In This Issue:• Functional MRI (fMRI)• Neuroimaging for Studying

Mental Illness• Ion Channels and Radiation Therapy• Women’s Issues in Epilepsy• GeneChip Analysis and Gliomas• Diffusion Tensor Imaging• High Magnetic Field MRI at Sycamore• Neuroscience Summer

Intern Program

Volume Three, Issue One2003

A Publication of The Wallace-Kettering Neuroscience Institute

This issue of the Wallace-Kettering (WKNI)Neuroscience Directions highlights several newertechnologies which are useful in a broad spectrum of neurologic disorders, especially in diseases where no clear structural abnormalities arefound, such as schizophrenia.• Functional MRI (fMRI) is a technique that images

physiologic changes in particular areas of the brainin response to specific activities, such as movementor visual tasks, and combines this information with structural MRI images, comparing normal toabnormal responses.

• Diffusion Tensor Imaging (DTI) is able to delineateabnormal changes in specific white matter axonaltracts by comparing them to normal pathways.Diagnosis, treatment planning, and research inschizophrenia, multiple sclerosis, strokes, tumors,epilepsy, and other brain diseases benefit from bothfMRI and DTI.

• Positron Emission Tomography (PET) scanning,which produces a functional image, is greatlyenhanced by using new radioactive tracers to locateand quantify abnormalities in specific regions of thebrain. This has been very useful for research onschizophrenia and other mental illnesses.

• GeneChip analysis can profile abnormal gene activity in brain tumors (gliomas) to improve thetreatment of these malignant tumors.

• Ion channel abnormalities in neurons are beingstudied for methods to enhance the death by radiation of malignant cells selectively, while sparing normal neurons.

Part V in the epilepsy series covers problems specific towomen. This fall, WKNI will hold a symposium onschizophrenia research. Details are included on page 21. As always, your comments and input are welcome. I can be contacted through WKNI. My e-mail is lgray26@aol.com.

From the Editor:

From the Executive Director, WKNI:

Lenora Gray, M.D., FAAN,

Editor, Co-Medical Directorof Sleep Disorders Center,

Kettering Medical Center

Gerald Szkotnicki,

Executive Director,Wallace-Kettering

Neuroscience Institute

The utility and value of a technical innovation arerarely clear from the outset. It takes the desire toimprove methods and outcomes for innovations tobecome changes in the way things are done day today. Maintaining this openness to change and continuously seeking new approaches are two of thehallmarks of medical centers of excellence, and arecore values held by the physicians and staff involvedin WKNI. As obvious as this might appear, itremains a challenge in the highly regulated andscrutinized environment that we, as healthcareproviders, work in. I would like to take this opportunity to thank the many benefactors ofWKNI, and the people behind the scenes, whomake this pursuit of beneficial change possible.

From the Medical Director, WKNI:

Fred Manchur, ChairmanDr. Theodore Bernstein, Vice-ChairmanFrank PerezRick Mace

WKNI GOVERNANCE COMMITTEE

Theodore Bernstein, M.D.,

Medical Director,Wallace-Kettering

Neuroscience Institute

Cover art by Bilal Ezzeddine, Ph.D.

It is said that nothing is moving faster than neuroscience, which is unique among clinical specialties due to the applicability and relevance ofa broad range of scientific and technical advances.These advances continue to occur in the field at anever-increasing rate, and they can become part ofthe routine regimen available to healthcareproviders only after overcoming a large number ofvalidation studies, and economic and other challenges. This gap between what is possible andwhat is available and practical is addressed onlythrough clinical trials and research, which meansthat only those facilities that have a research infrastructure can offer the latest options to

patients. In addition, since health insurance companies rarely cover the cost of new technology,sponsored research and clinical trials may be theonly way to access such care. This is why clinicallyrelevant research is integral to achieving the missionof the WKNI and Kettering Medical Center.

Dr. Gary KrausDennis HanaghanOscar BoonshoftDr. Kenneth PugarGerald Szkotnicki

Wallace-Kettering Neuroscience Institute (WKNI)is a recognized leader in defining the standards ofpatient care in the use of neuro-imaging techniques.WKNI possesses an array of state-of-the-artresearch tools for Magnetic Resonance Imaging.Physicians, scientists, nurses and technicians atWKNI are advancing the current standards of care,diagnosis and treatment for patients with brain andspine disease using sophisticated technology.Technological advancement at WKNI rivals thebest neuroscience centers in the world.

