Journal of Chemical Neuroanatomy 26 (2003) 233–242

Review

Human cholinergic basal forebrain: chemoanatomy andneurologic dysfunction

Elliott J. Mufsona,∗, Stephen D. Ginsbergb, Milos D. Ikonomovicc, Steven T. DeKoskyc

a Department of Neurological Sciences and Alzheimer’s Disease Center, Rush Presbyterian-St. Luke’s Medical Center,Tech 2000, 2242 West Harrison St., Suite 200, Chicago, IL 60612, USA

b Center for Dementia Research, Nathan Kline Institute, Departments of Psychiatry, Physiology and Neuroscience,New York University School of Medicine, Orangeburg, NY, USA

c Departments of Neurology, Psychiatry and Alzheimer’s Disease Research Center, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

Received 2 April 2003; received in revised form 28 June 2003; accepted 28 June 2003

Abstract

The human cholinergic basal forebrain (CBF) is comprised of magnocellular hyperchromic neurons within the septal/diagonal bandcomplex and nucleus basalis (NB) of Meynert. CBF neurons provide the major cholinergic innervation to the hippocampus, amygdalaand neocortex. They play a role in cognition and attentional behaviors, and are dysfunctional in Alzheimer’s disease (AD). The humanCBF displays a continuum of large cells that contain various cholinergic markers, nerve growth factor (NGF) and its cognate receptors,calbindin, glutamate receptors, and the estrogen receptors, ER� and ER�. Admixed with these cholinergic neuronal phenotypes are smallerinterneurons containing the m2 muscarinic acetylcholine receptor (mAChRs), NADPH-diaphorase, GABA, calcium binding proteins andseveral inhibitory neuropeptides including galanin (GAL), which is over expressed in AD. Studies using human autopsy material indicate anage-related dissociation of calbindin and the glutamate receptor GluR2 within CBF neurons, suggesting that these molecules act synergisticallyto induce excitotoxic cell death during aging, and possibly during AD. Choline acetyltrasnferease (ChAT) activity and CBF neuron numberis preserved in the cholinergic basocortical system and up regulated in the septohippocampal system during prodromal as compared with endstage AD. In contrast, the number of CBF neurons containing NGF receptors is reduced early in the disease process suggesting a phenotypicsilence and not a frank loss of neurons. In end stage AD, there is a selective reduction in trkA mRNA but not p75NTR in single CBF cellssuggesting a neurotrophic defect throughout the progression of AD. These observations indicate the complexity of the chemoanatomy of thehuman CBF and suggest that multiple factors play different roles in its dysfunction in aging and AD.© 2003 Published by Elsevier B.V.

Keywords: Human cholinergic basal forebrain; Chemoanatomy; Neurologic dysfunction

1. Introduction

A diverse group of telencephalic structures situated onthe medial and ventral aspects of the cerebral hemispherestogether compose the basal forebrain in the primate brain.This complex region contains structures including the septalarea, the vertical and the horizontal limbs of the diagonalband of Broca as well as the region termed the substantiainnominata (for review seeMufson and Kordower, 2000).Historically, this later region has been termed the nucleusof the ansa lenticularis (Meynert, 1872) and later renamed

∗ Corresponding author. Tel.:+1-312-563-3570;fax: +1-312-563-3571.

E-mail address: emufson@rush.edu (E.J. Mufson).

the nucleus basalis (NB;Kolliker, 1896). Part of the dif-ficulty in understanding this region is the complexity ofits chemoanatomical organization. These regions also havea diversity of cells displaying different neurotransmitters,morphologies and projection patterns (seede Lacalle et al.,1994; Mufson and Kordower, 2000for reviews). Thesecholinergic corticopetal projection neurons have receivedextensive attention due to their reduction in Alzheimer’s dis-ease (AD) and their close association with the neurotrophicsubstance nerve growth factor (NGF) and its high (trkA) andlow (p75NTR) affinity receptors which are involved in basalforebrain cell survival (seeMufson and Kordower, 1999;Mufson et al., 1997for reviews). This article will discuss thecentral cholinergic projection neurons of the human basalforebrain and their dysfunction in normal aging and in AD.

