Nicotinic versus muscarinic blockade alters verbal working memory-related brain activity in older women

Nicotinic Versus Muscarinic BlockadeAlters Verbal Working Memory-Related

Brain Activity in Older Women

Julie A. Dumas, Ph.D., Andrew J. Saykin, Psy.D.,Brenna C. McDonald, Psy.D., Thomas W. McAllister, M.D.,

Mary L. Hynes, R.N., Paul A. Newhouse, M.D.

Objectives: An important aspect of furthering our understanding of the central nervous

system function after menopause is to examine the cerebral circuitry that appears to be

influenced by cholinergic antagonist drugs in the presence and absence of estrogen. This

pilot study investigated the effects of two anticholinergic drugs on brain activation and

working memory performance in postmenopausal women not taking estrogen. This

approach simulates the effects of age- or disease-related neuroreceptor or neuronal loss

by temporarily blocking pre- and postsynaptic muscarinic and nicotinic cholinergic

receptors. Design: Six healthy postmenopausal women took part in three drug challenges

using the antinicotinic drug mecamylamine (MECA, 20 mg, oral), the antimuscarinic

drug scopolamine (SCOP, 2.5 �g/kg, IV), and placebo during functional magnetic

resonance imaging. The cognitive measure was a visually presented verbal N-back test of

working memory. Results: Neither MECA nor SCOP significantly impaired performance

on the verbal N-back. Functional magnetic resonance imaging results showed greater

increases in frontal lobe activation in the placebo condition relative to each drug

condition with different specific regional activation for MECA and SCOP. Conclusions:These preliminary results suggest that brain activation patterns are sensitive to cholin-

ergic modulation in postmenopausal women and that differential effects may be ob-

served following nicotinic versus muscarinic blockade. This approach offers a potentially

valuable method for modeling age-related changes in brain function, and the findings

may have implications for cholinergic contributions to normal and pathologic aging.

(Am J Geriatr Psychiatry 2008; 16:272–282)

Key Words: Working memory, postmenopausal women, fMRI, cholinergic system

Received March 23, 2007; revised August 17, 2007; accepted August 23, 2007. From the Department of Psychiatry (JAD, PAN), ClinicalNeuroscience Research Unit, University of Vermont College of Medicine; Department of Radiology (AJS, BCM), Center for Neuroimaging, IndianaUniversity School of Medicine; and Department of Psychiatry (AJS,TWM, BCM, MLH), Brain Imaging Laboratory, Dartmouth Medical School,Dartmouth, NH. Send correspondence and reprint requests to Paul A. Newhouse, M.D., Department of Psychiatry, Clinical NeuroscienceResearch Unit, University of Vermont College of Medicine, 1 South Prospect St., Burlington, VT 05401. e-mail: [email protected]

© 2008 American Association for Geriatric Psychiatry

Am J Geriatr Psychiatry 16:4, April 2008272

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Converging evidence from psychopharmacologi-cal, neuroimaging, and psychological studies

shows that the cholinergic system has a specific mod-ulatory role in cognitive processing. In humans, thecholinergic system has been implicated in many as-pects of cognition including the partitioning of atten-tional resources, working memory, inhibition of ir-relevant information, and improved performance oneffortful tasks.1 Prior research has shown that olderadults perform more poorly than younger adults in avariety of cognitive domains including workingmemory.2 Cholinergic deficits due to degeneration ofbasal forebrain cholinergic nuclei appear to be inti-mately tied to the cognitive deficits in attention,learning, and memory in patients with neurodegen-erative disorders such as Alzheimer disease andLewy body dementia.3 Further, the activity of thecentral nervous system cholinergic system may be aprimary determinant of the effectiveness of atten-tional, learning, and memory mechanisms.4 Intrigu-ingly, cognitive symptoms reported by women atmenopause also include difficulties in memory, at-tention, and word finding, and some studies haveshown an acceleration of cognitive problems of agingafter menopause.5 Women also have a higher riskthan men of developing cholinergic-related dement-ing disorders such as Alzheimer disease.6

What has not yet been fully elucidated is how theeffect of aging on neurotransmitter systems affectsthe cognitive processes these systems support. Thisstudy examined the effects of cholinergic antagonistson working memory performance and related brainactivation in cognitively normal postmenopausalwomen without hormone therapy. Specifically, theeffects of nicotinic and muscarinic blockade on work-ing memory performance and brain activation pat-terns were examined in older women.

