research report efferent projections of infralimbic and

Brain Research 888 (2001) 83–101www.elsevier.com/ locate /bres

Research report

Efferent projections of infralimbic and prelimbic areas of the medialprefrontal cortex in the Japanese monkey, Macaca fuscata

a , b b*Tanemichi Chiba , Tetsuro Kayahara , Katsuma NakanoaDepartment of Anatomy and Neurobiology, Chiba University School of Medicine, Inohana 1-8-1, Chuo-Ku, Chiba 260-8670, Japan

bMie University School of Medicine, Tsu 514-8507, Japan

Accepted 19 September 2000

Abstract

The infralimbic area (IL) and prelimbic area (PL) have been postulated as an autonomic motor region in the medial prefrontal cortex.The present study was conducted to reveal the projection sites of IL and PL of the monkey, Macaca fuscata, using biotinylated dextranamine as an anterograde tracer. IL and PL projected densely to the ventromedial caudate nucleus, the core and shell of the nucleusaccumbens (Acb), parvicellular lateral basal and magnocellular accessory basal nuclei of the amygdala, lateral preoptic area, ventromedialhypothalamic nucleus, tubero-mammillary nucleus (TM), medial part of the magnocellular and dorsal part of the parvicellular (MDpc)dorsomedial thalamic nuclei, reunience and medial part of the medial pulvinar nucleus, and dorso-lateral part of the periaqueductal gray(PAGdl) in the mesencephalon. Moderately to weakly projected areas were the intermediate and lateral parts of the agranular insularcortex, orbital part of area 12, agranular and dysgranular part of the temporal pole cortex (TPa-g), auditory temporal cortex, lateral andmedial (MS) septal nuclei, bed nucleus of the stria terminalis, diagonal band of Broca, substantia innominata, and medial preoptic area,dorsomedial, lateral, and posterior hypothalamic nuclei, magnocellular lateral basal and lateral amygdaloid nuclei, paratenial,paraventricular (PV), inter-antero-medial (IAM), reticular, central medial (CeM), parafascicular (PF) and limitans nuclei of the thalamus,lateral habenular nucleus, pedunculo-pontine nucleus, dorsal part of the lateral lemniscal nucleus, ventral tegmental area (VTA), dorsalraphe, superior central nucleus, medial and lateral parabrachial nuclei (PBl) and nucleus locus coeruleus (LC). A few scattered terminalswere observed in the perifornical nucleus of the hypothalamus and substantia nigra pars compacta. PL and area 24 were characterized byprojections to the entorhinal (Ent) and piriform (Pir) cortex as well as to the magnocellular part of the ventral anterior thalamic nucleus(VAmc). The morphology of the terminal arborization in each nuclei was different in appearance, perhaps reflecting the synapticinteraction between the nerve terminals and postsynaptic dendrites. PL projected uniquely to Ent, Pir and VAmc and IL projected uniquelyto TPa-g, MS, IAM, CeM, MDpc, PF, PBl and LC. IL projected more strongly than PL to the shell of Acb, amygdaloid nuclei, PV, TM,VTA and PAGdl. The present results support the hypothesis that IL is a major cortical autonomic motor area and PL integrates limbic andautonomic inputs in the primate. 2001 Elsevier Science B.V. All rights reserved.

