and adjoining internal
194
TROIANO
AND
SIEGEL
capsule, and which arise from lateral parts of the nucleus accumbens, synapse within the dorsolateral part of the substantia innominata and adjacent globus pallidus as well. Thus, the overall findings indicate that the nucleus accumbens and midbrain tegmentum share reciprocal connections that are topographically organized in both directions (5). Although the functional significance of this descending pathway to the tegmentum remains unknown, it is certainly possible that some of these fibers, at least, serve as an anatomic substrate underlying feedback regulation of basal forebrain dopamine levels. The present data raise a number of related experimental problems. First,
FIG. 6. Summary diagram illustrating topographical organization of descending fibers from the nucleus accumbens to the brain stem in the cat. Circles-fibers associated with the dorsomedial accumbens; squares-fibers associated with the ventromedial accumbens ; diamonds-fibers associated with the ventrolateral and dorsolateral accumbens.
Frc. nucleus medial lateral labeled site in grains
7. a-Photomicrograph of an injection site in the ventromedial aspect of the accumbens in case 3. X 13; h--presence of labeled axons of passage within the tip of the internal capsule in case 13 after an injection placed into the ventroaccumbens, X 230 ; c--dark-field photomicrograph indicating the presence of axon terminals kvithin the central tegmental field ipsilateral to the injection case 13, x 230; d--dark-field phototnicrogrnpll indicating abwnce of labeled in contralater:d central tegmeutnl ficltl in case 13, X 230.
196
TROIANO
AND
SIEGEL
it would be of interest to determine the nature of the transmitter associated with this pathway and its electrophysiological actions upon neurons in the tegmentum. Studies by Lorens et al. (13) have already suggested that serotonin may play a role in this system. It would be of further significance to compare the electrophysiologic effects of stimulation of the medial and lateral parts of the nucleus accumbens on tegmental neurons. Previous studies in our l.aboratory (6) and elsewhere (IO, 17) showed that lateral hypothalamic stimulation can inhibit or excite tegmental neurons. Therefore, it would also be important to compare the effects of medial forebrain bundle activation on mesencephalic neurons when this fiber system is driven from the nucleus accumbens in contrast to when it is driven from the lateral hypothalamus. Finally, a possible resolution to the issue of whether the nucleus accumbens is more closely associated with the limbic system or with the basal ganglia is evident from the present data. Because the medial aspect of the nucleus accumbens projects to the ventral tegmental area whereas the lateral region projects to the reticular division of the substantia nigra and central tegmental field, it is apparent that this nucleus contributes to the organization of both these major functional systems. The critical problem to be solved from this finding is to determine whether the nucleus accumhens is simply an anatomic zone of a topographically organized transition between two functionally discrete systems or whether the nucleus accumbens may represent a focal point for the integration of signals which regulate components of both the basal ganglia and limbic system. The ventral tegmental area sends projections to a variety of limbic structures such as the anterior cingulate gyrus (l), septal area (12, 19), amygdala (21) , and olfactory tubercle (21) , whereas fibers from the substantia nigra innervate motor nuclei of the thalamus [i.e., ventrolateral (2). Therefore, and ventral anterior nuclei (2) 1, and the neostriatum activation of the nucleus accumbens could then result in the integration of discharge patterns in portions of both the basal ganglia and limbic system. The significance of the possible relationship between these two systems may potentially be better understood within the context of the total “behavioral response” traditionally associated with the limbic system. Such responses include feeding, aggression, and sexual behavior and obviously require for their fullest expression an intimate relationship between the limbic system and structures such as the various components of the basal ganglia which modulate the sensorimotor cortex. REFERENCES 1. BECKSTEAD, R. M. 1976. Convergent thalamic and mesencephalic projections the anterior medial cortex in the rat. J. Cotnp. Ncurol. 166: 403-416.