Magnetic Resonance Imaging (MRI) and functionalMRI (fMRI) are imaging techniques that takeadvantage of the magnetic properties of atomicnuclei to create images of different bodily tissuesand compounds. MRI distinguishes different tissues like fat, bone, muscle, white and gray matter, and cerebrospinal fluid from each other. Incontrast to the structural images, fMRI imageschanging blood flow in the brain over time.

This change in blood flow correlates with thechange in activity of the neurons. When a region ofbrain is active, that region begins to use more oxygen, depleting the amount of oxygen in the surrounding blood vessels. In response, theseblood vessels dilate to increase blood flow and

oxygen supply to the working neurons (a processknown as the hemodynamic response). This hemodynamic response changes the properties ofsurrounding water molecules, which in turnchanges the signal that can be detected by the MRIscanner. The change in signal intensity is a relativeone, representing the blood flow that differs from abaseline blood flow. During a typical fMRI, a subject is first asked to fixate their eyes on a pointand to relax their mind. If the subject is able to dothis, the image of baseline blood flow will show theblood flow in the subject’s brain when it is notengaging in any particular kind of activity. Whilethe living brain is never inactive, certain cognitivetasks require that specific areas become significantlymore active than when they are at rest. Comparingresting images to activated images can be used tofind out what parts of the brain are needed to do aspecific task. The task may be as simple as tapping one’s finger or passively viewing images, orit may involve complex problem solving. When theimage of blood flow in the baseline condition issubtracted from the task-related blood flow, whatremains is a picture representing the areas whereblood flow changed in response to the task (Figure 1).

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Functional MRI at KMCN/WKNIMehdi Adineh, Ph.D., Bilal Ezzeddine, Ph.D.

Mehdi Adineh, Ph.D.,

is a Research Associate/MRI Physicist at WKNI. Dr.Adineh is a Ph.D.-level scientist with extensive expertise in the application of Magnetic Resonance techniques for biomedicalresearch.

Bilal Ezzeddine, Ph.D.,

is the Project Director ofMedical Imaging at KetteringMedical Center. He has dualPh.D.s in computer scienceand biomedical engineeringfrom the Ohio State University.He has 14 years of experiencein computer hardware andsoftware, specifically asapplied to medical image acquisition, processing, and visualization.

Figure 1.

The images produced by MRI and fMRI require the use of complex software and powerful computers. This equipment does mathematicaltransformations and statistical analyses of the rawdata from the scanner and transforms it into imagesthat look like brains. By themselves, fMRI imagessimply look like colored blobs, and give little or noindication of the location in the brain where bloodflow increased. To know precisely what parts of thebrain were active, the software must align the fMRIimage with an anatomical MRI, a process known asco-registration. The software can also now correctfor distortion in the images, correct for headmotion, and construct 3-D images of the wholebrain. WKNI scientists are working with GeneralElectric (GE) Medical System’s researchers to automate these aspects of fMRI data processing(Figure 2).

Advantages of fMRIAdvantages of fMRI over other functional mappingtechniques include:1. Repeatability and Safety – a single subject can

be scanned as often as needed without exposureto radioactive isotopes or harmful substances of any kind.

2. Short Scan Times – an fMRI session may last aslittle as 10 – 15 minutes.

3. High Resolution – functional images generallyhave an in-plane resolution of 1.5X1.5 mm.

Another advantage of fMRI is that the same subjectcan be studied multiple times in a longitudinal(long-term) design. Other functional-imaging techniques require injections of radioactive isotopes and cannot be repeated frequently.However, radioisotope functional activation techniques are valuable tools that serve many neuroscience applications.

Uses of fMRIFunctional activity of the brain determined by fMRI is being used for cognitive mapping ofanatomically distinct processing areas in the visualcortex and Broca’s area (associated with speechand language-related activities). In addition, fMRI isbeing used to view complex cognitive tasks such asrecognition, emotion, comprehension, memory,learning, motivation and attention.

