Genomic–anatomic evidence for distinct functionaldomains in hippocampal field CA1Hong-Wei Donga,1, Larry W. Swansonb, Lin Chenc, Michael S. Fanselowd, and Arthur W. Togaa

aLaboratory of Neuro Imaging and Department of Neurology, School of Medicine, University of California, Los Angeles, CA 90095-7334; bDepartmentof Biological Sciences, University of Southern California, Los Angeles, CA 90089; cDepartment of Genomic Science, University of Washington,Seattle, WA 98195; and dDepartment of Psychology and Brain Institute, University of California, Los Angeles, CA 90095-1563

Edited by Edward G. Jones, University of California, Davis, CA, and approved May 8, 2009 (received for review December 15, 2008)

Functional heterogeneity has been investigated for decades in thehippocampal region of the mammalian cerebral cortex, and evi-dence for vaguely defined ‘‘dorsal’’ and ‘‘ventral’’ regions is emerg-ing. Direct evidence that hippocampal field CA1 displays clearregional, laminar, and pyramidal neuron differentiation is pre-sented here, based on a systematic high-resolution analysis of apublicly accessible, genome-wide expression digital library (AllenBrain Atlas) [Lein et al. (2007) Genome-wide atlas of gene expres-sion in the adult mouse brain. Nature 445:168–176]. First, geneticmarkers reveal distinct spatial expression domains and subdo-mains along the longitudinal (dorsal/septal/posterior to ventral/temporal/anterior) axis of field CA1. Second, genetic markersdivide field CA1 pyramidal neurons into multiple subtypes withcharacteristic laminar distributions. And third, subcortical brainregions receiving axonal projections from molecularly distinctspatial domains of field CA1 display distinct global gene expressionpatterns, suggesting that field CA1 spatial domains may be genet-ically wired independently to form distinct functional networksrelated to cognition and emotion. Insights emerging from thisgenomic–anatomic approach provide a starting point for a de-tailed analysis of differential hippocampal structure–functionorganization.

genetics � genomics � hippocampus � learning and memory �neuroanatomy

The basic outlines of hippocampal architecture were establishedby the pioneering work of Ramon y Cajal (1) and his student

Lorente de No (2), who named field CA3 (with large pyramidalneurons and mossy fibers), field CA2 (with large pyramids but nomossy fibers), and field CA1 (with small pyramids). The hippocam-pus and other interconnected medial temporal lobe cortical areashave been known for some time to be critical for learning andmemory (3), and more recently the fact has become clear that otherfunctions are subserved as well (4–6). Overall, the evidence sug-gests that the ‘‘dorsal’’ (septal or in humans posterior) hippocampusis involved in navigation and related spatial memory, whereas incontrast the ‘‘ventral’’ (temporal or in humans anterior) hippocam-pus influences stress responses and motivated or emotional behav-iors. Experimental neuroanatomical work has established in ratsand primates that different transverse levels along the dorsoventralor longitudinal axis establish extrinsic axonal connections in a highlydifferentiated and topographic way (7–11). Nevertheless, the exactspatial arrangement of the dorsal and ventral hippocampus has not,until now, been clearly established. Here, we directly parcel fieldCA1 into multiple, spatially distinct molecular domains and sub-domains using robust gene markers selected from a comprehensivedigital gene expression library (Allen Brain Atlas [ABA], www.brain-map.org) (12) combined with hippocampal cytoarchitectonicanalysis. The evidence indicates that field CA1 is not a homogenouscortical area but instead displays clear regional and laminar spec-ificity. These basic structural insights immediately suggest genetic,physiological, and behavioral experiments to clarify exactly howthese molecular expression domains are differentially involved in

two of the most complex cerebral functions—cognition andemotion.