to
NITCLEI;S
AC’CL’MHl:NS
PROJECTIONS
107
2. CARPENTER, M. B., AND E’. PETER. 1972. h’igrostriatal and nigrothalamir fibers in the rhesus monkey. .I. (‘OIJI~. h’cwol. 144 : 93-116. 3. DE FRANCE, J. F.. AND H. YOSFIIHARA. 1975. Limbic input to the nucleus accumbens septi. Brain Kcs. 90: 159-163. 4. DILL, R. E., AND E. COSTA. 1977. Behavioral dissociation of the enkephalinergic systems of nucleus accumbens and nucleus caudatus. Nrlirnplrarirrncoiogy 16 : 323-326. 5. Do~r~srcrc, V. B., R. RI. BECKSTEAD, AND W. J. NANTA. 1976. Some ascending and descending projections of the substantia nigra and ventral tegmental area in the rat. Nczlvusri. .46&r. 2: 61. 6. EDIXGER, H. M., S. F. KRAMER, AND A. SIEXL. 1977. Effect of hypothalamic stimulation on mesencephalic neurons. Exp. Ncrtrol. 54 : 91-103. 7. Fox, C. A. 1940. Certain basal telencephalic centers in the cat. J. Cowp. hTczwol. 72: l-62. 8. GARDNER, C. R., AND S. W. PHILLIPS. 1976. Heterogeneity of inhibitory mechanisms in the nucleus accumbens and preoptic area of the rat. Br. J. Plramacol. 58: 267-268. 9. HEIMER, L., AND R. D. Wrtso~. 1975. The subcortical projections of the allocortex: Similarities in the new-01 associations of the hippocampus, the piriform cortex, and the neocortex. Pages 177-193 in M. Santini, Ed., Co& Cwtwrrial S~~rrposiw~ : Prrsprctivrs i~r ivrlrr~,hiolo~/~l. Raven Press, New York. 10. HL-ANG, Y. H., AIYD J. P. FLYNN. 1975. Unit activities in the hypothalamus and midbrain during stimulation of hypothalamic attack sites. Rwin Kcs. 93: 419440. 11. KOIKEGAMI, H., Y. HIRATA, AND J. OGREIIA. 1967. Studies on the paralimbic brain structures. I. Definition and delimitation of the paralimbic brain structures and some experiments on the nucleus accumbens. Folio Pqrhiat. Nrzrrol. Jap. 21: 151-180. 12. LINDVALL, 0. 1975. Mesencephalic dopaminergic afferents to the lateral septal nucleus in the rat. Brain Kcs. 87 : 89-95. 13. LOREITS, L. A., J. P. SOREKSEN, AND J. A. HARVEY. 1970. Lesions in the nuclei accumbens septi of the rat. J. Colrrp. Pkysiol. Ps~~cld. 73 : 284-290. 14. MEIBACH, R. C., AND A. SIEGEL. 1977. The efferent connections of the septal area in the rat: An analysis utilizing retrograde and anterograde transport methods. Bruin Krs. 119 : l-20. 1.5. NADTA, W. J. H. 1958. Hippocampal projections and related neural pathways to the midbrain in the cat. Bvni~ Kcs. 81 : 319-340. 16. POWELL, E, W., AND R. B. LEMAN. 1976. Connections of the nucleus accumbens, Brairr Rrs. 105: 389-403. 17. SUTIN, J., R. L. MCBRIDE, R. H. THALMANN, AND E. L. VAN ATTA. 1975. Organization of sotne brainstem and limbic connections of the hypothalamus. Plzarwncol. Biochrm. Bclrae~ 3: Suppl. 1, 49-59. 18. SWANSON, L. W., AND W. M. COWAN. 1975. A note on the connections and development of the nucleus accumbens. B&IL Rrs. 92: 324-330. 19. T’HOIAXO, R., AND A. SIEGEL. 1975. The ascending and descending connections of the hypothalamus in the cat. Exp. Ncurol. 49: 161-173. 20. TROIANO, R., AND A. SIEGEL. 1978. Efferent connections of the basal forebrain in the cat: The substantia innominata. E.zp. Nmrol. 61 : 198-213. 21. UNGIIRSTEDT, V. 1971. Stereotaxic mapping of the monoamine pathways in the rat brain. .-Zrttr )-‘h~si~~f. Smrtrl. 82 : Suppl. 367, l-48.