This approach has been used to investigate howhealthy subjects learn new complex tasks and,more importantly, to aid in early diagnosis of neurological diseases like stroke and epilepsy. The clinical relevance of fMRI has also beenstrengthened by recent experiments exploring theneurobiological basis of specific rehabilitation

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Figure 2.

strategies intended to aid the recovery of function.The use of fMRI for the pre-surgical mapping of eloquent cortex also suggests that it plays animportant role in the routine management ofpatients with neurological disease. fMRI has beenused to aid the understanding and treatment ofmultiple sclerosis, schizophrenia, depression, bipolar disorder, Parkinson’s disease and addictiveillnesses. Current research by Thickbroom et. al.(2003) involving auditory and visual stimuli isbeing used to map language areas with fMRI. The development of new methods of languagelocalization will lead to new insights into the braincircuitry underlying disorders like aphasia anddyslexia. Bonaffini et. al. (2002) found fMRI usefulfor distinguishing between potentially salvageableand irreversibly damaged tissues in the brains ofstroke and tumor victims (Figure 3).

Current Projects and the Future Direction of WKNI“Brainwave”A time-consuming and often subjective step in theuse of fMRI is the analysis of data. As previouslyoutlined in this article, the images must be reconstructed and corrected for variables like headmovement. Currently, the data output of an fMRIcould take up to a day for manual analysis. In aneffort to provide better care for patients and a morepractical tool for research, the Wallace KetteringNeuroscience Institute has been involved with GEMedical Systems to develop automated software.This software, known as “Brainwave,” efficientlyand accurately analyzes data that is acquired fromthe fMRI, and automatically runs it through thesteps necessary to produce clear, accurate picturesof brain functions. Researchers from surroundinguniversities, including the Ohio State University,the University of Cincinnati, and Wright StateUniversity (WSU), have visited WKNI to view thesecapabilities. WKNI has worked to become one ofthe primary locations in the U.S. for the display ofBrainwave software. The benefits of this software will revolutionize the use of fMRI in research andpatient care.

Schizophrenia Using the rich variety of brain-imaging resourcesfound at WKNI, and the cadre of scientists supportingthese technologies, our hope is to reveal new information about several areas of the brainthought to be involved in schizophrenia, includingthe striatum, the prefrontal cortex and the thalamus. The imaging technologies are capable ofrevealing the white-matter tract architecture of thebrain (responsible for relaying neural signals) and imaging gross neural anatomy and activity.Obtaining an assortment of images from schizophrenic patients and control subjects, andexamining them “in the context of” comprehensivepsychiatric and neurocognitive information, we willbe better equipped to characterize disease-relatedbrain abnormalities than ever before.

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Figure 3.

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Brain Mapping/Localizing Function (Figure 4)1) Epilepsy2) Alzheimer’s disease3) Schizophrenia4) Visual cortex5) Motor cortex6) Complex cognitive functions (i.e., memory,

learning, comprehension, attention, language).

Visual Motion Perception, fMRI and SchizophreniaThe ability to process motion is one of the mostimportant visual capacities that we have, and onethat may reveal something significant about schizophrenia. Schizophrenic patients exhibitimpaired smooth-pursuit eye tracking (the ability tosmoothly follow a moving target) and othermotion-processing deficits. Chen et. al. found evidence that schizophrenic patients have troublewith velocity discrimination when velocity cues areseparated from nonmotion cues like contrast.Perception of the direction of motion is also defective in schizophrenics, and a more recentstudy showed that these patients were impaired inestimating the speed and spatial trajectory of amoving target. This evidence suggests that some ofthe specific impairments affecting schizophrenic

patients are located within the network of motionprocessing. Both visual areas V5 and V1 are likelyinvolved in this task. Understanding the brainregions involved in motion processing and developing tasks that specifically tap into the processing of natural motion used for pursuit eye movements will help identify the nature of thedeficiencies found in motion processing of schizophrenia.

A current study undertaken at WKNI is using fMRI to look for the anatomical/physiological correlates of the observed perceptual abnormalities withinthe primary visual cortex (V1). The stimulus in thefMRI experiment will be a signal dot moving at a fixed velocity and direction through the center of gaze amid a background of random-motion dots. We expect to find a characteristic pattern ofactivation when normal patients are exposed to themoving stimulus, and a specific discrepancybetween the activation patterns of schizophrenicand control subjects.