Theme: Other systems of the CNS

Abbreviations: ABmc, accessory basal amygdaloid nucleus, magnocellular part; Acb, accumbens nucleus; BLmc, basolateral amygdaloid nucleus,magnocellular part; BLpc, basolateral amygdaloid nucleus, parvicellular part; BST, bed nucleus of stria terminalis; CeM, central medial thalamic nucleus;CDvm, caudate nucleus, ventromedial part; DB, nucleus of diagonal band; DMH, dorsomedial nucleus of hypothalamus; DR, dorsal raphe nucleus; Ent,entorhinal cortex; Hbl, lateral habenular nucleus; Iai, agranular insular cortex, intermediate part; Ial, agranular insular cortex, lateral part; Iam, agranularinsular cortex, medial part; IAM, interanteromedial nucleus of thalamus; Iapm, agranular insular cortex, posteromedial part; L, lateral amygdaloid nucleus;LC, nucleus of locus coeruleus; LH, lateral hypothalamic nucleus; Lim, limitans nucleus of thalamus; LLd, dorsal nucleus of lateral lemniscus; LS, lateralseptal nucleus; MDmc, dorsomedial nucleus of thalamus, magnocellular part; MDpc, dorsomedial nucleus of thalamus, parvicellular part; MS, medialseptal nucleus; NCS, superior central nucleus; NTS, nucleu of tractus solitarius; PAG, periaqueductal gray; PAGdl, periaqueductal gray, dorso-lateral part;PBl, lateral parabrachial nucleus; PBm, medial parabrachial nucleus; PF, parafascicular nucleus of thalamus; PFC, prefrontal cortex; PfH, perifornicalnucleus of hypothalamus; PH, posterior hypothalamic nucleus; Pir, piriform cortex; PM, medial pulvinar nucleus; POL, lateral preoptic nucleus; POM,medial preoptic nucleus; PPN, pdunculo-pontine nucleus; PT: pretenial nucleus of thalamus; PV, paraventricular nucleus of thalamus; PVH, paraventricularnucleus of hypothalamus; R, reticular nucleus of thalamus; Re, reuniens nucleus of thalamus; RPC, reticular parvicellular nucleus; SI, substantiainnominata; SNc, substantia nigra, pars compacta; TA, temporal auditory cortical area; TM, tubero-mammillary nucleus of hypothalamus; TP, temporopolarcortex; TPa-g, temporopolar cortex, agranular area; TPdg, temporopolar cortex, dysgranular area; VAmc, ventral anterior nucleus of thalamus,magnocellular part; VMH, ventromedial nucleus of hypothalamus; VTA, ventral tegmental area; ZI, zona incerta; 12o, area 12o of prefrontal cortex; 12l,area 12l of prefrontal cortex

*Corresponding author. Tel.: 181-43-226-2022; fax: 181-43-226-2025.

E-mail address: [email protected] (T. Chiba).

0006-8993/01/$ – see front matter 2001 Elsevier Science B.V. All rights reserved.PI I : S0006-8993( 00 )03013-4

84 T. Chiba et al. / Brain Research 888 (2001) 83 –101

Topic: Limbic system and hypothalamus

Keywords: Infralimbic area; Prelimbic area; Medial prefrontal cortex; Monkey; Biotinylated dextran amine; Efferent connection

1. Introduction used BDA as a marker useful for precisely identifying theinjection sites and also followed very thin axons of the

The medial and ventral parts of the frontal lobe of the autonomic neurons in the central nervous system for amonkey can modulate autonomic parameters [21,41,59] considerable distance.and the role of the ventromedial frontal lobe in autonomicfunction is that individuals with lesions of this area areunable to generate autonomic responses to emotional 2. Materials and methodsstimuli. Remarkably, they are also impaired in makingjudgments about the consequences of their actions in social Ten adult Japanese monkeys, Macaca fuscata, of bothsituations, despite possessing the knowledge necessary to sexes and 3.5–5.5 kg body weight were used in the presentmake the correct decision [9,18,19]. Damasio and his study. All animal protocols were reviewed and approvedco-workers [19] have suggested that these two deficits are by the Animal Studies Committee of Mie University. Therelated and that the sociopath-behavior of these patients is animals were anesthetized with intramuscular injection ofdue to their inability to generate ‘somatic markers’ that tag ketamine hydrochloride (10 mg/kg) and then with pen-behavioral options as desirable or not [10]. The frontal tobarbital (25 mg/kg). The animals were fixed on stereo-lobe damage would lead to loss of affective responsiveness taxic apparatus, and 0.1–1.0 ml31–3 times of 5%and foresight arising from interoceptive agnosia [32]. biotinylated dextran amine (BDA, 10 000 MW, MolecularDamasio [18] extended this concept to explain the ‘ac- Probes Inc.) in physiological saline was injected through aquired sociopathy’ of patients with bilateral orbitofrontal glass micropipette (inner diameter of the tip was 30–50damage. mm) using a pneumatic picopump, Model PV800, under an

The viscerosensory and visceromotor areas in the frontal operation microscope. A period of 14–21 days later, thelobe are suggested to be localized in the agranular insular, animals were anesthetized again and fixed by perfusioninfralimbic and prelimbic cortex (IL and PL). The vis- with 1 l /kg body weight of 8% formalin (3.2% formalde-cerosensory inputs reach specific areas within the agranular hyde), 0.2% glutaraldehyde in 0.1 M phosphate buffer (pHinsula in primates as in rodents [14]. Thus, IL and PL have 7.4) and 1000 ml of 10% sucrose in phosphate buffer.been postulated to be an autonomic motor area in the After the perfusion, the brain was removed and placed inmedial prefrontal cortex (PFC), as efferent and afferent the 25% sucrose in phosphate buffer at 48C for a few days.connections of IL in the rat were examined by several Serial coronal sections were then cut at 50 mm thicknessworkers and IL and PL were found to be reciprocally with a freezing microtome. The serial sections wereconnected with most central autonomic nuclei as far as the classified into three groups and processed as follows. (1)spinal cord in the rat [5,4,27,54]. The sections were rinsed with 0.1 M phosphate buffered-

