he dopamine transporter (DAT), located on the presynaptic dopamineneuron,is the principalmechanismforregulating synaptic dopamine levels in the synaptic cleft.The majority of dopamine is taken up into the neurons by

this processand is repackaged into presynaptic vesicles to bereleased in the next neurotransmission signal. The striatum

is a dopamine-richregion and consequentlyhashigh densities of DAT (1). As a result of its significant role in dopamineregulation, DAT is a targeted site for various psychoactivedrugs, including methylphenidate, cocaine and amphetamines (2). Most radiopharmaceuticals that target DAT havebeen labeled with positron-emitting isotopes (3) or therelatively expensive single-photon emitter 1231(4—6).Thegrowing interest in imaging studies on the dopaminergicsystem, particularly of the presynaptic site, DAT, and theavailability of 99mTc in every clinical nuclear medicinefacility have led to efforts to develop 99mTc@labeledligandsthat are suitable as routine imaging agents for SPECT. Theuse of 99mTcbasedsmall molecules as site-specific imagingagents has been reviewed (7) and several @°‘Tc-labeledtropanes have been reported (8—11).[99mTc]TRODAT@1(technetium, [2-[[2-[[[3-(4-chlorophenyl)-8-methyl-8-azabicyclo[3,2,1]oct-2-yl]methyll(2-mercaptoethyl) amino

]ethyl]amino]ethanethiolato (3-)]- oxo-[ 1R-(exo-exo)]), anovel 99mTc4abeledtropane derivative, was initially evaluated and characterized as a potential imaging agent for DAT(10, 12). Further pharmacological characterization of in vivobinding of [99mTc]TRODAT..1demonstrated that the bindingof this new 99mTccomplex in the brain was associated withDAT (13).

The postsynaptic dopaminergic receptors, dopamine D2/D3receptors, have been studied in vivo using SPECT with[123ljiodobenzamide (IBZM) (14) or [‘23I]iodobenzofuran(IBF) (15). Both iodinatedligandshave beenshownto bindreversibly with high affinity and high specificity to striatalD2/D3 receptors (16). Using [‘23I]IBFor [‘23I]IBZMinconjunction with SPECT, researchers have evaluated thestatus of the D21D3 receptors in a wide variety of diseases(14,17,18).

It isgenerallyrecognizedthatimagingpre-andpostsynaptic sites of the dopaminergic system is important forunderstandingthe statusof the dopaminergicsystemandthat the images provide critical information for patient

The central nervous system dopamine transporters (DAT5)anddopamine D@/D3receptors are implicated in a variety of neurological disorders. Both sites are also targets for drug treatment.With the successful development of [@Tc]TRODAT-1, singleisotope imaging studies using this ligand for DAT imaging canbe complemented by additional use of 1231-IabeledD@/D3receptor ligand co-injected to assess both pre- and postsynaptic

., sites of the dopaminergic system simultaneously. Methods:

TwelveSPECTscans of the brainwere obtained in two baboonsafter intravenous administration of 740 MBq (20 mCi) [@‘Tc]TRODAT-1 (technetium, [2-[[2-[[[3-(4-chlorophenyl)-8-methyl-8-azabicyclo[3,2,ljoct-2-yl]methyl](2-mercaptoethyl)aminojethyl]amino]ethanethiolato(3-)]- oxo-[1R-(exo-exo)])and 185 MBq (5mCi) [1@l]iodobenzamideor [1@l]iodobenzofuran.SPECT datawere acquired by a triple-head gamma camera equipped withultra-high-resolutionfanbeam collimators (scan duration = 210mm). Two sets of SPECT data were obtained using energywindows of 15% centered on 140 keV for @Tcand 10%asymmetric with a lower bound at 159 keV for 1231.Aftercoregistration with MRI, region-of-interest analysis was performed using predefined templates from coregistered MRI. Inblocking studies, baboons were pretreated with N-methyl-2@3-carbomethoxy-3@3-(4-fIuorophenyl)tropane(CFT,14 mg)or raclopride (14 mg) to block DAT or DID3 binding site, respectively.Results:Imagequalityofdual-isotopestudieswassimilartothatobtainedfrom single-isotopestudies.Whenone sitewas blockedwithCFTor raclopride,thebindingoftherespectiveligandtotheothersite was notaffected.Conclusion: This is the first examplethat clearly demonstratesthe feasibilityof simultaneousimagingof bothpre-and postsynapticsitesof the dopaminergicsysteminbaboons with dual-isotope SPECT studies. With or withoutcorrectionsfor cross-contaminationof 1231into the @“Tcwindow,striatum-to-cerebellum ratios (target-to-nontarget) of dualisotope experiments did not differ significantly from singleisotope experiments. This method may be a valuable andcost-effectivetool for gaining comprehensiveinformationaboutthe dopaminergicsystemin one SPECTimagingsession.