Characterizing specific activation pattern idiosyncrasies in schizophrenics could greatlyexpand the power of pursuit paradigms to aid inthe diagnosis and treatment of schizophrenia.Activation patterns could become useful in trackingdrug response, course of illness and differentialdiagnosis as well as important familial markers.

In the first phase of a long-term series of such activation studies, we have demonstrated the feasibility of performing such experiments. Our primary long-term objective is to integrate theadvanced neuropsychological findings relating tovisual motion perception with fMRI to better

Figure 4.

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understand aberrant neural networks characterizing schizophrenia. Our colleague at WSU, Dr.Watamaniuk, is a leader in the field of visual motionperception. He has developed a mathematical modelthat describes how cells in our visual system respondto motion. Prior to joining WSU, he was a researchfellow at the Smith-Kettlewell Eye Research Institutein San Francisco (Figure 5).

The Future of fMRINeurosurgery relies on a precise delineation of thestructural and functional aspects of the brain, makingfMRI very significant for neurosurgical planning. Thestructural and functional topography of the brain differsfrom individual to individual, and the need for individualized maps of brain function is enhancedwhen the presence of a tumor alters the typical functional role of a brain region, orwhen the tumor resides in an area witha function that is uncertain. These areasmay include association or language-related cortex. A wide variety of investigators has reported fMRI resultsconsistent with PET, cortical stimulationand magneto-encephalography, servingto confirm that fMRI does indeed provide a source of precise functionaland structural information for neurosurgery.

With the ability to image the entire 3-D volume of the brain, fMRI is capableof isolating many simultaneous andcoordinated brain events. Theseinclude aspects of higher-level cognitivefunctioning that have not previouslybeen scrutinized with such precisionand represent some of the remainingfrontiers in neuroscience.

fMRI has proven itself useful for neurosurgicalplanning, better assessment of risk for individualpatients, improved seizure localization and betterunderstanding of the physiology of neurologicaldisorders. We look ahead to these and other emerging applications as the benefits of this technology become incorporated into current andfuture patient care.

Figure 5.

Acknowledgment: The support of the United States Air Force, Air ForceResearch Laboratory (AFRL/HEOP), Air Force Material Command, undercooperative agreement F33615-98-2-6002 is gratefully acknowledged.

Expanding Neuroimaging to Study Mental IllnessBrad Christian, Ph.D.

The field of medical imaging has made a largeimpact on almost all specialties in medicine, allowingthe radiologist to visualize broken bones, the cardiologist to assess coronary blockage and theoncologist to track tumor growth. In the nearfuture, medical imagingmay arm the psychiatristwith the ability to objectively diagnose mental illness, see the “cleaving”of neural circuits, guidethe clinicians with themost effective course oftherapy and monitorimprovements in patientbrain function. PositronEmission Tomography(PET), which has alreadyestablished itself as aninvaluable tool in wholebody tumor imaging, isalso proving to be a powerful device in studying changes in brain chemistry often implicated in mental illnesses, suchas schizophrenia.

Schizophrenia is a devastating mental disorder thatafflicts approximately 1% of the world population,not discriminating for wealth, race or social status,and has been documented since the beginnings ofwritten history. Our understanding of this diseasehas improved greatly. Owing to scientific research,

schizophrenia is no longer believed to be deservinglycast upon an individual as punishment for sinfulways, or equally misguided, inflicted upon thosewith weak moral character unable to muster up thewill power to overcome their ailments. Modern

science has revealed thatschizophrenia is caused by a disruption in theneurochemical systems ofthe brain, though themechanisms and causesare largely unknown. Theability of PET imaging to study these minisculequantities of chemicals may provide valuableinsights for a greaterunderstanding of theunderlying processes.