As recent studies revealed, the cytoarchitectonic map saline (PBS) and incubated in a solution composed ofand histochemical characteristics of the monkey PFC 15–17 ml of streptavidine, 1 ml of 2.5% Triton X-100, andincluding IL and PL [14], we designed the present study so 4 ml of 0.1 M PBS overnight. After rinsing five times withas to reveal the projection sites of the medial PFC of the PBS, the sections were incubated for 3–5 h withmonkey, Macaca fuscata, using biotinylated dextran amine diaminobenzidine solution composed of 50 ml 0.05 M(BDA) as an anterograde tracer. The injection sites covered Tris–HCl buffer (pH 7.6), 10 mg diaminobenzidine andPL, IL and the adjacent medial PFC including area 14, and 125 mg nickel ammonium, and added with 80–100 ml ofarea 24b. We concentrated on the projection pattern of IL 0.3% H O . (2) The sections were processed as in (1), but2 2

compared with PL in this study. Part of the present studies nickel ammonium was omitted from the final reactionwere presented as preliminary reports on several occasions solution and they were observed under-dark field optics.[17,38,39]. During the preparation of this manuscript, two (3) The sections were rinsed with the phosphate buffer,important results of projection studies of the medial processed for Nissl staining, and observed by microscopeprefrontal cortex of macaque monkeys, Macaca fas- and used for reference to identify nuclear orientation. Acicularis and Macaca nemestrina, were reported [3,43]. total of ten animals were used for the present study andThe major results were similar to ours except for a few seven of them were used for the following data analysis.different findings and the present results were carefully Animal number, sex, body weight, and BDA injectioncompared and discussed with those of their studies. We method of each animal are shown in the following list.

T. Chiba et al. / Brain Research 888 (2001) 83 –101 85

the serial frontal sections and were projected to the saggitalAnimal Sex Body Injection method of BDA plane of the medial prefrontal cortex as illustrated in Fig.No. weight 1A and B. The injection area of each animal was de-

(kg) termined by referring to the atlas of the medial prefrontalplane by Carmichael and Price [14] to identify the approxi-M686 Female 4.7 Injected 3 times in adjacent sites with 1.0, 1.0mate cortical area. The injection sites of cases M686 andand 1.0 ml while withdrawing the micropipette.M707 corresponded approximately to area 25 and caudal

M707 Female 4.5 Injected 2 times, 1.0 ml in one site and 0.810.2 ml area 32. Those of case M723 corresponded to rostral areain the adjacent site. 32 and caudal area 10m and case M746 to rostral area 32.

The injection sites of case M719 and M776 correspondedM719 Female 3.9 Injected 3 times in three adjacent sites, 0.610.6 ml, to area 24b and case M738 to the middle of area 10m. The

0.5 and 0.6 ml. cortical area was identified by referring to the cytoarchitec-tonic and immunohistochemical map of the rhesus monkey

M723 Female 6.8 Injected 2 times in the same site with 0.610.6 ml. by Carmichael and Price [14]. Cases M707 and M686 wereused for analysis of the projection sites of area 25 and

M738 Female 3.0 Injected 2 times in two adjacent sites, 0.310.3 andM723 and M746 for that of the projection sites of area 32.

0.310.25 ml.The results of cases M719, M738 and M776 were used ascontrol injections for areas 24b, 10m and 24a respectively.M746 Male 4.0 Injected 2 times in the same site with 0.210.25 ml

and 0.3 ml in the adjacent site.