KeyWords:dopaminetransporters;dopamineD@/D3receptors;SPECTimaging;dual-isotopestudies

J NucI Med 1999;40:660—666

Received Mar. 24, 1998; revision accepted Sep. 22, 1998.For correspondence or reprints contact: Stefan H.J. Dresel, MD, Department

of Radiology,Universityof Pennsylvania,3700 Market St., Room 305,Philadelphia, PA 19104.

660 THE JOURNALOF NUCLEARMEDICI@ •Vol. 40 •No. 4 •April 1999

Simultaneous SPECT Studies of Pre- andPostsynaptic Dopamine Binding Sites in BaboonsStefan H.J. Dresel, Mei-Ping Kung, Xiao-Feng Huang, Karl Plössl,Catherine Hou, Sanath K. Meegalla,Gregory Patselas,Mu Mu, Janet R. SalTerand Hank F. Kung

Department ofRadiology, University ofPennsylvania, Philadelphia, Pennsylvania

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management. It is possible to achieve this by using 1231..labeled compounds in two separate sessions. The limitationof this is that two scansmust be separatedby several days toallow the radioactivity of the first scan to decay. During thatinterval, the patient's condition and medication may havechanged and the patient will most likely be in a differentpositiononthetable.Manypatients,particularlypsychiatricpatients, may have difficulty lying still for SPECT dataacquisition. Dual-isotope studies would reduce by half thetime spent on the table compared with two scansperformedindependently, and costs would be minimized. Simultaneousassessment of pre- and postsynaptic binding sites of thedopaminergicsystemcanfacilitate thediagnosisof adiseaseinvolving the dopaminergic system. The simultaneous SPECTstudy can avoid the pitfalls mentioned previously, therefore,full information about the status of the DAT and D2/D3receptors would be obtained in one SPECT imaging settingwith images that are in accurate spatial registration to eachother. The images of each energy window are in anatomicregistration within 1 mm. Therefore, the influence on dataanalysis from placement of regions of interest (ROIs) fromstudies performed on different days can be eliminated.Motion artifacts can also be reduced using the dual-isotopeapproach. These studies can be performed in most nuclearmedicine facilities with commercially available gammacameras.

There is a clear advantage to investigating DAT and D21D3receptors using dual-isotope imaging studies with [99mTc]@TRODAT-1 and [‘23I]IBFor [1231]IBZM. It has been shownthat dual-isotope imaging with 99mTcand 1231is feasible andinformationfrombothradionuclidescanbeacquiredsimultaneously from appropriately selected energy windows (19—21). The objective ofthis study was to evaluate this approachin vivo using [99mTc]TRODAT4and [1231]IBF/[1231]IBZMtosimultaneouslyimageDAT and D2/D3receptorsin healthybaboons. Additionally, the baboons were pretreated withN-methyl-2@3-carbomethoxy-3@3-(4-fluorophenyl)tropane(CFT; also known as WIN 35,428) or raclopride to blockpre- or postsynaptic dopamine binding sites, respectively,and thereby to mimic conditions that may occur in patientsdisplaying a loss of either DAT or D21D3receptors.

MATERIALS AND METHODS

Radiopharmaceuticals and DrugsPreparationof[@―Tc}TRODAT-l,[‘23I]IBFand[‘23I]IBZMwas

performed according to methods published previously (12,22,23).CF7fwas synthesized in our laboratory. Raclopride was purchasedfrom ResearchBiochemicals (Natick, MA).