ResultsMore than 4,000 genes are expressed in the hippocampal for-mation, based on data on the ABA web site. We first screenedexpression patterns of �2,000 genes in the hippocampal region,based on expression level, expression density, and gene cluster-ing. From this, we carefully analyzed expression patterns of 48genes in the category field CA1 pyramidal layer and 400 ‘‘return’’genes when we interrogated points in dorsal, intermediate,ventral, and ventral tip parts of field CA1 (Fig. 1 A–C). Althoughthese genes display very heterogeneous expression patterns, wefound that many display consistent regional specificities in fieldCA1. Using these genes as molecular spatial markers, we havedivided field CA1 into three distinct domains along the dorso-ventral or longitudinal axis (Fig. 1D and Fig. S1): dorsal (CA1d),intermediate (CA1i), and ventral (CA1v) domains. They arevisualized simultaneously in transverse mouse brain levels be-tween 2.7 and 2.9 mm caudal to bregma where the maximaldorsoventral extent of field CA1 is displayed (Fig. 1 D–F and Fig.S1B). At these levels CA1d, CA1i, and CA1v occupy approxi-mately the dorsal, intermediate, and ventral thirds of field CA1,respectively. The CA1d–CA1i border here is parallel to theventral edge of the dentate gyrus lateral blade, whereas theCA1i–CA1v border is approximately level with the dorsal edgeof the rhinal fissure (Fig. 1D and Fig. S1B). Rostral and dorsalto these transverse levels CA1d extends to the rostral (septal) tipof field CA1 (Fig. 1E and Fig. S1 A), whereas caudally domainCA1d is gradually replaced by pyramidal neurons of the dorsalsubiculum (Fig. S1 C and D), which is distinguished from domainCA1d by lack of a stratum oriens. In contrast, domain CA1vextends rostrally and ventrally for only a short distance (Fig. 1E).Dorsolaterally domain CA1v is progressively displaced by theventral subiculum, until it merges with domain CA1i at thecaudal end of field CA1 (Fig. 1E and Fig. S1 C and D).

The extent of domain CA1d is uniquely defined by strongexpression of Wfs1 (Figs. 1 D–F and 2 and Fig. S1 A–D) and anumber of other gene markers including Nov, Kcnh7, Ndst4, and2610017I09Rik, whereas domain CA1v is defined by strong andunique expression of many other gene markers, including Dcn,Grp, Htr2c, Col5a1, and Gpc3 (Figs. 1 D–F, 2, and 3 and Fig. S1A–D). (The full names of these genes and image series numbersof their corresponding full sets of gene expression images in theABA are listed in Table S1.) The middle third of field CA1(CA1i) is characterized most obviously by the absence or much

Author contributions: H.-W.D., L.W.S., M.S.F., and A.W.T. designed research; H.-W.D. andL.C. performed research; H.-W.D. analyzed data; and H.-W.D., L.W.S., L.C., M.S.F., andA.W.T. wrote the paper.

The authors declare no conflict of interest.

This article is a PNAS Direct Submission.

1To whom correspondence should be addressed. E-mail: hongwei.dong@loni.ucla.edu.

This article contains supporting information online at www.pnas.org/cgi/content/full/0812608106/DCSupplemental.

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weaker expression of the above markers (Fig. 1 D–F and Fig. S1A–D). However, many genes have expression patterns that helpto distinguish domain CA1i more directly. For example, Zbtb20and Gpt2 are expressed in both domains CA1d and CA1i but notin CA1v, with Zbtb20 expressed more strongly in CA1i thanCA1d (Fig. S1 B–D) and the reverse occurring for Gpt2. Offundamental importance, although the expression of many genesin field CA1 may not be restricted to CA1d, CA1i, or CA1v, theexpression does consistently respect one of the boundariesbetween them. Because genes expressed in domain CA1i aremore commonly also expressed in CA1d but not CA1v, CA1i ispossibly simply a subdomain of CA1d.