3.2. Case M686M776 Female 2.9 Injected 3 times in the same site with 0.110.111.15

ml.DAB was injected into three adjacent locations of area

25 in this case as illustrated in Fig. 1. The distribution ofprojection nerve terminals was illustrated in Fig. 2. The

3. Results densest network of nerve terminals with many varicositieswas observed in the ventro-medial caudate nucleus

3.1. Injection sites (CDvm), core and shell of the nucleus accumbens (Acb),magnocellular accessory basal and parvicellular lateral

From a series of BDA injections into the medial basal (ABmc, BLpc) nuclei of the amygdala, and theprefrontal cortex of monkeys, seven cases were selected magnocellular part of the mediodorsal (MDmc) nuclei offor the present analyses of the projection pattern of the the thalamus. Moderately to weakly projected areas weremedial prefrontal cortex including the anterior cingulate the lateral septum (LS), bed nucleus of the stria terminaliscortex (area 24b), IL (area 25) and PL (area 32). Injection (BST), diagonal band of Broca (DB), substantia in-sites were identified by the distribution of pyramidal cells nominata (SI), dorsomedial, ventro-medial, lateral andlabeled by the uptake of injected BDA, reconstructed from posterior hypothalamic nuclei (DMH, VMH, LH, PH),

Fig. 1. Schema demonstrating the location of injection sites of BDA in seven monkeys used in the present analysis. (A) Injection sites of seven monkeyswere shown reconstructed from serial prefrontal sections and projected to the parasagittal plane. (B) The injection sites of BDA in the present study weresuperimposed on the map of the cytoarchitectonic cortical area of the prefrontal cortex of Carmichael and Price [14].


Fig. 2. A series of frontal sections of M686 demonstrating the distribution of nerve terminals labeled by BDA.

tubero-mammillary nucleus (TM), magnocellular part of central medial, reunience, parvicellular part of mediodor-basolateral and lateral amygdaloid nuclei (BLmc, L), sal, medial part of the medial pulvinar and limitans nucleiparatenial, paraventricular, reticular, inter-antero-medial, of the thalamus (PT, PV, R, IAM, CeM, Re, MDpc, PM,


between the nerve terminals and postsynaptic dendritesmay have been responsible. Typical photomicrographs ofsuch terminals were observed in the shell and core of Acb(Fig. 4B), BL and AB amygdaloid nuclei (Fig. 4A), TMhypothalamic nucleus (Fig. 4B), MDmc (Fig. 4C) and PMthalamic nuclei (Fig. 4C) and TP (Fig. 4A).

3.4. Case M746

The injection site in this case was localized in the rostralpart of area 32 as shown in Fig. 1. Moderately to weaklylabeled networks of nerve terminals were observed in Iai,12o of the frontal cortex, TA of the temporal cortex, coreand shell of Acb, DB, LS, CDvm, SI, as well as Ent andPir cortex. The nerve terminals were also seen in PT, PV,R, Re, MDmc, PM and Lim of the thalamus, POL, DMH,VMH, LH, PH and TM of the hypothalamus, Hbl, PPN,LLd, VTA, DR, NCS and PAGdl of the midbrain (Fig. 5).

Fig. 2. (continued) 3.5. Case M723

DAB was injected into the rostral part of area 32 andcaudal part of area 10m as illustrated in Fig. 1. Moderately

Lim), ventral tegmental area (VTA, A10), substantia nigra to weakly labeled nerve terminals with varicosities werepars compcta (SNc), dorsolateral part of the periaqueductal distributed in Iai, 12o and the medial part of the agranulargray (PAGdl) and nucleus locus coeruleus (LC). insular cortex, TPdg and TA of the temporo-polar cortex,

the core of Acb, DB, BST, LS, CDvm, SI, as well as Ent3.3. Case M707 and Pir in the frontal cortex, ABmc, BLmc, BLpc and L of

the amygdala, PV, R, Re, VAmc, MDmc, PM and Lim ofDAB was injected into area 25 and the caudal part of the thalamus, POL, DMH, VMH, LH, TM, PfH of the

area 32 as seen in Fig. 1. Projection sites of the case M707 hypothalamus, Hbl, VTA, DR, NCS, and PAGdl of the thewere schematically demonstrated in Fig. 3. The densest midbrain (Fig. 6).network of nerve terminals with many varicosities wasobserved in the CDvm, core and shell of Acb, BLpc andABmc, lateral preoptic area (POL), VMH, TM, medial part 3.6. Case M719of MDmc and dorsal part of MDpc, Re and PM of thethalamus, PAGdl, pedunculo-pontine nucleus (PPN) and DAB was injected almost exclusively in area 24b in thisdorsal lateral lemniscal nucleus (LLd). case as illustrated in Fig. 1. Medium to weakly labeled