0891-0618/$ – see front matter © 2003 Published by Elsevier B.V.doi:10.1016/S0891-0618(03)00068-1

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2. Materials and methods

2.1. Subjects

The human brain material discussed in this compilationreport was harvested from several brain banks includingRush Presbyterian St. Luke’s Medical Center, Harvard Med-ical School Alzheimer’s Disease Center, University of Pitts-burgh Alzheimer’s Disease Center and Sun City ResearchInstitute. In general, brains are removed and soaked in 4%paraformaldehyde and cryoprotected (Mufson et al., 1989c,1998) overnight and cut at 40�m thickness on a freez-ing sliding microtome. Material used for biochemical assaywas snap frozen in liquid nitrogen. Pathological designa-tions were based on the CERAD Criteria (Mirra et al., 1991),Braak scoring (Braak and Braak, 1991), and NIA-ReaganDiagnostic Criteria (NIA, 1997). Clinical evaluation of theROS and University of Pittsburgh ADRC populations havebeen published elsewhere (Bennett et al., 1997, 1999, 2002;DeKosky et al., 2002; Gilmor et al., 1999; Kordower et al.,2001; Mufson et al., 1999a, 2000b, 2002b).

3. Immunocytochemistry

Immunohistochemistry was performed for each antibodyaccording to previously reported procedures (for cholineacetyltrasnferease (ChAT) (seeMufson et al., 1989c); fortrkA (see Mufson et al., 1997); for p75NTR (seeMufsonet al., 2002b); for galanin (GAL) (seeMufson et al., 1993);for m2 receptor (seeMufson et al., 1998); for calbindin (seeGeula et al., 2003); for glutamate receptors (seeIkonomovicet al., 2000)). ER� (Zymed Labs, San Francisco, CA) stain-ing was performed at a dilution of 1:1000 over night at roomtemperature.

4. Choline acetyltransferase activity

ChAT activity was measured from snap frozen tissue sam-ples obtained from the anterior cingulate, superior frontal,superior temporal cortex, and hippocampus using radioac-tive carbon-14 labeled acetyl Co-A (New England Nuclear,Boston, MA) (seeFonnum, 1975). Protein content was de-termined by a BCA protein assay kit (Pierce, Rockford, IL).Calculations were based on net counts per minute (CPM) ofsamples multiplied by specific activity of labeled substrateand divided by protein concentration to yield ChAT activityin �mol/h per g

¯protein.

5. Single cell RNA amplification

Procedures for single cell micro aspiration of NB neu-rons, RNA amplification and analysis used fixed tissue asdescribed previously (Ginsberg and Che, 2002; Ginsberg

et al., 1999; Hemby et al., 2003; Mufson et al., 2002a).Briefly, terminal continuations (TC) amplified RNA probesgenerated from individual NB cholinergic basal forebrain(CBF) neurons were hybridized to custom-designed cDNAarrays, which consisted of nylon membranes (Hybond XL,Amersham Biosciences, Piscataway, NJ) adhered with cD-NAs (1 �g each) selected to focus on mRNAs that encodeproteins implicated in the degeneration of NB neurons dur-ing AD. TC hybridization signal intensity is detected byphosphor imaging. Specific signal intensity of TC amplifiedRNA bound to each linearized cDNA is expressed as a ratioof the total hybridization signal intensity of the array (seefor detailsGinsberg and Che, 2002; Ginsberg et al., 1999;Hemby et al., 2003).

6. Quantitative stereology

Total neuron counts were derived using an unbiased, stere-ologic cell counting method (seeKordower et al., 2001;Mufson et al., 2000b; West et al., 1996). The total number ofneurons was calculated by using the formula N= NV × VSN,where NV is the numerical density and VSN is the volumethe NB (West et al., 1996). Variability within groups was as-sessed by mean of the coefficient of error (CE:West et al.,1996).

7. Anatomy of the human cholinergic basalforebrain subgroups

Nissl stained preparations reveal that the main cell groupsof the CBF exhibit a distinct magnocellular appearance(Fig. 1A). Studies using the non-specific cholinergic markeracetylcholinesterase (AChE), the synthesizing enzymefor acetycholine, ChAT, the vesicular acetylcholine trans-porter, and the high-affinity choline transporter have delim-ited a continuum of cholinergic neurons within the CBF(Fig. 1B–D and G; seeGilmor et al., 1999; Kus et al., 2003;Mesulam et al., 1983, 1984, 1986; Mufson et al., 1989b).Cholinergic neurons (25–30× 25–30�m) of the medial sep-tum (Ch1) compose 10% of this region. The diagonal bandof Broca comprises the Ch2 (vertical limb;Fig. 1D) andCh3 (horizontal limb) subgroups. Ch2 neurons are fusiformin shape, 20–25× 30–40�m in size and approximately 70%of these perikarya are cholinergic. Ch3 has the most diffuseboundaries in the primate brain and contain fusiform shaped(15–20× 40–50 �m) neurons approximately 1–2% ischolinergic. The largest group of cholinergic neurons (90%)constitutes the Ch4 cells of the NB (Ch4;Fig. 1B, G), whichare fusiform to multipolar in shape and 40–50× 60–70�min size. Topographically Ch4 can be separated into anterior(Ch4a), intermediate (Ch4i) and posterior (Ch4p) subfields.The anterior portion can be divided into medial (Ch4am)and lateral (Ch4al) sectors while the intermediate subfieldis divided into dorsal (Ch4id) and ventral (Ch4iv) regions