The concept of subdomains is an important one, especially indomain CA1v where four are obvious (Figs. 1D and 2 and Fig.S1B): dorsolateral (CA1vd), intermediodorsal (CA1vid), inter-medioventral (CA1viv), and ventromedial tip (CA1vv). The tinyCA1vv is very distinctive because of weak Grp expression (Figs.1D and 2 and Fig. S1 B–D) and much stronger, localized Htr2cand Col5a1 expression (Figs. 1 D–F and 2 and Fig. S1 B–D). Inaddition, a number of other genes such as Gpr101 (Fig. 2) andDlk1are expressed almost exclusively in CA1vv. Genes expressedin the other three subdomains display more or less overlappingor gradient-type patterns. For example, Tc1568100 andLoc432748 are weakly expressed in subdomain CA1vv, areheavily expressed in CA1viv, and become progressively morerestricted to a single sublayer of pyramidal neurons in CA1vid

and then CA1vd (Fig. 2). Many other genes, including Zdhhc7and Pole4, are expressed progressively more strongly in CA1vid,CA1vd, and even CA1i—but not in CA1viv and CA1vv.

The size and shape of field CA1 pyramidal neurons anddifferences with field CA3 pyramids are firmly established (1, 2),and boundaries between these two hippocampal cortical areasare clearly visualized with molecular markers (13, 14). However,these pyramidal neuron populations have long been consideredrelatively uniform within an area. The expression pattern het-erogeneity described above next led us to reexamine carefullyfield CA1 cytoarchitecture based on high-resolution Nissl-stained images presented in the Allen Reference Atlas (ARA)(15). We found that field CA1 cytoarchitecture is clearly heter-ogeneous both regionally and with respect to pyramidal neuronlayers (Figs. 1D and 3 and Fig. S1 A–D). Following Swanson inrats (16), we first divided the pyramidal layer into superficial anddeep sublayers. The cytoarchitecture of domain CA1d is char-acterized by a remarkably darkly stained, densely packed super-ficial pyramidal layer (CA1d-sps) and a thin deep layer withmany fewer pyramids (CA1d-spd; Fig. 1D1 and Fig. S1 A–D), asoriginally described by Lorente de No (2). The CA1d-spd isconsiderably thinner rostrally, with mostly only one row ofpyramids (Fig. S1 A and B), but progressively thickens caudallyinto 3 or 4 rows of loosely arranged pyramids (Fig. S1 C and D).Nevertheless, expression patterns for most genes (such as Novand Ndst4) are very consistent from rostral to caudal in trans-

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Fig. 1. (A–C) Overall gene expression ‘‘heat maps’’for ‘‘seed voxel’’ points in dorsal, intermediate, ventraldomains of field CA1. (D) One transverse level of theARA (15) displaying the maximal dorsoventral scope offield CA1 and its corresponding Nissl-stained histolog-ical images. Distributions of five marker genes, Wfs1(blue), Zbtb20 (green), Dcn (purple), Htr2c (red), andGrp (yellow) are plotted on this level to reveal threemolecular domains of field CA1 (CA1d, CA1i, andCA1v). (D1, D2, D3) High-resolution Nissl images thatcontain CA1d, CA1i, and CA1v, respectively (see textfor details). (E) A three-dimensional model of Am-mon’s horn (in the context of the whole mouse brain).The overall shape of CA1 and its molecular domainsrevealed by CA1 gene markers Wfs1 (blue), Dcn (pur-ple), and Htr2c (red) occupy the outside surface of theC-shaped cylinder of Ammon’s horn, with the dorsalend (CA1d) extending much more rostral than theventral end (CA1v). These two domains merge at thecaudalmost end of CA1. (F) Three-dimensional expres-sion patterns of 4 representative genes, Wfs1 (blue),Grp (yellow), Dcn (purple), and Htr2c (red), in onetransverse plane of field CA1. These genes show dis-tinct regional specificities and clearly define the spatialextent of CA1d (Wfs1), CA1v (Grp, Htr2c, and Dcn), andCA1i (revealed here by lack of these gene markers).Three-dimensional images of CA1 were generated inBrainExplore (http://www.brain-map.org), one three-dimensional model of the ARA (15, 40). See Fig. S1 formore detailed mapping of these genes in 4 represen-tative levels of the ARA. CTX, cerebral cortex; DGlb,dentate gyrus, lateral blade; ENTl, lateral entorhinalarea; PERI, perirhinal cortical area. (Scale bars: D, 1,310�m; D1, D2, D3, 212 �m to match the same magnifi-cations of the same images on ABA web site.)