In the working memory models of Baddeley7 andNorman and Shallice,8 the central executive or su-pervisory attention system is a limited capacity pro-cessing resource that coordinates necessary opera-tions for a task. Warburton and Rusted9 proposedthat the cholinergic system modulates processes thatare supported by a limited capacity central executiveand influences information processing during tasksthat engage the control processes for the allocation ofcognitive resources. Additionally, age-related changesin cognition may occur as a result of impairments inallocating necessary resources to a task, thereby im-

plicating negative alterations in the cholinergic sys-tem as one of the main factors in cognitive aging.Thus, there is a link between cholinergic dysfunctionand age-related cognitive dysfunction.

Prior research by Sunderland and Newhouse us-ing scopolamine (SCOP), a muscarinic antagonist,and mecamylamine (MECA), a nicotinic antagonist,have shown that these agents can be used to simulateage- and disease-related cognitive impairments.10–14

These studies have shown that muscarinic and nico-tinic blockade impairs performance on initial sensoryprocessing, attention, and psychomotor function,thereby implicating a role for the cholinergic system inthe initial processing and encoding of information intomemory. Thus age- and disease-related differences incholinergic system integrity may influence sensitivityto antinicotinic challenge on cognitive performance.

Actions of the cholinergic system in cognitive pro-cessing have also been revealed by neuroimagingstudies (see Ref. 15 for a review). Increased corticalactivity after physostigmine, a cholinesterase inhibi-tor, compared with placebo has been shown during aworking memory task in extrastriate and intrapari-etal areas during encoding, but not during retriev-al.16 Physostigmine administration also facilitated vi-sual attention by increasing activity in the extrastriatecortex during a repetition priming task.17 A numberof studies have examined the effect of SCOP on brainactivation during cognitive task performance inyounger adults18–22 and in older adults.23 In general,when SCOP impaired performance, there was also adecrease in brain activity relative to placebo in re-gions required for task performance. The tasks uti-lized in these studies were auditory conditioning,19

repetition priming,20 associative encoding,18 de-layed-matching-to-sample,21 recognition memory,22

and object location learning.23 Nicotinic blockadewith MECA has not been examined in humans usingfunctional magnetic resonance imaging (fMRI), norhave the effects of nicotinic or muscarinic blockadeon working memory.

The current study examined the brain activity as-sociated with working memory and interaction withthe cholinergic system after menopause. A priorstudy by Green et al.24 showed that high-dose mus-carinic and combined muscarinic and nicotinic cho-linergic blockade can impair performance on the N-back test of working memory in younger adults. Thecurrent study examined the effects of nicotinic and

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muscarinic cholinergic blockade separately on N-back performance and associated brain activation incognitively normal older postmenopausal womenutilizing fMRI. Given the proposed role of the cho-linergic system in central executive function and theknown age-related changes in working memory, weutilized an anticholinergic challenge paradigm to ex-amine the effects of MECA and SCOP on workingmemory performance and brain activation in olderpostmenopausal women. We hypothesized that an-ticholinergic drugs would result in decreases in fron-tal cortical activation relative to placebo during aworking memory task.

METHODS

Subjects

Subjects were six cognitively normal women, aged51–72, M � 58.8 (SD � 9.1) who were postmeno-pausal for an average of 9.9 years since their lastmenses (SD �11.0). Subjects had an average of 16.6years of education (SD � 1.4). Subjects were requiredto be postmenopausal, without menses for 1 year,have follicle-stimulating hormone (FSH) levels �30mIU/mL, without surgically induced menopause,and without the use of hormone therapy for at least1 year. These requirements for participation wereutilized to ensure a homogeneous sample with re-gard to hormone status, which has been shown toaffect brain activation (i.e., Ref. 25). Exclusion criteriaincluded history of breast cancer, smoking, heavyalcohol or coffee use, significant cardiovascular dis-ease, asthma, active peptic ulcer, hyperthyroidism,pyloric stenosis, narrow angle glaucoma, epilepsy, orcurrent or past Axis I psychiatric disorder.

Initial screening and study procedures took placeat the University of Vermont General Clinical Re-search Center. After signing informed consent docu-ments, subjects gave a medical history and under-went a physical and laboratory tests assessinghematopoietic, renal, hepatic, and hormonal func-tion. Subjects were cognitively evaluated using theMini-Mental State Exam (MMSE),26 Brief CognitiveRating Scale,27 and the Mattis Dementia Rating Scale(DRS)28 to establish a Global Deterioration Scale(GDS) score rating the degree of cognitive impair-

ment.27 Subjects were required to have an MMSEscore greater than or equal to 27, a Mattis DRS scoreof 123 or greater, and a GDS score of 1 or 2.