Moderately to weekly projected areas were the lateral networks of nerve terminals were observed in Iai, Ial andpart of the agranular insular cortex (Ial), orbital part of areas 14 and 13 of the frontal cortex, TA of the temporo-area 12 (12o), agranular and dysgranular part of the polar cortex, core and shell of Acb, CDvm, DB, BST, LS,temporal pole cortex (TPa-g, TPdg), auditory temporal MS, SI and Pir in the cortex, ABmc, BLmc and BLpc ofcortex (TA), medial septal nucleus (MS), BST, DB, SI, the amygdala, PV, R, IAM, CeM, Re, VAmc, MDmc.medial preoptic area (POM), DMH, LH, PH of the MDpc, Lim of the thalamus, POL, POM, DMH, VMH,hypothalamus, Blmc, L of the amygdala, and magnocellu- LH, PH, TM, and PfH of the hypothalamus, Hbl, VTA,lar part of the ventral-anterior (VAmc), parafascicular (PF) SNc, DR, NCS, PAGdl, PBl and LC of the midbrain (Fig.and PT, PV, R, IAM, CeM and Lim of the thalamus, lateral 7).habenular nucleus (Hbl), VTA, dorsal raphe (DR), superiorcentral nucleus (NCS), medial and lateral parabrachialnuclei (PBm and PBl) and LC. 3.7. Case M776

Only a few scattered terminals were observed in theperifornical nucleus of the hypothalamus (PfH) and SNc. DAB was injected in the caudal part of area 24b and aThe morphology of the terminal arborization in each nuclei small part of area 25a in this case as depicted in Fig. 1. Awas different in appearance for which synaptic interaction strongly labeled nerve terminal network was observed in


Fig. 3. A series of frontal sections of M707 showing the distribution of nerve terminals labeled by BDA. Note that retrogradely labeled nerve cells are alsoobserved in this particular case (indicated by triangles).


Fig. 3. (continued)

CDvm, core and shell of Acb and rostral cingulate cortex. as well as ABmc, BLmc, BLpc in the amygdala. MediumThe distribution of labeled terminals was more intense in intensity of labeling was also seen in DR, NCS and PAGdlthe core than the shell of Acb. Moderately labeled termi- of the midbrain.nals were seen in the Iai, Ial, and 12o of the prefrontalcortex, and TPa-p segment of the temporo-polar cortex. 3.8. Case M738

Medium density of nerve terminals was seen in DB, LS,MS, SI and PV, MDmc, PF, PM, IAM and CeM of the The injection site in this case was small and localized inthalamus. Medium to weak labeling was seen in POL, area 10m as shown in Fig. 1. Distribution of labeled nervePOM, DMH,VMH, LH, PH, and TM of the hypothalamus, terminals was very limited, being mainly found in the


Fig. 4. Representative photomicrographs of nerve terminals and cells labeled by biotinylated dextran amine injected into the medial prefrontal cortex. Anetwork of fine axons and small varicosities is seen in accessory basal (AB) and basolateral (BL) nuclei of amygdala (upper panel, M686). A column-likedistribution of labeled nerve terminals and some pyramidal cells in the layer V is observed in the temporo-polar cortex (TP). The surface of the cortex isdirected to the left (lower panel, M707). Scale bar550 mm in AB and BL; 200 mm in TP. A dense network of fine nerve terminals and varicosities is seenin both core and shell of the nucleus accumbens (upper panel, Acb M686). A dense network of axon terminals and varicosities is observed in thetuberomammillary nucleus of the hypothalamus (lower panel, TM M707). Scale bar5100 mm in Acb and 20 mm in TM. A dense network of fine nerveterminals and axon varicosities is seen in the magnocellular part of the dorsomedial nucleus (MD) and medial part of the medial pulvinar nucleus (PM) ofthe thalamus (lower panel, M707). Scale bar550 mm.