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(Fig. 1G). The posterior subfield (Ch4p) is located up to thelevel of the lateral geniculate nucleus. The total number ofcholinergic neurons located within the NB is approximately210 000 per hemisphere (Arendt et al., 1985; Gilmor et al.,1999). These CBF neurons provide the major cholinergicinnervation to the hippocampus (Ch1-2); olfactory system(Ch3): the amygdala and cortical mantle (Ch4) (Fig. 1F;Kitt et al., 1987; Mesulam et al., 1983; Selden et al., 1998;Mesulam and Geula, 1988).

8. NGF, estrogen receptors, gabaergic andglutaminergic markers in CBF neurons nerve growthfactor receptors

Immunocytochemical and in situ hybridization studieshave shown that the high (trkA) and low (p75NTR) affinityNGF receptors as well as retrogradely transported NGFare localized within human CBF neurons (Fig. 1B–E;Schatteman et al., 1988; Strada et al., 1992; Boissiere et al.,1997; Chu et al., 2001; Hefti, 1983, 1986; Hefti and Mash,1989; Kordower et al., 1988, 1989a,b; Mufson et al., 1989b,1999b, 1997, 1996, 2000b). In humans, virtually all (>95%)p75NTR containing CBF neurons contain ChAT (Mesulamet al., 1983; Mufson et al., 1989b). The functional im-portance of these proteins will be discussed below in thesection dealing with AD.

9. Calcium binding proteins

Calbindin and ChAT colocalize within human CBF neu-rons of the NB (Ch4) (Wu et al., 1997; de Lacalle andSaper, 1997; Geula et al., 2003, 1993). Geula et al. (2003,1993)have shown an age-related loss of calbindin stainedhuman NB neurons (compareFig. 1H and I) suggesting aloss of the phenotypic expression of calbindin in CBF neu-rons which may be a potential mechanism for the selectiveloss of cholinergic neurons in aging and AD (Geula et al.,2003).

10. Glutamate receptors

Co-localization experiments reveal that ChAT- andp75NTR-positive CBF neurons express glutamate recep-tor 1 (GluR1) in aged humans (Fig. 1J; Ikonomovic andArmstrong, 1996; Ikonomovic et al., 2000). In contrast,the GluR2 subunit appears to undergo an age-relateddown-regulation within the CBF of elderly humans (Fig. 1K,L; Ikonomovic and Armstrong, 1996; Ikonomovic et al.,2000). It is interesting to note that the presence of the GluR1subunit, and the absence of GluR2, is associated with in-creased Ca2+ influx, which over time affects the abilityof a neuron to regulate intracellular calcium homeostasis.Therefore, a loss of the GluR2 subunit (Ikonomovic et al.,

2000) can lead to a greater Ca2+ influx and increasedsusceptibility to glutamate toxicity, resulting in neuronaldegeneration in “vulnerable” brain regions such as the CBFin aging and AD (for review seeArmstrong et al., 1998;de Lacalle et al., 1991; Lowes-Hummel et al., 1989; Mannet al., 1984; McGeer et al., 1984; Szenborn, 1993). GluR2may be controlled by a switch turning “on” or “off” normalgene expression during aging (Pellegrini-Giampietro et al.,1992). Similar to GluR2, calbindin undergoes a selectiveage and AD related reduction (Wu et al., 1997; Geula et al.,2003). This reduction is found particularly within the morehighly vulnerable posterior CBF subfield (Mufson et al.,1989a; Vogels et al., 1990). We suggest that increased Ca2+influx (due to GluR2 loss) and an impaired ability of thecell to buffer increased intracellular Ca2+ (due to calbindinloss) may act synergistically to induce excitotoxic CBF celldeath during aging and AD.