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verse sections of field CA1d, although a few genes (including Lct)show much stronger expression in rostral levels of CA1d than inmore caudal levels, and a few others (including Kit) present thereverse gradient. Wfs1 and Nr4a1 are expressed in both CA1d-sps and CA1d-spd sublayers, Nov and Nr3c2 tend to express morespecifically in CA1d-sps, and Ndst4 and Astn2 are expressedpreferentially in CA1d-spd (Fig. 3). Some neurons in the stratumoriens also express Astn2. Compared with domain CA1d, thesuperficial pyramidal layer of CA1i (CA1i-sps) is much lessdense and more lightly stained, whereas the deep layer (CA1i-spd) is relatively thicker with 3 or 4 rows of pyramids (Fig. 1D2and Fig. S1B).

CA1v displays even more complex cytoarchitecture. In addi-tion to superficial (CA1v-sps) and deep (CA1v-spd) pyramidallayers, a middle sublayer (CA1v-spm) containing smaller pyra-mids appears to be sandwiched between the other layers insubdomains CA1vd and CA1vid (Fig. 1D3 and Fig. S1B). In theprogressively more ventral CA1viv, the middle pyramidal sub-layer disappears, and pyramidal neuron morphology becomesless distinguishable between the superficial and the deep layers(Fig. 1 D and D3 and Fig. S1B). On reaching subdomainCA1vv—the ventral tip of field CA1—pyramids align in anapparently uniform, evenly stained, thick layer of approximately7–9 neuronal rows (Fig. 1D and Fig. S1B). This cytoarchitectonicsublamination pattern in CA1v is substantiated by gene expres-sion patterns. First, Grp is expressed strongly in both CA1v-sps

and CA1v-spd but weakly in CA1v-spm (Figs. 2 and 3 and Fig.S1B); Dcn is expressed strongly in CA1v-sps but weakly inCA1v-spd and CA1v-spm (Fig. 3 and Fig. S1 B–D), whereasHtr2c and Gpc3 are strongly expressed in CA1vv, but only in thedeep layer of CA1viv, CA1vid, and CA1vd (Figs. 1 D–F and 3 andFig. S1 B–D). Second, Tc1568100 and Loc432748 are expressedstrongly in the CA1viv-sps, but only the CA1vid-spm and CA1vd-spm (Fig. 2), whereas Prss23 is expressed specifically in the deeplayer of CA1viv, CA1vid, and CA1vd and in CA1i (Fig. 3). Andthird, Prss12 is expressed specifically only in the striatum oriensof domain CA1v (Fig. 3). Interestingly, some genes displaylaminar differences in different domains. For example, Calb1 isexpressed selectively in the superficial layer of CA1d and CA1vivbut is expressed more strongly in the deep layer of CA1i, CA1vd,and CA1vid (Fig. 3).