AnimalsA series of 12 SPECT scans was performed on two female

baboons(Papio anubis) under protocols approvedby the localanimal care committee and consistent with guidelines of theNationalInstitutesof Health.Theintervalbetweeneachsuccessivestudy was at least 2 wk. After fasting overnight, animals wereimmobilized with an intramuscularinjection of Saffan (1 mg/kg)(0.9% w/v alfaxalone,0.3% alfadoloneacetate;Pittman-Moore,

Middlesex, UK). During the scanningprocedure,anesthesiawasmaintained by intravenous (tibial vein) infusion of a solution ofSaffan (2.4 mg/mL) at a rate of 45 mL/h. Passive inhalation ofoxygen (1.5 IJmin) was also maintained to keep the oxygensaturation >95%. An additional intravenous line in a superficialbrachial vein was usedfor hydration (0.9% physiological salinesolution, 5 mL/kg/h), as well as for ligand administration. Acylindrical polycarbonatepositioning device equipped with acustomized foam head-holder enabled reproducible placement ofthe animal in the SPECT camera. The core body temperature wasmaintained using a circulating warm water pad maintained at 37°C.A single-bolusinjection of [@‘°Tc]TRODAT-land [‘231]IBF(or[‘23I]IBZM)was administered immediately before each scan with ameanbolusinjectionof 729.64±45.88MBq (19.72±1.24mCi)[99mTc]ThODAT1and 183.89±19.61MBq (4.97 ±0.53 mCi)[‘23I]IBF(or [‘231]IBZM).A total of eight blocking studies wereperformed; four with the additional injection of raclopride (1mg/kg) and another four with the additional injection of CFT (1mg/kg). These compounds were injected 5 mm before the administrationof theradiopharmaceuticals.

Image Acquisition and ProcessingFor SPECT image acquisition, we used a triple-head gamma

camera equipped with ultra-high-resolution fanbeam collimators(Picker Prism 3000S, Picker International, Cleveland, OH). Theacquisition parameters comprised a rotational radius of 14 cm, a15%energywindowcenteredon 140keV anda 10%asymmetricenergy window with a lower bound at 159 keV, 120 projectionanglesover 360°and a 128 X 128 matrix with a pixel width of 2.11mm in the projection domain. Data collection started 20 mm afterinjection.Three10-mmscans(15 s/step)followedby two40-mmscans(60 s/step)were obtainedover a total of 210 mm. Theprojectionimageswerereconstructedby filteredbackprojection.Then a three-dimensional, count-rate dependent postprocessingfilter (Wiener filter, Picker International) with a modulationtransformer function specific to both @mTcand 1231was applied.For uniform attenuation correction, Chang's first-order method wasused. MR images were acquired on a 1.5-T instrument (GeneralElectric Medical Systems, Milwaukee, WI) using a repetition timeof 3000 ms and echo times of 30 and 80 ms with 5-mm spacing thatproduces0.97 x 0.97 X 1 mm voxels.Sliceswere interleavedsothat there was no interslice gap. The MR images were resized andreslicedin planesparallelto theonecontainingtheanteriorandposterior commissures. ROIs for the nucleus caudate and theputamen were drawn directly onto the MR image inside the outeredgeof each structure to minimize the effects of volume averaging.Coregistration of the SPECT scans and the MRI was achievedusing a customized software package developed to rotate andtranslateeachSPECTscanwith respectto a fixed MRI dataset.Fusion images of the SPECT and MR images were generated tovisually validatethe accuracyof the coregistrationin the sagittal,coronalandtransverseplanes.To assessspecifictraceruptakeinthe striatum, we usedthe ROI template created with the MM. Meanspecific uptake in the striatum region (STR) was calculated bysubtracting the mean counts per pixel in the cerebellum asbackground(BG)fromthemeancountsperpixelin theSTRanddividing theresultby themeancountsperpixel in theBG, (STR —BG)/BG.Uptakevalueswereexpressedascorrectedvaluesonthebasis of the results of the phantom studies for cross contamination.The @“@Tcscanswerecorrectedfor down contaminationof 1231bysubtracting 14.9% of the 1231counts. 1231scans were corrected by

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subtracting0.54% of the @mTccounts.For statisticalanalyses,analysisof variance(ANOVA),regressionanalyses(least-squaresmethod) and the Student t test were used. Monoexponential orlinear fitting was applied to all data. Data were considered to bestatistically significant with a P value of <0.05. Results werecomparedwith single-isotopeSPECTscansof [99mTc]ThODATI,whichwereacquiredpreviously.