Behavioral screening consisted of a partial Struc-tured Clinical Interview for Diagnostic and StatisticalManual of Mental Disorders, Fourth Edition (DSM-IV)-Text Revision29 to establish the presence or absenceof Axis I psychiatric disorders. In addition, subjectscompleted the Beck Depression Inventory (BDI).30 Acutoff score of 10 was used for the Beck DepressionInventory, and subjects scoring over this criterionwere discontinued from further participation. Allsubjects met required criteria for the cognitive andbehavioral screening.

Cholinergic Challenge Procedure

After screening at University of Vermont, subjectstook part in three cholinergic challenges and fMRItesting sessions at Dartmouth-Hitchcock MedicalCenter. At each visit, subjects performed a baselinemotor skill sobriety test to have as a comparison witha second test before discharge in the afternoon. Anintravenous line (IV) was inserted and baseline vitalswere assessed. A double-blind, double placebomethod of administration of the challenge drugs wasfollowed. Subjects received one of the followingmedications: 2.5 �g/kg SCOP (calculated as thebase), 20 mg MECA (calculated as the salt), or pla-cebo. SCOP was administered intravenously andMECA was administered orally. At time 0, a capsulewas administered containing MECA or placebo.Thirty minutes later, an injection of SCOP or salineplacebo was administered through the IV. On eachday, only one of the drugs was active or both wereplacebo. The order of the drug administration acrossthe 3 days was counterbalanced. Ninety minutes af-ter the injection and 2 hours after oral pill adminis-tration, the fMRI session began at a running time of120 minutes. Structural and functional MRI studiestook approximately 70 minutes, after which subjectsand the experimenter completed behavioral assess-ment measures. Subjects completed the Profile ofMood States (POMS),31 Stanford Sleepiness Scale,32

Subjective Visual Analogue Scale (SVAS),14 and aPhysical Symptom Checklist (PSCL). The experi-menter completed the Brief Psychiatric Rating Scale(BPRS)33 and Objective Visual Analogue Scale(OVAS14).

fMRI in Older Women During Anticholinergic Challenge



Vital signs and pupil diameter were assessed at sixtime points throughout the session at running timesof 0, 30, 60, 120, 210, and 240 minutes. At the end ofthe study day, after passing the sobriety test to thesatisfaction of the research nurse and covering phy-sician, subjects were discharged.

fMRI Working Memory Task

We used a visually presented verbal N-back taskto probe working memory circuitry, wherein subjectssaw a string of consonant letters (except L, W, and Y),presented in upper or lower case to control for pat-tern recognition, one every 3 seconds. Four condi-tions were presented: 0-back, 1-back, 2-back, and3-back. The 0-back control condition had a minimalworking memory load; subjects were asked to decideif the current letter matched a single target letter thatwas specified before the epoch began. In the 1-backcondition, they were asked to decide if the currentletter matched the previous one. During the 2-backcondition, the task was to decide whether the lettercurrently presented matched the letter that had beenpresented two back in the sequence; the more diffi-cult 3-back condition followed the same pattern. Sub-jects responded to all items by button press throughan MRI compatible fiber optic button response sys-tem (LUMItouch, Lightwave Medical Industries Ltd.,Vancouver, British Columbia, Canada) to indicatewhether the item matched the target condition. Stim-uli were delivered through an MR-safe goggle andheadphone system (Resonance Technology, Inc.,Northridge, CA). Experimental tasks were presentedby computer interface and were programmed usingthe Presentation software package; the computer re-corded subject responses and reaction times. Thistask is highly robust in producing bilateral frontaland parietal activation in healthy young and elderlycontrols and is sensitive to differences in patientgroups.34–38

fMRI Scan Procedure and Preprocessing

All scans were acquired using the same GE Signa1.5 T Horizon LX scanner with echo speed gradientsusing a standard head radio frequency (RF) coil.fMRI parameters were repetition time (TR) 2,500 mil-liseconds, echo time (TE) 40 milliseconds, field ofview (FOV) 24 cm, NEX 1, yielding 29 contiguous 5

mm sagittal slices in a 64 � 64 matrix with 3.75 mm2

in-plane resolution. Initial volumes before spin satu-ration were discarded. Spatial realignment was per-formed on all raw scan data before further analysisto remove any minor (subvoxel) motion-relatedsignal change. All volumes for each subject werenormalized into standardized Montreal NeurologicalInstitute (MNI) atlas space using SPM2 (Wellcome De-partment of Cognitive Neurology, University College,London). During spatial normalization all scans wereresampled to 2 mm3 isotropic voxels. Spatial smooth-ing to a full width half maximum of 10 mm was per-formed before statistical analysis.