Next, we examined global spatial gene expression patterncorrelations between these field CA1 molecular domains andother brain regions (Fig. 4; all correlation coefficients providedin Table S2). We found that domain CA1d exhibits a high levelof intrastructural correlation between different transverseplanes of CA1d, but lower correlations with domain CA1v,confirming their molecular specificities across the dorsoventralrather than the rostrocaudal axis. As expected from earlierresults (above), domain CA1i expression patterns are correlatedmore with those of CA1d than with CA1v. Most interestingly,probabilistic coexpression correlations of domains CA1d and

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Fig. 2. Digital images of six representative genes expressed specifically in three molecular domains of field CA1: CA1d (Wfs1), CA1i (Zbtb20), CA1v (Grp), andthe CA1v subdomains, CA1vd, CA1vid, and CA1viv (Col5a1 and Tc1568100) and CA1vv (Gpr101 and Col5a1) at two different rostrocaudal levels (A, B) of field CA1.CA1v subdomains are labeled: a (CA1vd), b (CA1vid), c (CA1viv), and d (CA1vv). Images in (C) are gene expression heat maps of corresponding images in (B) (fordefinition of heat map, see ABA web site, http://www.brain-map.org) that display relative intensities of gene expression signals. All images were downloadedfrom the ABA web site. (Gene names and the image series numbers of their original digital images are listed in Table S1.) Overall expression patterns of Wfs1,Zbtb20, and Grp in the hippocampus are schematically mapped in more detail in Fig. S1. All images in the same panels have the same magnifications. Scale barsare displayed in the lower right corner of Gpr101 images.

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Fig. 3. Gene markers showing regional and laminarspecificities of field CA1 pyramidal neurons. For high-resolution Nissl-stained cytoarchitecture and delineationof pyramidal neuron sublayers, see Fig. 1 and Fig. S1.CA1v subdomains are labeled: a (CA1vd), b (CA1vid), c(CA1viv), and d (CA1vv). All digital images were down-loaded from the ABA gene expression library. (Genenames and the image series numbers of their originaldigital images are listed in Table S1.) Images in the samerows have the same magnifications.

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CA1v with other brain regions are very distinctive. For example,although both domains exhibit very high correlation values withthe cortical mantle as a whole, domain CA1v shows highercorrelation values than CA1d with cortical amygdalar areas andthe basolateral amygdalar complex, both of which share directbidirectional axonal connections with CA1v (10, 11, 17, 18). Interms of subcortical regions, CA1v displays strikingly highercoexpression correlations than CA1d with neuron populationsassociated with autonomic, neuroendocrine, and affective be-haviors, including the central and medial amygdalar nuclei,intermediate and ventral parts of the lateral septal nucleus, bednuclei of the stria terminalis, and hypothalamus—all of whichshare massive direct and indirect neuronal connectivity withCA1v (9, 10, 17–19). These obvious correlations strongly suggestthat CA1d and CA1v are genetically wired independently withdifferent functional specificities.

Finally, Risold and Swanson (9, 16) proposed the existence ofa series of structure–function domains along the longitudinalaxis of field CA1 and adjacent subiculum that send topograph-ically organized projections to the three major parts of the lateralseptal nucleus that in turn share bidirectional connections with

hypothalamic structures mediating the expression of differentclasses of innate motivated behavior. To explore potentialgenetic mechanisms underlying this topographic wiring pattern,we compared coexpression patterns of CA1 domain-specificgenes with those of the lateral septal nucleus. We found that anumber of CA1 domain-specific genes are also expressed in thelateral septal nucleus and that their coexpression correlationsrespect the topologically ordered dorsal-to-dorsal and ventral-to-ventral organization of their axonal projection connectivity.For example, genes that show strong expression in domain CA1d,such as Wfs1 and Matn2, are also strongly expressed in the caudalend of the caudal and rostral lateral septal nucleus (LSc, LSr),which are topologically dorsal (Fig. 4D). In contrast, Dlk1 andGpr101—the CA1vv (and ventral subicular) marker genes—alsoshow strong expression patterns in the ventral lateral septalnucleus (LSv). Genes that show significant expression in moredorsal subdomains of CA1v are most likely also to coexpress inventral levels of the LSr. Thus, Loc432748, a specific markergene for CA1viv, presents specific signal only in a specific zoneof the LSr. Gpc3 and Htr2c, which are strongly expressed inCA1vv and the rest of domain CA1v, are strongly expressed in