RESULTS

A total of 12 dual-isotope SPECT studies were performed:4 control studies, 4 studies with pretreatment of CFT and 4studies with pretreatment of raclopride. Preliminary studiesindicated that no statistically significant difference wasobservedfor [99mTc]TRODAT.iin both dual-isotopeandsingle-isotope scans (data not given). The average specificuptake values for [99mTc]TRODAT@l at 210 mm afterinjection were 2.18 ±0. 11. The slope of the uptake curvesdid not show any significant differences between single- and

dual-isotope experiments, and the uptake ratios correlatedwell (0.95). Figure 1 displays typical SPECT scans of[‘231]IBFand [99mTc]TRODAT4 at 170—210mm postinjection and the coregistered MR images. Similar scans ofdual-isotope studies with [1231]IBZMand [99mTc]TRODAT..lwere observed (data not given). Both SPECT scans shownormal uptake and homogeneous binding of the radiopharmaceutical to the binding site of the baboon striatum.Specific uptake of [‘231]IBFwas calculated as 9.59 ±0.77and of [‘231]IBZMas 2.91 ±0.19. Under pretreatment withraclopride ( 1 mg/kg), a specific blocker for D21D3receptors,the specific binding of [1231]IBF in the nucleus caudateputamen area of the baboons was dramatically diminished(Fig. 2A), while the simultaneously acquired scanof [99mTc]..TRODAT-l remained unchanged (Fig. 2B). Similar resultswere observed for [1231]IBZM scans under pretreatment with

raclopride (data not given). The semiquantitative analysisrevealed a comparable specific uptake value for [99mTc]@TRODAT-l (reflecting DAT uptake) in normal andraclopridetreated baboon studies (Fig. 3A), whereas the specific uptakefor [1231]IBF or [‘23I]IBZM,unlike the untreated ratios,

FIGURE1. Dual-isotopeSPECTscanwith[@Tc]TRODAT-1and [1231]IBF170—210mmpostinjection. (A) [1231]IBFcoregistered to Ti -weighted MR image in sagittal, transverseand coronalviews displays normalbindingofligand to dopamine D@/D3receptors. Specificbinding of 9.6 is demonstrated. (B) [@Tc]TRODAT-1coregistered to Ti -weighted MRimage in sagittal, transverse and coronalviewsrevealsregularbindingof radiopharmaceutical to DAT. Specific binding is 2.2 forcrosstalkcorrectedimage.

increased only slightly over time, resulting in a markedlydiminished specific binding of 1.02 ±0.03 and 0.68 ±0.05for [‘23I]IBFand [123I]IBZM, respectively (Fig. 3B), inraclopride-treated baboons. The results of pretreatment withCVF are shown in Figure 4 for the presynaptic receptor99mTc4abeledcompound and for the postsynaptic 1231..labeled compounds (IBF and IBZM, respectively). Becauseof the high nonspecific binding of [@Tc]TRODAT-l compared with [123I]IBF, the blockade of the DAT binding sitesdoes not reveal an “emptyshell―image like the one of

[‘231}IBFin a raclopride-pretreatedbaboon. However, theSPECTimagesof the 99mTcwindow revealeda remarkablereduction of specific uptake of [99mTc]TRODAT..l with aspecific uptake value of 0.32 ± 0.06 for the crosstalkcorrected scans at 210 mm postinjection (Fig. 5). Thespecific uptake ratio curve of CFF-treated studies increasesonly slightly over time, suggesting the blockade of specificbinding of [99mTc]'fl@ODAT4 to DAT. This result is inconcordance with the effect of CFT pretreatment on [@Tc]TRODAT- 1 binding to DAT in rats (24). The binding of[123I]IBF (or [‘23I]IBZM)to D2/D3 receptors in CFFpretreatedbaboons,as shown in Figure SB, however, doesnot significantly differ from the scans without any pretreatment.