11. Steroid receptors in the CBF

CBF neurons may be influenced by sex hormones. Andro-gen receptors are present on CBF neurons and their expres-sion is altered during aging (Ishunina and Swaab, 2001). Twoestrogen receptors (ERs) have been identified and termed al-pha (ER�; Koike et al., 1987) and beta (ER�; Kuiper et al.,1996). Immunohistochemical studies indicate that ER� doesnot co-localize with ChAT, p75NTR or trkA containing CBFneurons, although scattered ER� positive nuclei were foundwithin Ch2 and Ch4a in the monkey (Blurton-Jones et al.,1999; Sendera et al., 1999). Recently, the protein and mRNAfor both ERs has been reported in human NB neurons (seeFig. 1M, present report as well asIshunina and Swaab,2001; Osterlund et al., 2000). Epidemiological studies indi-cate that treatment with estrogen decreased the risk of ADin postmenopausal women (Tang et al., 1996) and enhancedthe response to the cholinesterase inhibitor tacrine in theseindividuals (Schneider et al., 1996). However, recent stud-ies have questioned the putative therapeutic effects of estro-gen (Sherwin, 2003). In this light, estrogen plus progestintherapy has been reported to increase the risk for proba-ble dementia while not preventing mild cognitive impair-ment (MCI) in postmenopausal women aged 65 years andolder (Shumaker et al., 2003). The mechanisms underlyingsteroid-receptor action upon neuronal function is an area re-quiring continued investigation.

12. m2 muscarinic receptor neurons within the CBF

Several metabotropic muscarinic acetylcholine recep-tors (mAChRs) have been identified by differential affini-ties for antagonists, and it is established that five distinctgenes m1–m5, encode highly related muscarinic recep-tor subtypes (Bonner et al., 1987; Hulme et al., 1990).It was assumed that m2 subtype of mAChR is the gene

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Fig. 1.

E.J. Mufson et al. / Journal of Chemical Neuroanatomy 26 (2003) 233–242 237

product which functions as a presynaptic autoreceptor toinhibit ACh release (Levey et al., 1991; Mash et al., 1985;Mash and Potter, 1986; Pohorecki et al., 1988) and waslocalized to CBF neurons. In humans, we found that m2receptor protein is expressed primarily in non-cholinergicmultipolar neurons located primarily outside the Ch1-4subfields (Fig. 1N; Mufson et al., 1998). Dual-stained sep-tal/diagonal band sections revealed m2 in only 14% ofChAT-labeled neurons, whereas, 13% of m2-stained neu-rons were co-labeled with ChAT. Within the NB, ChATwas found in 35% of the m2-labeled neurons, whereasonly 6% of ChAT stained perikarya were dual-labeled form2. Co-localization of m2-positive/ChAT-immunonegativeneurons and the cholinergic inhibitory neuropeptide GALwas not observed, although GAL fibers coursed in closeapposition to m2-immunopositive cells. This suggests thegalaninergic fibers may modulate m2 receptor activity inhumans. Cell counts of the relative number of m2 im-munoreactive neurons within the NB from aged controlsand AD patients revealed that the m2 neurons are spared(Mufson et al., 1998).

13. Non-cholinergic neurons within the CBF

In addition to the magnocellular cholinergic neuronslocated within the Ch4 subfields, there are many smallernon-cholinergic perikarya (seede Lacalle and Saper, 1997;Mufson and Kordower, 2000). Although the protein andmRNA for the inhibitory neuropeptide GAL are not foundin human CBF neurons, small GAL-positive interneuronsare admixed within CBF perikarya (Fig. 1O) (Benzinget al., 1993; Chan-Palay, 1988; Kordower et al., 1992;Kordower and Mufson, 1990; Mufson et al., 1993; Walkeret al., 1991). In AD, there is an hyperinnervation of gala-ninergic fibers upon remaining CBF neurons (Fig. 1P) aswell as an increase in galanin receptor expression in thelate stages of AD (Mufson et al., 2000a). These findingssuggest that GAL down-regulates acetycholine production