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Fig. 4. Global gene coexpression ‘‘energy’’maps result from setting seed voxel points indomain CA1d (A), CA1i (B), and CA1v (C) atthree representative transverse levels of themouse brain at approximately bregma �2.739(a, b, c), �1.656 (a�, b�, c�), and 0.029 (a�, b�, c�).The heat map energy definition is shown at thebottom. The coexpression correlation coeffi-cients of these CA1 molecular domains withother brain structures are listed in Table S2. Fora detailed description of AGEA spatial geneexpression correlation map, see Ng et al. (24).(D and E) Correlated gene expression patternsof domain CA1d marker gene Matn2 and CA1vmarker gene Gpc3 in the lateral septal nucleus.(Scale bars: D and E, 421 �m.)

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the LSv and LSr but not in LSc (Fig. 4E). Man1a and Zdhhc7,which are expressed throughout field CA1 (except in CA1vv),express strongly in the LSc and LSr (but not LSv). By comparisonof the topology and topography of these gene expression cor-relations with the hippocampal functional domains defined byRisold and Swanson in rat (9, 16), CA1vv defined here appearsto correspond well to their Domain 5—the ventral tip of fieldCA1 and subiculum, which sends projections specifically to theLSv (9)—one of the primary sources of cerebral inputs to thehypothalamic neuroendocrine zone and region controlling feed-ing behavior. The progressively more dorsally located CA1viv,CA1vid, and CA1vd may correspond to their Domains 2–4.These CA1v subdomains send topographic projections todifferent zones of the LSr, which in turn project to thehypothalamic medial column that controls two basic classes ofsocial behavior—reproduction and defense (9). Finally, CA1dand CA1i together may correspond to their Domain 1, whichpresumably innervates LSc and LSr levels that in turn project to thetheta-rhythm-controlling supramammillary nucleus (8). The dens-est outputs of dorsal CA1 have been shown to be to the dorsalsubiculum (11)—the major source of the postcommissural fornixpathway innervating the medial and lateral mammillary nuclei andanterior thalamic complex. As a matter of fact, this dorsal CA1 tosubiculum projection is the major hippocampal component thatprocesses spatial memory (20–22).

DiscussionMany gene markers have been used as reproducible landmarksfor defining nervous system divisions at various stages of devel-opment (23) and in the adult (15). This is an extension of therevolutionary chemical neuroanatomy approach that began inthe 1950s with cholinesterase histochemistry, followed by histo-f luorescence and then immunohistochemistry (see ref. 15). Theresulting chemoarchitectonic data greatly refined classical cyto-architectonics based on Nissl staining. The present study, com-bined with two recent reports (24, 25), supply proof-of-principleexamples of how the genomic–neuroanatomic approach canfurther refine architectonic maps and identify neuronal pheno-types amenable to selective experimental genetic manipulation.Although raw data about these expression patterns (digitizedimages) are freely available on the ABA web site and powerfulonline informatics tools enable users conveniently to identifyregionally specific gene expression patterns, synthesis and inter-pretation of this information relative to existing structural,functional, and behavioral knowledge is still required. Specifi-cally, we provide here a testable model to guide and stimulatefurther analysis of hippocampal structure–function in health anddisease.

Our genetic–anatomic analysis and the current axonal con-nectivity literature (7–11, 17–19, 26) suggest that domains CA1dand CA1i lie in approximately the dorsal half of the hippocam-pus, and they display strong global gene expression correlationsprimarily with other parts of cerebral cortex and subcorticalregions innervated by dorsal hippocampus and related to theta-rhythm modulation and navigation. In contrast, domain CA1vlies in approximately the ventral half of the hippocampus, andthis domain shares strong gene coexpression patterns and pro-jections with cerebral cortex and subcortical regions mediatingneuroendocrine, autonomic, and goal-oriented or emotionalbehavioral responses. The natural boundary between CA1i andCA1v in mice coincides with the rhinal fissure’s dorsal border, aboundary shown by certain CA1 domain-specific genes and byother cortical marker genes, including Col5a1, which is stronglyexpressed in isocortical layers 6a and 6b (Fig. 2). The evidencesuggests that the dorsal and ventral halves of field CA1 aregenetically wired independently to support cognitive and emo-tional responses, respectively (4–6).