It is now well known thatdrugs that are effective for

treating the psychotic symptoms accompanyingschizophrenia, may block the dopamine receptorsin the brain. This has lead to the “dopaminehypothesis of schizophrenia,” predicting overactivetransmission in the brain’s dopamine receptors. Ourresearch in studying schizophrenia has focused onusing neuroimaging methods to study thedopamine systems of the brain. By attachingpositron-emitting atoms to a particular chemical ordrug (termed as a radiotracer) and imaging the brain

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Brad Christian, Ph.D.,

is a Medical Physicist in thePET/Nuclear Medicine

Department at Kettering MedicalCenter Network. His work focuses

on 3-D PET imaging and thedevelopment of mathematical

models to study neuroreceptors.His Ph.D. is from the University of

Wisconsin, which was followedby a fellowship at HarvardUniversity with one of the

pioneering PET groups.

I felt a cleaving in my mind –As if my brain had split –

I tried to match it – seam by seam –But could not make them fit.

The thought behind, I strove to join Unto the thought before –

But sequence ravelled out of soundLike balls – upon a floor.

– Emily Dickinson

Figure 1. A transaxial view of [F-18] fallypride distribution in the human brain. The PET image (center)

is co-registered to an MRI image (left) and displayed as an overlay image (right).

with a PET scanner, it is possible to follow the pathof the radiotracer in the body and see where it isincorporated into a biochemical process (Figure 1).

By developing chemicals that are exquisitely selectivefor the dopamine receptors in the brain, it is nowpossible to visualize “dopamine functional regions”of the brain that have never been seen before(Figure 2). When PET imaging of the dopaminereceptors is used in combination with antipsy-chotic medications, it is possible to directlymeasure the effect of the medication at thedopamine regions in the brain. This will allowthe psychiatrist to fine tune the best course oftherapy for each patient (Figure 3).

Continued advances in neuroimaging technology,and the rapid development of new radiotracers forstudying brain chemistry, place us at a very excitingtime for studying the neurochemical basis of schizophrenia. Hopefully, the day is close when neuroimaging will allow us to truly visualize thesplit in the brain’s wiring, and a course of therapycan be chosen to sew it back together.

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Figure 3. The effects of antipsychotic medications on the uptake of the PET radiotracer([F-18] fallypride) binding. The image on the left shows normal uptake of the radiotracer(baseline) and the three images on the right reveal that less uptake of radiotracer is seenbecause of competition by the antipsychotic medication.

Figure 2. Regions of the brain with very smallamounts of dopamine receptors can

now be visualized using [F-18] fallypride PET imaging.

Acknowledgment: The support of the United States Air Force, Air ForceResearch Laboratory (AFRL/HEOP), Air Force Material Command, undercooperative agreement F33615-98-2-6002 is gratefully acknowledged.

Ongoing WKNI Research ProjectsEpilepsy Imaging ProjectRobert Simkins D.O., FACN

We are studying the utility of 11C-flumazenil PET scanning in temporal lobe epilepsy. Patients with medicallyintractable partial epilepsy who are studied with 11C-flumazenil PET scans are also undergoing extensivediagnostic investigations, including video/EEG monitoring, high-resolution MRI scanning and 18FDG PETscans, as part of a pre-surgical evaluation of their medically intractable epilepsy. Flumazenil attaches to thebenzodiazepine receptors in the brain. The benzodiazepine receptors are a component within the GABAreceptor complex. While we do not have a way of directly imaging the actual GABA receptors with PET scanning, this technique does give us an indirect way of imaging the GABA receptors. GABA receptors areknown to be affected in temporal lobe epilepsy. The absolute numbers of GABA receptors and/or their functional binding characteristics are diminished in epileptic temporal lobe tissue. We are studying the utility of 11C-flumazenil PET scanning for localization of epileptic foci. In addition, we are investigatingchanges in 11C-flumazenil binding distant from the primary epileptogenic focus. Specifically, where are theremote changes in binding located and what are their relationships to the primary epileptogenic focus? Arethe remote changes reversible after resection of the alleged primary epileptogenic focus? Does the finding of abnormalities in 11C-flumazenil binding remote from the suspected primary focus imply additional independent epileptogenic foci and a poor prognosis for seizure control?

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James Olson, Ph.D.,

is a Professor and the Research Director in theDepartment of EmergencyMedicine and also a Professor of Anatomy and Physiology at Wright State University. In addition to funding from WKNI,Dr. Olson’s research is supportedby two grants from the NationalInstitutes of Health. He graduatedfrom Cornell University with aBachelor of Science in EngineeringPhysics and obtained his Ph.D. inBiophysics from the University ofCalifornia at Berkeley. Followingpostdoctoral training at StanfordUniversity Medical School, he tooka faculty position at TulaneUniversity Medical School. Dr. Olson moved to his presentposition at Wright State Universityin 1986.