fMRI Analyses

fMRI analysis included statistical parametricmapping on a voxel-by-voxel basis using the gen-eral linear model approach39 as implemented inSPM2. This procedure involves deriving one meanimage per individual for each relevant contrast inthe activation task (e.g., 3 � 0 back) after account-ing for the hemodynamic response function. Thesecontrast images were then used for the secondlevel multisubject or between-group random ef-fects analyses.40 These contrast images were fur-ther analyzed using standard paired t test andanalysis of variance procedures in SPM2. Giventhe preliminary nature of this study and smallsample size, the critical significance level for grouplevel analyses was based on clusters of activatedvoxels with the probability threshold set at pcorr

�0.05 and a minimum cluster extent (k) of threecontiguous voxels. We chose to use p values cor-rected for searching the whole brain volume toattempt to minimize Type I error, given the smallsample size. Interpretation of imaging data fo-cused on differences in targeted brain regionsknown to be involved in working memory process-ing based on prior functional imaging and lesionstudies, including the prefrontal and parietal cor-tices and interconnected components of the atten-tional network shown to be activated in positronemission tomography (PET) and fMRI studies.41,42

Focusing on differences only in regions which con-stitute the brain network activated by this task andtargeting our hypotheses to these areas also pro-vides an additional means of minimizing Type Ierror.

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RESULTS

Performance Data

N-back scores were adjusted for false alarms withthe following formula: adjusted score � proportion(hits) � proportion (false alarms). This formula cor-rects for subjects adopting a strategy where theyendorse all items as targets. Data were analyzed witha 2 (drug versus placebo) � 4 (working memoryload) analysis of variance for SCOP and MECA sep-arately. The expected effects of working memoryload were found in both the SCOP (F[3,15] � 4.19,p �0.05) and MECA (F[3,15] � 24.0, p �0.001) analy-ses. No interactions of drug and working memoryload were found for SCOP (F[3,15] � 0.44, p �0.70) orMECA (F[3,15] � 0.33, p �0.80). Neither SCOP norMECA impaired performance relative to placebo onthe N-back task (F[1,15] � 0.14, p �0.7, F[1,15] � 0.95,p �0.3, respectively).

Neuroimaging Data

Performance was examined on the most difficultcondition of the N-back task, the 3-back condition,compared with the 0-back control condition, as themore difficult working memory load conditions haveproved most sensitive to group differences in ourprior studies. Activation related to SCOP and MECAchallenges was examined separately as was done forthe performance data. Across drug conditions(SCOP, MECA, placebo), the expected pattern of bi-frontal, biparietal, and bicerebellar activation wasseen for activation during 3-back relative to 0-back,consistent with our prior work.34–38 To determine theeffects of anticholinergic challenge on working mem-ory circuitry, we examined brain activation patternsfor each drug relative to placebo in three different

ways. First, we examined the drug-induced reduc-tion in brain activation by comparing placebo rela-tive to each drug condition. We hypothesized thatregions with decreased activation on drug relative toplacebo represent those areas where anticholinergicchallenge results in diminished working memory-related brain activation. Second, we examinedwhether regions existed with increased activationunder each drug relative to placebo suggesting com-pensatory activation of neural circuitry recruited tomaintain task performance during anticholinergicchallenge. Third, we directly compared MECA withSCOP to examine the differences in antinicotinic andantimuscarinic drug challenges on working memory-related brain activation. These contrasts are de-scribed below, with results from paired t test com-parisons between drug conditions (MNI coordinates,cluster extent, and region descriptions) presented inTable 1. As noted above, results discussed below andin Table 1 include only those regions with pcorr �0.05.Although other clusters of activation can be observedin the figures, these did not survive statistical correc-tion for the whole brain search volume.