The model of three major CA1 molecular domains presentedhere shows interesting correlations with the 9 even more strikingmolecular regions recently reported along the longitudinal axisof mouse field CA3 (25). The observations of Thompson et al.,together with our own in fields CA1 and CA3, suggest that fieldCA3—and thus Ammon’s horn as a whole—is also divided intothree major molecular domains: (i) a dorsal domain (CA3d)including their regions 1–3 that is accompanied by domainCA1d; (ii) an intermediate domain (CA3i) including their re-gions 4 and 5 that is accompanied by domain CA1i; and (iii) aventral domain (CA3v) including their regions 6–7 that isaccompanied by domain CA1v. The domains CA3d and CA3i(regions 1–5) form approximately the dorsal half of field CA3,whereas the domain CA3v (regions 6 and 7) forms approximatelythe ventral half.

Ramon y Cajal first noted structural differences along thehippocampal longitudinal axis, related to what he called thesuperior and inferior perforant paths (1) and what would laterbe referred to loosely as the dorsal and ventral hippocampus,respectively (9, 27). Lorente de No (2) vaguely divided the‘‘Ammonic system’’ into three longitudinal segments based onnow discredited connectional data and also divided field CA3and field CA1 into three parallel, longitudinally running zones(a, b, and c for both). Lorente de No’s (2) subfields CA3a, CA3b,and CA3c appear to correspond well with three molecularregions identified by Thompson et al. (25): region 3 (Mas1expression) with CA3a, region 2 (Fmo1 expression) with CA3b,and region 1 (Ttn1 expression) with CA3c. In field CA1, at leastsome genes (such as Lct and Kit) were found here to showdifferential expression patterns across the topological medial tolateral (transverse) axis, but correspondences with Lorente deNo’s subfields CA1a, CA1b, and CA1c are not yet clear.

Extensive work since the early 1970s established that intra-hippocampal circuitry, typified by the classical trisynaptic circuitfrom entorhinal area to dentate gyrus to field CA3 to field CA1,runs transverse to the longitudinal axis and that specific extrinsicinputs and outputs are associated with specific regions along thelongitudinal axis of the hippocampal formation (8–11, 17–19,28–33). The dorsal half of field CA1 has a high concentration ofplace neurons (e.g., ref. 34) and sends most of its extrinsiccortical projections, either directly or indirectly via dorsal sub-iculum, to the cortical retrosplenial area (11, 35), which isinvolved in cognitive processing of visual sensory information atleast in part for spatial and contextual memory. The dorsalsubiculum receives its strongest input from dorsal CA1 and sendsmassive projections to the mammillary and anterior thalamicnuclei, two regions containing abundant navigation neurons(36). These two diencephalic regions in turn project back to thedorsal hippocampus and retrosplenial area (35). This and otherevidence (see refs. 4–6, 20, and 21) suggest that the dorsal halfof field CA1 (domains CA1d and CA1i) is critically involved incognitive spatial memory processing, at least in rodents.

The most distinguishing feature of connectivity associatedwith the ventral half of field CA1 (domain CA1v and its 4subdomains) is dense bidirectional connections with amygdalarregions implicated in emotional behaviors (10, 11, 17, 18).Ventral but not dorsal field CA1 also projects directly (19) to thehypothalamic periventricular region and medial zone, whichintegrate neuroendocrine, autonomic, and somatic motor re-sponses associated with three basic classes of motivated behav-iors common to all animals: ingestive (feeding and drinking),reproductive (sexual and parental), and defensive (fight orflight) (26). These direct descending projections from ventral notdorsal CA1 are reinforced by multiple indirect pathways involv-ing relays in the ventral subiculum, amygdalar region, lateralseptal nucleus, and bed nuclei of the stria terminalis (9–11,17–19, 26, 33, 37), all of which display gene coexpression patternscorrelated with those of CA1v as shown here.