Fig. 1. Photomicrographs and drawing showing the chemoanatomy of the human NB in the aged and AD brain. (A) Human nucleus basalis Nissl stainedneuron showing the magnocelluar phenotype of CBF perikarya. Scale bar= 50 �m. (B and C) NB neurons immunoreacted for choline acetyltransfereaseand trkA, respectively. Scale bars= 50 �m in B and C. (D) p75NTR immunoreactive vertical limb diagonal band neuron. Scale bar= 60 �m. (E) NBneuron immunoreactive for regrogradely transported NGF. Scale bar= 50 �m. (F) Artist’s drawing showing the cholinergic innervation from the medialsepatl/vertical limb of the diagonal band (MS/VDB; Ch1-2) to the hippocampus (red) and the nucleus basalis of Meynert (NBM) to the cortex andamygdala (blue). (G) Photographs showing the location of the four major cholinergic cell groups (Ch4am; Ch4al, Ch4id and iv and Ch4p) within the NB.Scale bar= 500 �m. (H and I) Concurrent immunostaining of ChAT (brown reaction product) and calbindin (CB; granular blue-black reaction product) inthe Ch4am in a 50-year-old (H). Note that most Ch4am neurons contain ChAT and CB. Panel I from a 93-year-old showing Ch4 ChAT positive neuronsand reduced CB staining. Figures H and I were reproduced with permission from Geula C., Bu J., Nagykery N., Scinto L.F., Chan J., Joseph J., ParkerR., Wu C.K. Loss of calbindin-D28k from aging human cholinergic basal forebrain: relation to neuronal loss 455, 249–259, 2003. Scale bar= 40 �m inH and I. Panels H and I are reproduced with permission fromGeula et al. (2003). (J) Ch4p neurons dual stained for GluR1 and p75NTR, scale bar= 25�m. (K and L). GluR2/3 immunoreactive Ch4i neurons from a 33- and 76-year-old humans. Note the age related reduction in the intensity of staining(arrows). Figures J, K, L were reproduced with permission from Ikonomovic M.D., Nocera R., Mizukami K., Armstrong D.M. Age-related loss of theAMPA receptor subunits GluR2/3 in the human NB of Meynert 166, 363–375, 2000. Scale bars= 40 �m. (M) Estrogen beta immunoreactive nuclei withinCh4 neurons. Scale bar= 25 �m. (N) Section from Ch4 dual stained for m2 (black) and p75NTR. Note the m2 single and the double (m2/p75NTR; blackarrows) neurons. Open arrow shows a single p75NTR lightly immunoreactive neuron. (O) Ch4 section dual stained for galanin (blue) and p75NTR (brown)from an aged brain. Scale bar= 10 �m. (P) Hyperinnervation of galanin (black) fibers) upon a cholinergic (light brown) Ch4 neuron in AD. Abbreviations:AC, anterior commissure; BST, bed nucleus of the stria teriminalis; gpe, external globus pallidus; gpi, internal globus pallidus Scale bar= 80 �m.

in CBF neurons during the late stages of AD (Bowser et al.,1997; Chan-Palay, 1988; Mufson et al., 1993).

NADPH-diaphorase, a putative neurotransmitter/neurotoxin(Dawson et al., 1991), is not found in the monkey (Geulaet al., 1993) or human (Benzing and Mufson 1995; Ellisonet al., 1987) CBF neurons. In AD, there is an increase in thein the number of small non cholinergic NADPH-d neuronssuggesting that this enzyme may play a role in cholinergiccell degeneration via its neurotoxic properties (Benzingand Mufson, 1995). The increase in NADPH-d containingneurons may be related to the re-expression of this enzymeby perikarya that contained this substance during neuronaldevelopment (seeMufson and Kordower, 1992). The pre-cise mechanism underlying this phenomenon remains tobe determined. Other perikarya and fibers immunoreactivefor somatostatin, neuropeptide Y (NPY), enkephalin orneurotensin, pro-opiomelanocortic peptides, substance P,CCK, vasoactive intestinal polypeptide (VIP) and oxytocininterdigitate with cholinergic neurons in the basal forebrain(Candy et al., 1985; Desjardins and Parent 1992; Haber andElde, 1982; Haber and Watson, 1985; Mufson et al.,1988). These observations suggest that peptidergic neuronsmodulate the actions of cholinergic neurons in aging anddisease.