DISCUSSION

The dual-isotope SPECT imaging performed in this studyin baboons provided similar information and image qualityas the corresponding single-isotope studies. We also used theadditional pretreatment of drugs that are known to selectively block one binding site to mimic the loss of this sitewith the other site remaining intact. The blocking studieswith CFT or raclopride, specific for DAT or dopamine D2/D3receptors,respectively,indicatedthat there is no effect onthe radiopharmaceutical binding to the sites that were notblocked. These findings are in concordance with the experimental phantom studies (data not given) and may represent

AII-12311BFSPECT/MRI

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662 Ti-ui JOURNALOF NUCLEARMEDICINE a Vol. 40 •No. 4 •April 1999

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.lTc-99m]TRODAT-1dualisotope study

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dual isotope study afterpretreatmentwith raclopride

ALI-12311BFSPECTIMRIraclopride blocking

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FIGURE2. Dual-isotopeSPECTscanwith[@“Tc]TRODAT-1and [1@l]lBF 170—2i0mmpostinjection. Baboon was pretreated withraclopride(1 mg/kg)5 mmbefore injectionofradiopharmaceuticals.(A) [1@l]lBFcoregistered to Ti-weighted MR image in sagittal,transverse and coronal views displaysmarked reduction of specific binding of Iigand to dopamine D@/D3receptors.There isalmostno uptakevisible in striatalarea.Scanreveals specific binding of i .02. (B) [@Tc]TRODAT-i coregisteredto Ti -weighted MRimage in sagittal, transverse and coronalviewsshowsregularbindingof radiopharmaceutical to DAT as seen in Figure i B. Specific binding is 2.i8 for crosstalk correctedimage.

FIGURE 3. Dual-isotopeSPECT studieswith [@mTcJTRODAT-iand [‘23l]IBF(or[1@I]lBZM).Baboons were pretreated withraclopride (1 mg/kg) 5 mm before injectionof radiopharmaceuticals.(A) Specific binding is displayedfor dual-isotope[@“Tc]TRODAT-i SPECT scans corrected forcrosstalk for control baboons and baboonsthat were pretreated with raclopride. Datafor blockingwith raclopridedoes not significantly differ from data obtained withoutblocking. (B) Specific binding is displayedfor dual-isotope SPECT scans of [1@l]lBFand [123l]IBZMin baboonswith and withoutpretreatment with raclopride. Binding of[1231]IBF(or [1231]IBZM)to dopamine D@/D3receptorsis markedlydiminished.

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conditions that may appear in patients suffering from variousdiseases.

There is considerableinterest in using in vivo imagingtechniques to study the dopaminergic system in the humanbrain. Mozley et al. (25) found a strong correlation betweenDAT density and age. A close relationship between striatal[‘@I]N-(3'-iodopropen-2'-yl)-2@3-carbomethoxy-3@3-(4-chlo

rophenyl)tropane(IPT) or [123I]@3-(1R)-2@3-carbomethoxy-3@3-(4-iodophenyl)tropane(CIT) bindingandclinical featuresofpatients suffering from Parkinson's disease was observed,suggesting that this method can be used to objectivelyfollow the course and progression of the disease (5,26). Thepotential of imaging the DAT will be further enhanced andbroadened with the availability of a 99mTc..labeled ligand,

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FIGURE4. Dual-isotopeSPECTscanwith[@TclTRODAT-i and [1231]IBZMi70—2i0mm postinjection. Baboon was pretreatedwithCFT(i mg/kg)5 mmbeforeinjectionofradiopharmaceuticals. (A) [1@l]IBZMcoregistered to Ti -weighted MR image in sagittal,transverse and coronal views displays normal and homogeneous specific binding ofligand to dopamine D@/D3receptors. Scanreveals specific binding of 2.93. (B) [@“TcJTRODAT-icoregisteredto Ti -weightedMRimage in sagittal, transverse and coronalviews shows heterogeneous and markedlyreduced binding of radiopharmaceuticaltoDAT.Note relatively high nonspecific backgrounduptake,whichisvirtuallyindistinguishable from any uptake in striatum area. Specific binding is 0.32 for crosstalk correctedimage.

which may readily be used in every nuclear medicinefacility. There are numerous clinical advantages of [@mTc]@labeled ligands compared with [‘231]-basedcentral nervoussystem receptor imaging compounds. Most important, @°Tcis the radionuclideof choicefor nuclearmedicinebecauseit

FIGURE 5. Dual-isotopeSPECT studieswith [@TcJTRODAT-i and [123I]IBF (or[1@l]lBZM).BaboonswerepretreatedwithCFT (i mg/kg) 5 mm before injection ofradiopharmaceuticals.(A) Specific bindingis displayed for the dual-isotope [@Tc]TRODAT-1 SPECT scans corrected forcrosstalkwith andwithoutpretreatmentwithCFT. Uptake is significantly reduced compared with SPECT scans in untreated baboons. (B) Specific binding is displayedfordual-isotopeSPECT scans of [1231]IBFand[1@l]lBZMwith and without pretreatmentwith CFT.CFT pretreatmentof baboonsdidnot affectbindingof [123l]IBFand [‘23l]IBZM.