Ion Channels and Improved Radiation TherapyJames Olson, Ph.D.

All cancer treatments are designed to damage orremove tumor cells, while allowing normal tissue tosurvive. Radiation treatment focuses beams of high-energy particles on the tumor, with a reduced doseof radiation given to adjacent normal cells.Chemotherapy exploits slight differences in the biochemistry of normal and cancerous cells inorder to selectively poison the tumor. However, normal cells are often in the path of the high-energyradiation beam or may be adversely affected by the chemotherapeutic treatment. Thus, to avoidexcessive damage to normal cells, the magnitude ofthe therapeutic dose delivered to the patient is limited. As the dose is reduced, a larger number ofviable cancerous cells will remain after the radiationor chemotherapy treatments, thus increasing therisk that the tumor will reappear.

Research at the Wallace-Kettering NeuroscienceInstitute (WKNI) and other centers has suggestedthat combining radiation and chemical treatmentmethods may enhance the selective killing of tumorcells, and thus provide more effective and safe cancertherapies. This interaction between therapeuticmodalities is explored in detail in a basic researchproject headed by Drs. James Olson and Viney Jainand funded by the Wallace-Kettering NeuroscienceInstitute. The researchers are examining novel drugtreatments that may specifically sensitize tumorcells for damage by radiation treatment or protectcells from injury.

Drs. Olson’s and Jain’s research builds on recentlydiscovered information about the biochemicalmechanisms of cell death. Cells injured by radiationand other causes initiate a built-in self-destruct program called apoptosis (Figure 1). During thisprocess, the outer cell membrane opens specific ion

Figure 1A. Phase contrast photograph of brain cancer cells.

Figure 1B. The nucleus of each cell in Figure 1A is identified with a blue fluorescent stain.

Figure 1C. Cells injured by oxidative stress (simulating damage produced by radiation) are identified by a green fluorescent stain, to indicate which cells have begun the process of programmed cell death called apoptosis.

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channels. This causes the cell to lose intracellularelectrolytes and shrink in size. Subsequently, thecell activates enzymes that digest its internal proteins and chop up its long DNA molecules intoshorter pieces. Eventually these processes bring themetabolic machinery of the cell to a halt and thecell dies. Several researchers have shown that byinhibiting the initial opening of the membrane ionchannels, the programmed progression to celldeath is blocked.

To bring this new information to bear upon thetreatment of patients with brain tumors, Drs. Olsonand Jain are examining membrane ion channels ofhuman cells from normal brain and from braintumors. Their attention is focused on channelsinvolved in cell death following radiation treatment.Because the regulation of membrane channels innormal and tumor cells is different, the researchersanticipate that drug treatments can be designed to

block the function of channels in normal tissue, butnot those of tumor cells. Alternatively, certain drugs may selectively increase the activation of membrane channels in irradiated tumor cells. Suchtreatments would protect the normal brain cellsfrom injury, while increasing the number of tumorcells to undergo the programmed cell death ofapoptosis. For example, after protecting normal cells from apoptosis with a drug treatment, the doseof radiation used to kill cancerous cells may beincreased, thus causing greater damage to thetumor and reducing the risk of it reappearing.These results would directly impact patients at Kettering Medical Center Network – andthroughout the world – by providing a more selectivemeans to destroy tumor cells while maintaining theviability and function of normal brain tissue.

Ongoing WKNI Research ProjectsNew High Magnetic Field MRI Coming to SycamorePeter Roe, B.S., Mehdi Adineh, Ph.D., John German, M.D., Bilal Ezzeddine, Ph.D.