Fig. 4. (continued)

prefrontal cortex as Iai, Iam, IL and 12o as well as in TA 3.9. A summary of the efferent projections of the IL andof the temporo-polar cortex. Weakly labeled nerve termi- PL from the seven cases examined in the present studynals were scattered in the core and shell of Acb, CDvm,LS, IAM and PF of the thalamus and POL of the In the present study, the projection sites of areas 25 andhypothalamus. 32 could be clearly differentiated by comparing the


Fig. 4. (continued)

observations of M686 and M707 versus M746 and M723. hand, projections to Ent, Pir and VAmc were observedProjections to TPa-g, MS, IAM, CeM, MDpc, PF, PBl, and from area 32 but not from area 25. Projection sites of areaLC were observed from area 25 but not from the area 32. 24 seemed to be similar but much weaker than that of areaFurther, projections to TPdg, TA, CDvm, DB, BST, LS, 25.SI, core and shell of Acb, ABmc, BLmc, BLpc, PT, PV, R, In summary, the projection sites of IL were classifiedRe, MDmc, PM, Lim, VMH, LH, PH, TM, VTA, and into four major categories (Fig. 8). The first was the limbicPAGdl were stronger from area 25 than 32. On the other sensory integration area including the amygdaloid, tem-


Fig. 5. A series of frontal sections of M746 showing the distribution of nerve terminals labeled by BDA.

poro-polar cortex, PM and midline nuclei of the thalamus. concerned with the coordinated regulation of autonomicSecond was the autonomic relay nuclei such as LH, PAGdl motor with behavioral outputs such as shell and core ofof the mesencephalon, and parabrachial nuclei. The third Acb, ventral pallidum, ventral tegmental area and MD. Thewas the nuclei related with feedback loop in parallel to the fourth was amine containing neuronal cell groups such ascortico-striato-pallido-thalamic motor loop that is histaminergic TM, dopaminergic VTA, serotonergic DR,


Fig. 5. (continued)

and noradrenergic LC that are most likely concerned with 14 of mPFC as shown in Fig. 1. The areas in the mPFCthe regulation of the synaptic activity in various target were determined by referring to the map of the monkeynuclei of these amine neurons. described by Carmichael and Price [14]. The actual shape

of the brain of the monkeys used in this study varied fromanimal to animal; also, the species used in this study was

4. Discussion the Japanese monkey, Macaca fuscata, which belongs tothe same class but in different subclass from Macaca

4.1. Methodological considerations nemestrina, fascicularis and mulatta used by Carmichaeland Price [14]. The location maps of the areas were not as

The present study using BDA as an anterograde tracer exact as those determined cytoarchitectonically or histo-clearly demonstrated the distribution of labeled perikarya chemically. The results, however, clearly showed theand dendrites in the injection sites of mPFC, and axons and difference in projection patterns after the tracer wasterminal arborizations with varicosities in the projected injected in area 25 compared to the other cases in whichareas. The axons of autonomic and limbic nervous systems the marker was injected in rostral area 32, areas 10, 24bwere extremely thin and it was difficult to identify the and 14.labeled ones with a low magnification microscope, butwere able to observe them at higher magnifications even if 4.2. Projection to prosencephalonthey were scattered in a few fiber bundles or a network ofaxon terminals. Dark-field optics also helped the observa- Projections from IL and PL were observed in Iai, Ial andtion of thin bundles and terminal axon networks. The 12o of the frontal cortex and TPa-g, TPdg and TA of TP.optimal duration of survival time after the injection of Iai and Ial have been determined as presser related areas ofBDA was presumed to be 2 or 3 weeks, although success- PFC in the rat [60] and have reciprocal connections withful labeling seemed to be intimately related to the amount IL and PL of mPFC [54,60]. IL projects to area 14 (Gyrusof uptake of tracers rather than the survival period of the rectus) [37] and PL projects to IL, and areas 14, 24, 9 andanimals. We could follow projection axon terminals as far 10 [42].as the medulla oblongata in this study, and only a few TPa-g, TPdg and TA in TP are also reciprocallyscattered nerve terminals were found in NTS. The loca- connected with IL, PL, and areas 24, 12 and 13 in thetions of the injection sites of BDA were reconstructed from monkey [15,42] and the distribution of nerve terminals hadserial frontal sections, superimposed on the parasagittal a column-like appearance (Fig. 4A). Auditory inputsplane of mPFC and distributed in areas 25, 32, 24b, 10 and predominate in the dorsolateral part and visual inputs



become prominent only in the ventral portion of this temporal cortex are concerned with object recognitionregion. Olfactory inputs are directed mostly to the medial memory and IL also has additional access to this in-part of the temporal pole where extensive projections from formation via reciprocal connections with the medialthe amygdaloid nuclei also converge. Afferents from MDmc in rhesus monkeys [6,42]. IL, PL, and areas 13 andlimbic and paralimbic regions are directed mostly to the 24 are also connected with the temporal proisocorticalagranular and dysgranular sectors of TP [36]. The medial area, as well as the perirhinal and entorhinal regions and