The potential impairing effect of the drugs wasassessed by examining brain regions with less acti-vation during each drug challenge relative to pla-cebo. Diminished activation during MECA challengerelative to placebo was seen in the right medial fron-tal gyrus and right superior frontal gyrus (Figure 1).A similar effect was found for SCOP challenge inwhich there was less activation for SCOP relative toplacebo in the right middle frontal gyrus and the leftprecuneus (Figure 2). Overall, the distribution of re-gions with diminished activation on drug (SCOP orMECA) relative to placebo was similar, though morepronounced for SCOP than for MECA.

The potential for compensation during anticholin-ergic drug challenge was assessed by examining re-

TABLE 1. Placebo (PLC) and Drug Paired t Tests: MNI Coordinates, Cluster Extent, Region Descriptions (Brodmann’s area, BA)and Corrected p Value

Contrast MNI Coordinates X Y Z Cluster Extent Region Description pcorr

PLC � MECA 4 �22 60 1,005 Right medial frontal gyrus (BA 6) �0.00018 62 38 562 Right superior frontal gyrus (BA 9) 0.002

PLC � SCOP 24 60 26 1,767 Right middle frontal gyrus (BA 10) 0.031�12 �58 30 4,220 Left precuneus (BA 31) �0.0001

MECA � PLC 10 �40 �4 502 Right parahippocampal gyrus (BA 30) 0.005MECA � SCOP �26 �10 46 1,324 Left middle frontal gyrus (BA 6) �0.0001SCOP � MECA 34 �56 54 947 Right superior parietal lobule (BA 7) 0.002

�6 �44 42 640 Left cingulate gyrus (BA 31) 0.034



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gions with greater activation during drug challengerelative to placebo. Significantly greater activationwas found in the right parahippocampal gyrus dur-ing the MECA challenge relative to placebo. No otherregions showed significantly greater activation ondrug (either MECA or SCOP) than placebo.

The differential effects of nicotinic versus muscarinicchallenge on working memory-related brain activationduring the 3-back were examined using a paired t testin which two images per subject were entered into themodel. Comparing the 3 � 0 back contrast image forthe MECA and SCOP challenges allowed direct com-parison of brain areas activated after nicotinic versusmuscarinic blockade. In these analyses, greater activa-tion on MECA relative to SCOP was found in the leftmiddle frontal gyrus, whereas greater activation onSCOP relative to MECA was found in the right supe-rior parietal lobule and in the left cingulate gyrus (Fig-ures 3A, B). Thus, both muscarinic and nicotinic block-ade resulted in specific alterations in brain activation inworking memory circuitry relative to placebo, and theeffects of these anticholinergic drugs appeared to bedissociable.

Behavioral Measures

Questionnaires were completed by the participantsand experimenter after returning from the MRI suite toassess whether there were any negative effects of theanticholinergic drugs or fMRI session on mood andphysical symptoms. Similar to the performance dataanalysis, the behavioral data were analyzed separatelyfor SCOP and MECA relative to placebo challenge.Overall, there were no effects of either SCOP or MECArelative to placebo on the subjective measures: the POMS,the Stanford Sleepiness Scale, the SVAS, and the PSCL.On the experimenter-completed OVAS the expectedeffects of SCOP were observed with subjects beingrated as more drowsy (t[5] � 3.79, p �0.05), more fa-tigued (t[5] � 2.61, p �0.05), and less alert (t[5] � 2.60,p �0.05) relative to placebo. There were no effects ofMECA relative to placebo observed on the OVAS.

Vital Signs

Blood pressure, pulse, and pupil diameter weremonitored at six time points throughout the challenge

FIGURE 1. Statistical Parametric Maps Showing Areas with Greater Activity on Placebo Relative to MECA Challenge (p <0.05)Displayed Over the MNI Template Surface Rendering. See Text for Results of Statistical Analyses. R: Right, L: Left,A: Anterior, P: Posterior

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day. Analyses were conducted on the maximumchange score from the baseline measurement for eachvariable. Only two significant changes were observed,both of which were expected. SCOP was associatedwith a higher pulse rate relative to the placebo condi-tion (t[5] � 4.05, p �0.01). MECA was associated with asignificantly greater decline in systolic blood pressure rel-ative to the placebo condition (t[5] � 3.21, p �0.05).