14. CBF dysfunction Alzheimer’s disease

Several reports demonstrate a significant reduction in bothCBF neurons (Whitehouse et al., 1982) and cortical ChATactivity (Wilcock et al., 1982; DeKosky et al., 1992; Perryet al., 1977) in human aging and AD. Most postmortem stud-ies examining CBF system changes in AD were derived fromend-stage patients with severe dementia. In contrast, there isa preservation of cortical ChAT activity (Fig. 2A) and NBneurons (Fig. 2B) in subjects with MCI and early AD (Daviset al., 1999; DeKosky et al., 2002; Gilmor et al., 1999). Sur-prisingly, ChAT activity is increased in the hippocampus ofpeople with MCI but not mild AD (Fig. 2B; DeKosky et al.,

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Fig. 2. (A) Histograms showing stability of ChAT activity in anterior cingulate and superior temporal cortex in people with mild cognitive impairment(MCI) and mild Alzheimer’s. By comparison there was a significant up regulation of ChAT activity in the superior frontal cortex and the hippocampusin MCI. Composite histogram showing the phenotypic differences in the number of choline acetyltransferase (ChAT), vesicular acetylcholine transporter(VAChT), trkA and p75NTR-ir neurons in MCI and AD individuals. Note a significant reduction in NGF receptor and not ChAT containing neurons withinthe basal forebrain of people with MCI and AD. This difference is not acerbated during the transition from MCI to AD. Reproduced with permissionfrom Mufson et al. (2001).

2002; Ikonomovic et al., 2003). This suggests that the sep-tohippocampal projection system is capable of neural reor-ganization during the prodromal stage of AD, manifestedby its sprouting in response to the loss of glutamatergic en-torhinal input to the hippocampus (DeKosky et al., 2002;Ikonomovic et al., 2003). In contrast, there is a marked re-duction in the number of CBF neurons containing the proteinand mRNA for trkA and p75NTR early in the disease process(seeFig. 2B; Mufson et al., 2000b, 2002b) suggesting a phe-notypic silence for different chemicals during the early stageof the disease. Studies of the molecular mechanisms asso-ciated with CBF neuron dysfunction using single cell RNAamplification and custom array technology indicate changesin functional classes of mRNAs in cholinergic NB neurons

including neurotrophin receptors, synaptic proteins, proteinphosphatases, and amyloid-related proteins in AD (Mufsonet al., 2002a). Interestingly, trkB and trkC mRNAs were sig-nificantly down regulated whereas p75NTR mRNA levels re-mained stable in end stage AD. TrkA mRNA was reducedby approximately 28% but did not reach statistical signifi-cance (Fig. 3A). In addition, there was a down-regulation ofsynaptophysin, synaptotagmin (Fig. 3B), and protein phos-phatases PP1� and PP1� mRNAs whereas, cathepsin DmRNA was up regulated in AD. Thus, cholinergic NB neu-rons undergo selective alterations in proteins and classes ofgenes in AD. These findings suggest differences in the neu-robiologic machinery related to cholinergic neurotransmis-sion, neurotrophins and synaptic transmission early in the

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Fig. 3. (A) Representative gene array (right) and histogram (left) illus-trating down-regulation of the genes for trkB (38±11% of control) andtrkC (47±9%) but not p75NTR in single cholinergic NB neurons in ADas compared with aged controls. Note the trend for a reduction of trkAmRNA in AD. *, P = 0.05 via ANOVA with Neuman–Keuls post hoc testfor multiple comparisons. (B) Representative gene array (top) and his-togram (bottom) illustrating down-regulation of synaptophysin (25±7%of control) and synaptotagmin (23±6%) expression in AD cholinergicNB neurons as compared with aged controls. No alterations in expressionlevels were seen for synapsin-1, synaptobrevin, or SNAP-29. *,P = 0.01via ANOVA with Neuman–Keuls post hoc test for multiple comparisons.Reproduced with permission fromMufson et al. (2002a,b).

disease process. Therefore, CBF neurons may be viable, al-beit, dysregulated, at this stage of AD.

In conclusion, the present discussion indicates the com-plex nature of the chemical anatomy of the human CBF in theaging and AD brain. Recent, studies indicate that the CBFsystem is capable of maintaining its cholinergic phenotypeas well as up-regulating ChAT activity in the hippocampusand superior frontal cortex in the prodromal stage of AD(DeKosky et al., 2002). Therefore, the cholinergic hypothe-sis suggesting that CBF dysfunction is an early pathologicalfeature of cognitive decline requires re-evaluation at least atthe level of ACh synthesis. Future investigations are neededto more fully evaluate other components of cholinergic func-tion (e.g. nicotinic receptors and the high affinity cholinetransporter) during aging and the progression of AD.

Acknowledgements

Supported by AG14449, AG16765, AG09466 andAG10161, AG10668, NS43939 and the Alzheimer’sAssociation.

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