Fig. 6. (continued)

the parahippocampal cortex [6,56]. The mPFC reciprocally limbic, autonomic and motor behavioral outputs in re-connects with the piriform cortex in the rat [20] and sponse to sensory inputs [22,34,35].projects to the parahippocampal cortex in the monkey [53].Thus, the temporal pole is most probably a site wheresensory and limbic inputs converge, sending integrated 4.3. Projection to diencephalonoutputs to mPFC which also sends feedback projections toTP reciprocally. The medial part of PM is connected reciprocally with

IL projects to the lateral capsular subdivision of the PFC including areas 9, 12l, 10, 24, 25, 32 as well as withcentral amygdaloid nucleus, corticomedial amygdaloid TP, rostral superior temporal gyrus and sulcus, amygdalanuclei, medial, anterior subdivision of the cortical- and anterior cingulate cortex. Medial PM with connectionsperiamygdaloid cortex, ventromedial subdivision of L, with dorsomedial PFC, auditory cortical regions of theaccessory basal amygdaloid nucleus and anterior superior temporal gyrus, polymodal processing areas of theamygdaloid area, while PL projects to the lateral capsular superior temporal sulcus and the amygdala might play asubdivision of the central and medial portion of BLmc and role in auditory, auditory-spatial, or other attentionalthe adjacent portions of L in the rat [11,33]. Porrino et al. processes [49]. Projection from mPFC to PM was strong[46] concluded that the ventromedial region of the frontal and indicated that this area of the pulvinar is intimatelycortex receives both direct amygdalo-cortical and indirect related with autonomic and limbic functions in connectionamygdalo-thalamo-cortical input from the amygdala in the with integrated audio–visual information.monkey and Aggleton et al. [1] provided evidence of As mentioned above, Acb has been postulated as a sitereciprocal projections between frontal cortex and of functional interface between the limbic and motoramygdala. An intimate reciprocal connection has also been systems [22,34,35]. The present study revealed that PL andreported between mPFC and the amygdaloid nuclear IL of mPFC projected to VAmc which is under the controlcomplex including periamygdaloid cortex, AB, BL and L of GABA neurons in the reticular part of substantia nigraof the primate [2]. [28] and is intimately related to the motor system and

The shell of Acb received projections primarily from IL projects to PFC including PL [8]. We suggest that VAmc isand PL of mPFC in the monkey. Projections from the an another site of interaction between the motor andventral striatum are represented topographically in the autonomic / limbic control systems of the cortico-basalventral pallidum and non-topographically in the substantia ganglia-thalamus-cortical loops [39]. Ventral precallosalnigra, pars compacta [26]. Projections to the shell predomi- area 14 and subcallosal area 25 and the ventral, subcallosalnated from IL rather than PL, and the core of Acb received part of area 32 receive projections from the mediodorsalmore projections from PL than IL in the present study, portion of MDmc and caudodorsal part of MD [47]. Thesupporting the previous suggestion that the shell is related dorsal, precallosal part of PL receives projections from theto autonomic and limbic functions and the core to motor dorsal portion of MDpc. Area 24 receives additional inputcontrol; further, the core is a kind of interface between from the anterior medial nucleus and midline thalamic



nuclei [6]. TP projects to the ventral part of the mid- and also has reciprocal connections with the amygdala androstrocaudal level of MDmc in the monkey [50]. ‘hypothalamic area controlling emotional responses’, cor-

¨ ¨Ongur et al. [43,44] examined projections from areas 25, responding with PfH and the most medial portion of LH32 and 24b to the hypothalamus in Macaque monkeys. and could be regarded as a center controlling the car-They found heavy projections from area 25 to POL, POM, diovascular responses accompanying emotion [52]. His-VMH, LH, PH, zona incerta (ZI) and SI. They observed taminergic neurons of TM receive inputs from PFC [23],moderate to weak projections from area 32 to the anterior preoptic area and septum, and project to almost all parts ofhypothalamic area, LH, DMH, VMH and PH as well as the brain; they may transmit information originating fromfrom 24b to POM, LH and PH. The present study the limbic system to most parts of the brain, releaseconfirmed similar projections and observed dense nerve histamine non-synaptically, and regulate the activities ofterminals in TM. Ventromedial PFC projects to LH and ZI widely divergent regions of the brain [58].