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The observation that the dorsal half and ventral half of fieldCA1 display distinct global gene expression correlations withbrain structures that they project to either directly or indirectlyis striking. Although the mechanistic significance of this rela-tionship between CA1 molecular domains and their extrinsiccircuitry is not yet clear, from the above the dorsal and ventralhalves are obviously genetically wired independently for differ-ent functional specializations. The former is selectively involvedin cognitive aspects of the learning and memory associated withnavigation, exploration, and locomotion, whereas in contrast thelatter is part of the temporal lobe associated most directly withmotivational and emotional aspects of the classic Kluver–BucySyndrome. In fact, at least two CA1v marker genes, Grp andHtr2c, have been implicated in psychiatric disorders (38, 39).

This and similar work (25) are part of a systematic examinationof relationships between gene expression patterns and neuronalnetworks to clarify genetic contributions to brain structure andfunction. However, the detection sensitivity and specificity formany genes, especially those of low abundance, probably werecompromised in the large-scale, high-throughput process ofABA data production (12). This would make the list of domain-specific markers assembled here rather incomplete, and manygenes critical for determining adult brain structure and connec-tions likely are expressed during development not in the adult(23). Even for the domain-specific markers listed here (TableS1), information in Gene Ontology suggests many diverse (andunknown) functions whose possible role in neuronal develop-ment and adult structure–function remain to be investigated.

Relative to isocortex, very little is known about laminar andregional differences in field CA1 pyramidal cell subtypes. Both

Ramon y Cajal (1) and Lorente de No (2) observed deep andsuperficial pyramids and noticed that they establish uniquerelationships with basket cells. However, the addition of multiplegenetic markers for differentiation of field CA1 pyramids opensmany new possibilities for establishing analytical distinctions.

Materials and MethodsThis study was based on a systematic, high-resolution analysis of �4,000 genesexpressed in the hippocampal formation through a publicly accessible onlinegene expression digital library—the ABA. We first manually screened expressionpatterns of �2,000 genes under the Anatomic Search tool category ‘‘hippocam-pal region,’’ based on expression level, expression density, and gene clustering,and carefully analyzed expression patterns of 48 genes in the category ‘‘field CA1pyramidal layer’’ under ‘‘fine structure search.’’ We then used the Gene Findertool of the Anatomic Gene Expression Atlas (AGEA) application (Fig. 1 A–C) tofilter candidate genes showing regional specificities. Detailed methodology ofthe AGEA has been described in a recent report (24). In brief, because all ABAimage-based, in situ hybridization data are spatially registered to the commonanatomic framework of the ARA (15) with a standard coordinate system andhierarchical ontology, the AGEA in principle enables users to view a comprehen-sive list of genes expressed in any given three-dimensional voxel (‘‘seed voxel’’) ofthe mouse brain atlas using the Gene Finder application (24). With this strategy,we carefully analyzed the expression of �400 ‘‘return’’ genes when we set seedvoxel points in dorsal, intermediate, and ventral parts of field CA1. AGEA alsoallowsusers toviewglobalexpressioncorrelationsassociatedwithanyseedvoxel.With this tool, we have also analyzed gene expression correlations of differentdomains of the CA1.

ACKNOWLEDGMENTS. This work was supported in part by National Institutesof Health Grants MH083180 (to H.-W.D.), RR013642 (to A.W.T.), NS16686 (toL.W.S.), and MH62122 (to M.S.F.).

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Dong et al. PNAS � July 14, 2009 � vol. 106 � no. 28 � 11799

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