As technology races ahead, changing the world faster and faster, the world of MRI is forever changing andimproving. Sycamore Hospital will be installing a new high magnetic field MRI system this summer. The newmagnet will be a 1.5 Tesla short bore system, which will feel much like an open magnet but with superior imaging qualities. The opening of the magnet is flared and larger than many MRI systems and combines the accuracy of a tunnel scanner with the comfort of an open MRI. The integrated panoramic arrayallows simultaneous scanning with up to four coils. The advantage is revolutionary: you do not have to take the patient off the table to change coils for more than 95% of all studies. Given the improvedtechnology in the gradient strengths and imaging coils, this magnet will provide Sycamore with state-of-the-art MRI capabilities, and high-resolution images when they are needed.

Clinically this new MRI can provide higher resolution capability with stronger and more precise images for diagnosis. It includes new arrays of neuroimaging sequences to allow greater diagnostic capability and accuracy. This more powerful system produces images faster, which adds to the comfort of the patient.

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Women’s Issues in Epilepsy, Part V of VPhillip White, M.D.

Phillip A. White, M.D.,

is board certified in neurology and clinical neurophysiology. Hereceived his medical degree fromBaylor College of Medicine,Houston, Texas. In addition, hereceived his neurology residencyand postdoctoral training in clinical neurophysiology andepilepsy from Baylor College ofMedicine. Dr. White was anAssistant Professor of Neurologyand ran the Epilepsy Clinic atBaylor College of Medicine.

to handle calcium and vitamin D differently, resulting in a higher risk of osteoporosis. The newer antiepileptic medications do not affect theliver in the same way and are now preferred bymost epilepsy experts treating women.

Valproic acid is the “traditional” choice for patientswith primary generalized epilepsy. However, thisagent may cause considerable weight gain, makingit less desirable for many women. Also, valproicacid carries the strongest association with severebirth defects, limiting its use in women of child-bearing years. And thirdly, though still uncertain, there is concern that valproic acid maycontribute to the development of polycystic ovariandisease in some women. While valproic acid is anexcellent agent for primary generalized epilepsy, forwomen of child-bearing years, one of the newermedications, such as Lamotrigine, Topiramate,Levitiracetam or Zonisamide is a better choice.

Until recently, very little attention has been given tothe special concerns of women with epilepsy. Drugstudies largely exclude women because of theunknown effects of new agents on pregnancy. Fromanecdotal experience, there does not appear to beany clear differences in women’s compared to men’sresponses to epilepsy medications; but there areclear differences in side effects, which make someagents better than others for certain groups ofwomen. Approximately one-half of women who suffer from epilepsy will experience some change inthe frequency or timing of seizures throughout thechanges in their menstrual cycle, during pregnancy,and after menopause. We think these differencesare due to estrogen’s effects on seizures, but this isstill a very complicated subject with no clearanswer. One of the primary concerns of women of child-bearing age is the effects of these medications on the developing child.Unfortunately, while we know that there is anincreased risk of birth defects with the older medications, there is still much that we do notknow about the newer medications. This articleaddresses some of these concerns.

Antiepileptic Medications for Women With EpilepsyOther articles in this series have addressed theproper use and potential side effects of both newerand older antiepileptic medications. While there isno evidence that epileptic syndromes in women differ from men, there are reasons to consider using other medications than the older “traditionalagents” for certain women. Older agents, such as Phenytoin, Carbamezepineand Phenobarbital, are the traditional choices for partial onset seizures.However, these medicinesinduce the liver to breakdown other medicationsand nutrients. In particular,they cause the liver tometabolize estrogens rapidly,decreasing the effectivenessof estrogen-containing birth control pills. Threeoptions to counteract this problem would be to use(1) higher dose estrogen-containing birth controlpills, (2) progesterone (such as DepoProvera shots)or (3) an alternative antiepileptic drug. In additionto estrogens, these older agents also cause the liver

Seizures and the Menstrual CycleBetween 30%-40% of women report a catamenialpattern (seizures associated with the menstrualcycle) to their epilepsy. Numerous animal studiesshow that estrogen lowers the threshold forseizures, thereby making seizures more likely tohappen. This may account for seizures occurringaround the time of menses and explain why estrogenreplacement increases seizure frequency in somewomen. However, progesterone treatment is onlyoccasionally effective for control of catamenialseizures, hysterectomy is ineffective for control ofseizures, and seizure frequency may increase,decrease, or stay the same during pregnancy andmenopause. The interactions between hormonesand seizures are, therefore, unclear. We are alsouncertain how knowledge of these interactions willtranslate into effective therapies. At this point, weshould probably view the menstrual cycle as anorganizer of when seizures occur rather than acause of their occurrence.