Fig. 7. (continued)

4.4. Projection to mesencephalon midbrain aversive system, and stimulation in this areaproduces intense fear with autonomic activation in man

DA inhibits pyramidal neurons of IL and PL of mPFC [40,61] and aversive behavioral responses with sympatho-that project to subcortical targets through D DA receptor activation in animals ([12] for reviews, [30,31]).2

[51], and in addition to direct inhibition of cortical IL, insular cortex, BST, perifornical region, TM, dorsalprojection neurons, DA inhibits pyramidal cells indirectly hypothalamic area,VMH, PVH, POL,M, ZI, raphe nucleus,

¨by augmenting GABA release from interneurons [48]. cuneiform nucleus, Kolliker–Fuse nucleus, PBl, PBm,An et al. [3] reported that PFC projected densely to the NTS and ventrolateral part of PAG project to corticotropin-

periaqueductal gray (PAG) of Macaque monkeys, and that releasing hormone-rich neurons of the pontine micturitionareas 25, 32 and 10m projected predominantly to the center, known as Barrington’s nucleus in the rat [55].bilateral dorsolateral columns of PAG. The present resultswere similar, showing dense bilateral projections to PAGdl 4.5. Projection to medulla and spinal cordwith ipsilateral predominance. PFC projects to PAG whichin turn projects to intermedio-lateral nucleus of the spinal Neurons in the rostral ventrolateral medulla area projectcord [52]. Layers V and VI of IL and PL project to PAGdl, to IL of the rat [62]. IL and PL projected to NTS, dorsala visceral motor area of midbrain in the rat [63]. The motor nucleus of vagus, nucleus ambiguus, and thelateral and dorsolateral region of PAG is considered as a ventrolateral medulla as well as lamina I and intermedio-


Fig. 8. A schematic diagram summarizing the results of the present study showing nuclei which received projections from the infralimbic area (IL, area25) of the medial prefrontal cortex.

lateral nucleus of the spinal cord in the rat [27]. The that IL and PL projected indirectly to the spinal cord in thevasomotor center in the mPFC of the rat corresponded to monkey.the PL by Krettek and Price [29] or Cg3 by Paxinos andWatson [45] projecting bilaterally and directly to the 4.6. Structure and function of mPFCcentral autonomic area of the thoracic spinal cord throughthe dorsal cortico spinal tract [5]. The cytoarchitectonic and histochemical atlas of the

As a control study to reveal projections to the spinal prefrontal cortex of the monkey has clearly identified areascord from IL or PL, we injected CTb into the caudal 25, 32 and 24 of mPFC [14]. Gabbott and Bacon [24,25]medulla oblongata of the monkey (unpublished data). We further showed histochemical characteristics in relation tocould observe retrogradely labeled neuronal perikarya in calcium binding proteins, GABA and nitric oxide in mPFCthe cingulate cortex, area 14, and latero–dorsal aspect of and differentiated IL, PL and anterior cingulate cortices ofthe frontal cortex. Many labeled neurons were also found the monkey.in PPN, pretectal nucleus, DR, LC, interstitial nucleus of The PL in the rat receives a direct input from theCajal, Darkschewitch nucleus, nucleus of Bechterew, hippocampus, a connection that is essential for spatialnucleus subfascicularis, ZI, substantia nigra, central memory, and Group 1 mGluRs are present on layer Vamygdaloid nucleus, PVH and LH. We injected WGA– neurons of PL participating in the production of long-termHRP into NTS of the medulla oblongata (unpublished data) potentiation [57]. mPFC of the rat has been implicated asand found retrogradely labeled neurons in the cingulate an autonomic motor area by many investigators mor-cortex, opercular cortex, and supplementary motor area. phologically [5,16,27,41,54] and functionally [13,41,42].Many labeled neurons were found in the hypothalamus Electrophysiological stimulation of the gyrus orbitalis(LH, DA, PfH, PVH), central nucleus of the amygdala, corresponding to area 13m, 13l, Iam, Iai, Iapm of Carmich-Acb, BST, ZI, DR, pretectal nucleus, LC and PBm. We ael and Price [14], which corresponds to the agranularcould observe only a few labeled terminals in NTS after insular cortex of the rat [60], resulted in inhibition ofBDA injection into mPFC and, further, we could find few respiration, rise of blood pressure and decrease in the tonuslabeled perikarya in IL and PL after injection of retrograde of the gastric musculature of the monkey, Macaca mulattaneuronal tracers into the medulla oblongata, and concluded [7]. Although no functional studies have been performed


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