DISCUSSION

This study is the first to directly compare the effectsof anticholinergic blockade of nicotinic and musca-rinic systems during a working memory task usingfMRI. A pattern of diminished activation on drugrelative to placebo was found for both MECA andSCOP with differing specific frontal regions affectedby each of the drugs. Overall, this may demonstratethat the activity of brain regions involved in perfor-mance of the N-back task was impaired by anticho-linergic blockade. Performance on the N-back taskwas not significantly impaired by the cholinergic

antagonists. It is possible that the lack of a signifi-cant effect of the drugs on performance may besecondary to the dose of SCOP particularly and therelatively small sample size. Our performancefindings are somewhat different from that of Greenet al.,24 who saw significant effects of SCOP andcombined SCOP-MECA on N-back performance inyounger adults. However, Green et al.24 used adose of SCOP that was on average 2.5 times largerthan that used here and their sample size wastwice as large. Thus, taken together with priorstudies, these data lend support to a model thatimpairment of the cholinergic system leads to re-duced ability to activate frontal regions typicallyinvolved in working memory processing.

We propose that in our postmenopausal samplethe recruitment of additional frontal brain areas maybe a result of activity of the cholinergic system. Ourdata showed that when the cholinergic receptors arepartially blocked, frontal regions are less active rel-ative to placebo. We are unable to differentiatewhether the frontal deficits seen in this study are aresult of age- or menopause-related effects on brain

FIGURE 2. Statistical Parametric Maps Showing Areas with Greater Activity on Placebo Relative to SCOP Challenge (p <0.05)Displayed Over the MNI Template Surface Rendering. See Text for Results of Statistical Analyses. R: Right, L: Left




FIGURE 3. Statistical Parametric Maps Showing Areas with Greater Activity on MECA Relative to SCOP [A] and SCOP Relative toMECA [B] (p <0.05) Displayed Over the MNI Template Surface Rendering. See Text for Results of StatisticalAnalyses. R: Right, L: Left

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activity. If menopause has similar or additive effectsto aging on cholinergic system activity and subse-quent influences on task-related brain activity, thensimilar deficits may be produced.

The data in the current study can be comparedwith data from Saykin et al.,37 who examined theeffects of donepezil, a cholinesterase inhibitor thatincreases both nicotinic and muscarinic signaling, onN-back performance in subjects with mild cognitiveimpairment (MCI). They found increases in frontalactivation in MCI subjects after donepezil treatment.In these patients with MCI, frontal regions wererecruited during this task with the aid of the procho-linergic drug, suggesting that pharmacological ma-nipulation counteracted dysfunction of the cholin-ergic system to allow successful performance of theworking memory task. Taken together, data from thecurrent study and Saykin et al.37 support the ideathat the cholinergic system is important for success-ful working memory performance and illustrate thatcholinergic influences on task-related effortful pro-cessing can be examined with fMRI.

In addition, we found a dissociation for these ac-tivation patterns for nicotinic versus muscarinic chal-lenges. Studies comparing nicotinic and muscariniceffects on the same cognitive process have been few.In a series of studies, Nathan and colleagues24,43,44

and Little et al.45 have shown that muscarinic block-ade consistently produces larger effects than nico-tinic blockade on working memory performance, butcombined antagonism showed a greater magnitudeof impairment than either drug alone, suggestinginteraction of muscarinic and nicotinic mechanismsfor optimal working memory performance. The po-tential mechanism(s) for these effects and interactionhave been suggested by two prior neuroimagingstudies. In a xenon133 inhalation study, Gitelman andProhovnik46 showed that SCOP produced frontalcortex flow reduction that correlated with SCOP-induced memory deficits and MECA produced aperfusion deficit in parietotemporal cortex. In a PETstudy of visual processing, Mentis et al.47 were ableto demonstrate that muscarinic activities predomi-nated in primary and secondary visual processingareas (striate cortex, lateral visual association areas),whereas nicotinic stimulation appeared to affect ar-eas consistent with attention to the visual stimulus(thalamus and inferior parietal regions).