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is readily available, is relatively inexpensive and gives lowerradiation exposure compared with 1231. It has been established by in vivo and in vitro studies that [@Tc]TRODAT-1reliably assesses the DAT status of healthy baboons andhumans and yields high-contrast SPECT imaging at 2—3h

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664 THE JOURNALOF NUCLEARMEDICINE•Vol. 40 •No. 4 •April 1999

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after injection (10, 13). The recently completed phase I studyof [@mTc]TRODAT@ldemonstrated that this ligand is safeand suitable for human imaging (27).

Studies of the dopamine D2/D3receptors with [123I]IBZMSPECTimaginghavebeenperformedsince1988.Thereis awealth of information on its clinical applications based onmore than 150 articles published in the literature. Currently,[‘@IIIBZMis commercially available in Europe, and [‘231]IBFis undergoing phase III clinical trial in Japan. Because oftheir potential clinical application in the future, we chosethese two D2/D3 receptor imaging agents for this study. Itwas reported that the most effective use of [‘231]IBZMSPECTstudiesis to differentiatethe idiopathicandnonidiopathic parkinsonian syndrome (14). The occupancy ofdopamineD2/D3receptorsin schizophrenicpatientsundertreatment with neuroleptics and its relationship with extrapyramidal side effects has been described in several studies(17,18,28). The coexistence of both Parkinson's diseaseandschizophrenia hasbeen reported. The comorbidity makes thedifferential diagnosis difficult. These patients are usuallytreated with neuroleptics that might cause the parkinsoniansyndrome by binding to the D21D3 receptor (18,29—31).Simultaneous assessment of both sites of the dopaminergicsystem will significantly simplify the correct diagnosis.Furthermore,the involvementof thedopaminergicsysteminmany other diseases associated with movement disordershas not yet been fully explored. It has been reported that inpatients suffering from Huntington's disease the function ofthedopaminergicsystemmight becompromised(32).Othermovement disorders, such as amyotrophic lateral sclerosis,multiple sclerosis, multisystem atrophy, Wilson's diseaseand progressive supranuclear palsy, have been evaluated foralteration of either the dopamine transporter or D2/D3receptor system (33,34). The simultaneous dual-isotopeimaging technique proposed in this article may provide aneven more powerful tool for improving the diagnosis ofthese complex disorders.

In general, SPECT imaging with [@mTc]TRODAT@1giveslower target-to-nontarget ratios (2—3)than 1231tropanederivates (10—15for @3-CITand 1FF). This may be due to itslower binding affinity to DAT and a faster in vivo metabolism (35). In addition, the down contamination of 123! into

the 99mTcwindow may further contribute to the highbackground activity. It is important to note that the slope ofthe uptakeandretentioncurveswith [@Tc]TRODAT-l didnot differ significantly between single- and dual-isotopestudies. The in vivo studies in primates further confirm theresults obtained from phantom studies. The cross contamination of 1231into the @°‘Tcwindows is consistent over a widerange of activities and therefore does not change relativeuptake ratios significantly. The target-to-background ratiosof the uncorrected images have shown a trend to be slightlyhigher compared with the corrected data due to the contamination of the other isotope. However, this difference was notstatistically significant (P < 0.05). The loss of one bindingsite (mimicked by drug pretreatment) can be imaged accu

rately in the presenceof unchanged binding of the other sitesin the striatum. The crosstalk corrections in each energywindow did not change the outcome measure significantly.

To our knowledge,thereis no previouslypublishedstudyon simultaneous dual-isotope imaging of the dopaminergicsystem. However, there are reports of studies using @mTc@and 1231-labeledradiopharmaceuticals on cardiac imaging(36) and brain blood-flow imaging (19—21,37).There is amajor difference between the blood-flow studies and thespecific binding site imaging presented in this article. Thedistribution mechanisms of both agents used for the bloodflow studies are similar. Generally, they are nonspecificsimple diffusion mechanisms. The ligands used in this study,however, target different sites, pre- and postsynaptic dopaminergic binding sites, which are functionally distinct andseparated by the synaptic cleft about a few microns apart.Thus, subtle changes in physiological conditions of onebinding site may or may not directly influence the otherbinding site. The binding sites for [@mTc]TRODAT@1and[‘231]IBF(or [‘231]IBZM)in the dopaminergic system arefunctionally different, even though their localization in thestriatum is identical. Partial volume effects in SPECTimaging are of importance, but it is expected that the effectson 123!and @‘@Tcare similar. The net effects on the specificbinding ratios are not dramatically different between thesetwo isotopes. In contrast, the differences in blood-flowimages, for example, during monitoring cognitive activationor measuring minor increases in regional cerebral blood flowafter stimulation, may be more subtle and not as characteristic and anatomically well defined as in SPECT scans of thestriatum (19).

In this study, no effort was made to obtain quantitativekinetic information. The blocking studies were performedusing a relatively high blocking dose: raclopride (1 mg/kg)and CET (1 mg/kg) for D21D3receptors and DAT sites,respectively. One should view the data in this article aspreliminary proof of principle in using simultaneous dualisotope SPECT to study pre- and postsynaptic dopaminebinding sites. A more detailed kinetic study, including a fullevaluation of input function and in vivo metabolites, will benecessary to validate the quantitative aspect for the bindingsite measurements. In addition, there will be shades of gray

(38) on changesof binding sites in normal and diseasestates;therefore, a careful consideration of detection sensitivity ofthis imaging technique is required. It is encouraging that

both of these issues have been addressed previously usingsingle-isotope SPECT imaging technique for D2/D, receptors and DAT binding sites. At tracer levels, both of theligands used for measuring pre- and postsynaptic dopaminebinding sites behave independently without interfering witheach other. Therefore, it is likely that the kinetic modeling ofboth ligands will follow a similar model as that reported forsingle-isotope studies. A dual-isotope kinetic modelingstudy in vivo will be evaluated and published in the future.

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16. Kung HF, Alavi A, Chang W. et al. In vivo SPECT imaging of CNS D2 dopaminereceptors: initial studies with iodine-123-IBZM in humans. J Nuci Med. I990;31:573—579.

17. Pilowsky LS, Costa DC, ElI PJ, Murray RM, Verhoeff NPLG, Kerwin RW.Clozapine, single photon emission tomography, and the D2 dopamine receptorblockade hypothesis of schizophrenia. Lancet. 1992:340:199—202.

18. Dresel 5, Tatsch K, Dthne I, Mager T, Scherer J, Hahn K. Iodine-l23-

iodobenzamide SPECF assessment of dopamine D2 receptor occupancy inrisperidone-treated schizophrenic patients. JNucIMed. 1998:39:1138—1142.

19. O'Leary DS, Madsen MI, Hurtig R, et al. Dual isotope brain SPECT imaging formonitoring cognitive activation: initial studies in humans. Nuci Med Commun.1993;14:397—404.

20. DevousMD Sr. PayneJK, Lowe JL. Dual-isotopebrain SPECI' imaging withtechnetium-99m and iodine-123: clinical validation using xenon-133 SPECI'. INuciMed. l992;33:19l9—l924.

21. Devous MD Sr. Lowe JL, Payne JK. Dual-isotope brain SPECT imaging withtechnetium-99m and iodine-123: validation by phantom studies. J Nuci Med.1992:33:2030—2035.

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CONCLUSION

We have demonstrated dual-isotope imaging of the dopaminergic system using [99mTc]TRODAT.i and [‘231}IBF(or[1231]IBZM) in nonhuman primates. Simultaneous imagingof both pre- and postsynaptic binding sites of the dopaminergic system is feasible and produces images of comparablequality as single-isotope scans. To our knowledge, this is thefirst example of dual-isotope SPECT imaging of the DATs

and dopamine D2/D3 receptors in the living brain.

ACKNOWLEDGMENTS

The authors thank SusanWest and Laura Danich for theirassistance in preparing the manuscript. This work wassupported by grants from the National Institutes of Health(NS24538 and NS 18509) and the Department of Energy(DOE-ER-61657).

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