Prior functional imaging studies of nicotinic stim-ulation have strongly suggested nicotinic modula-tion of attentional processing. Thiel et al.48,49 andLawrence et al.50 showed a stimulus-specific modifi-cation of neural activity in parietal cortex consistentwith improved signal-driven attentional perfor-mance on a visual spatial orienting task49 and visualprocessing tasks following nicotine. Interestingly,Kumari et al.51 found that administration of nicotineduring the performance of a verbal N-back task alsoshowed parietal activity modulation and, as with ourresults with MECA, found alterations in a distrib-uted neural network which the authors proposed isrelated to online task monitoring and attention. In-creased activation in the superior parietal regionseen in the placebo versus MECA contrast from thisstudy (Figure 1) is consistent with this interpretation.A hypothesis suggested by prior neuroimaging studiessuggests that nicotinic and muscarinic systems may beresponsible for different aspects of task performance;e.g., nicotinic stimulation may affect attentional modu-lation, rather than simply acting as a general signal gainenhancing system, whereas muscarinic effects may bemore tied to stimulus processing or encoding.15,20,47

The results of the present study tend to support thishypothesis, as the patterns of cortical activity shift weredifferent between muscarinic and nicotinic antago-nisms compared with placebo rather than simply analteration in the intensity of task-related activation.These results are also consistent with the proposal ofSarter et al.4 that the cholinergic system optimizes sig-nal-driven detection (bottom-up) processes as well astop-down knowledge-based detection and filtering ofirrelevant information. Nicotinic and muscarinic mech-anisms may influence separate components of theseprocesses and attentional changes in normal andpathologic aging.

Finally, working memory has been shown to be af-fected by the loss of circulating estrogen after meno-pause.52 The loss of estrogen during menopause andassociated alterations in cognitive functioning may be aresult of the effect of estrogen loss on cholinergic sys-tem functioning. We have shown that 3 months ofestrogen treatment attenuated the effects of anticholin-ergic-induced impairment on cognitive task perfor-mance.53 Thus, future studies should directly examinethe effects of the estrogen-cholinergic interaction onbrain activation in postmenopausal women.



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Some caveats should be considered with the inter-pretation of the current results. Although the designof this study was an intensive within-subjects design,this study had a small sample size, thus limiting thepower to detect some effects of anticholinergic drugson performance and related brain activity. Addi-tional brain areas, particularly in the drug versusplacebo comparison, may have survived correctionwith a larger sample size. In addition, we did not usea range of doses of the anticholinergic drugs. Alarger range of doses would allow for differentiationof brain and performance effects of the differentantagonists. Further studies will benefit from using arange of doses to better establish correlates betweenperformance and brain activity. Also, we were notable to separate effects of menopause from aging inthis study. To do so would require studying womenon and off hormone therapy. Additionally, as oursubjects were a small sample of healthy, highly ed-ucated postmenopausal women not on hormonetherapy, these results do not generalize to the popu-lation of all older women. Finally, the N-back task isa complex working memory task that does not allowfor the dissociation of attentional, storage, and ma-nipulation components of the processes necessary toperform the task. Tasks that allow for these dissoci-ations will need to be examined to differentiate theinvolvement of nicotinic and muscarinic systems onattention and memory processes.

In summary, this preliminary study of the effectsof anticholinergic drug challenge on brain activity inpostmenopausal women showed areas of decreasedactivation in the frontal lobe under cholinergic block-ade which were different for muscarinic versus nic-otinic blockade. We interpret these data to suggest

that both nicotinic and muscarinic systems are im-portant for the performance on working memorytasks and contribute differentially to task perfor-mance. Our subjects were postmenopausal womenwho were not taking hormone therapy and thus hadlow levels of circulating estradiol. We have previ-ously shown that estrogen treatment attenuates theeffects of anticholinergic challenge on tests of atten-tion53 and thus estradiol status may be important ininfluencing the activity of the cholinergic system. Arecent study by Smith et al.54 demonstrated the in-fluence of estrogen and progesterone treatment inpostmenopausal women on brain activation during aworking memory task. This study found differencesin brain activation between the hormone and placebocondition in frontal regions typically involved inworking memory tasks. Thus, estrogen effects onbrain activation patterns are important to examine inpostmenopausal women, and future studies shouldexamine effect of the estrogen-cholinergic system in-teraction on specific cognitive operations using fMRI.

The authors thank the research nursing staff of theUniversity of Vermont General Clinical Research Centerfor their hard work and support of this study and to ourvolunteers for their dedication to clinical research.

This work was supported by NIA F32 AG023430 (toJAD), NIA R01-AG021476 and FAHC Trustees Fundgrant (to PAN), NIA R01-AG19771 (to AJS), andNCRR-00109.

A portion of this work was previously presented as aposter at the Society for Neuroscience annual meeting, SanDiego, CA, 2005, and the annual meeting of the Society forBiological Psychiatry, Toronto, ON, Canada, 2006.

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Nicotinic versus muscarinic blockade alters verbal working memory-related brain activity in older women

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