Epilepsy and PregnancyThe overwhelming majority ofchildren born to women withepilepsy are normal andhealthy. However, there is anincreased risk of birth defectsand need for special educationin children born to womenwith epilepsy, even when nomedicine is taken. For womentaking antiepileptic medication,

the risk for birth defects and mental slowing intheir children is even higher. The older medicationsincrease these risks by about 4-6%, with valproicacid being the most likely drug to cause a major

fetal malformation. Phenytoin, Carbamezepine,Phenobarbital, and Primidone have all been associated with greater risks of birth defects; andtaking more than one medicine also increases thisrisk. The newer medications appear to have a lowerrisk of birth defects in animal studies, but we do notyet know the full impact on humans. Of the neweragents, we have the most information aboutLamotrigine during pregnancy. This medicationappears to be safer than the older drugs, but this isonly preliminary information at best. All of thenewer agents should be considered “unknown”regarding their effects during pregnancy. Despiteour uncertainties, the newer antiepileptic agents(particularly Lamotrigine) are generally preferred.

During pregnancy, seizure frequency increases inone-quarter to one-third of women with epilepsy.Should a woman have a generalized tonic-clonicseizure, there is a risk of her baby being injured, notreceiving enough oxygen, or being spontaneouslyaborted. Because of these concerns, convulsions(tonic-clonic seizures) are considered more dangerousto both mother and child than exposure toantiepileptic medications. Despite the concerns ofbirth defects expressed above, treatment appearsbetter than the alternative.

All antiepileptic agents are present in breast milk.The concentration in breast milk is, however, muchlower than in the blood stream (the concentrationthe baby was exposed to during pregnancy), evenwith Lamotrigine, which may be concentrated at ahigher level than other medications. Every mothermust decide for herself if she wishes to breast-feed;but there is no absolute reason to avoid it becauseof using antiepileptic medication.

Generally, treatment of women with epilepsy doesnot differ significantly from that of men. There are,however, special concerns that must be taken intoaccount when treating women, especially duringchild-bearing years. Most physicians avoid valproicacid and try to use one of the newer antiepilepticagents. All women of child-bearing age should takefolic acid to help minimize the risk of birth defects.Pregnant women should be treated with a singleantiepileptic medication when possible.

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Although surgery and radiation for patients withmalignant cerebral gliomas can prolong survival fora few months, the long-term outlook is dismal. Aspart of a multi-center phase II clinical trial,researchers at the Wallace-Kettering NeuroscienceInstitute (WKNI) at Kettering Medical Center areevaluating a therapy that uses a glucose analogue(2-deoxyglucose, 2DG) to make tumors more sensitive, and surrounding tissue less sensitive, toradiation. Although the therapy has shownpromise, there is wide variation in tumor response.

To better understand the variation in tumorresponse, WKNI has supported studies led by Dr.Viney Jain, WSU Emergency Medicine, and Dr. Steven Berberich, Director of the GeneExpression Laboratory at WSU’s School ofMedicine. Using Affymetrix “GeneChips” – whichcontain small DNA oligomers representing thousands of human genes (Figure 1) – theseresearchers have been systematically examininghow 2-deoxyglucose affects the changes of geneexpression in cultured human gliomas. They hopethe gene expression changes they uncover will beuseful in future clinical trials investigating correlations between treatment-induced changesand clinical response in a large patient population.Establishing a significant correlation would lead todevelopment of predictive assays for therapyresponse and individualization of therapy for optimal benefit to the patients, according to theresearchers.

This novel technology of determining changes ofgene expression using DNA arrays or GeneChips isone of the new technologies that has developed outof the completion of the human genome project. The pharmaceutical industry has begun touse the technology to develop new drugs, as well aspotentially tailoring drugs based on a person’sgenetic makeup. In a few years, pathologists andother physicians are likely to be using DNA arraysfor diagnosing diseases and selecting treatmentsmost appropriate for individual patients.

Figure 1. Affymetrix GeneChip™.

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Steven Berberich, Ph.D.,

is the current Director of the Gene Expression Laboratory atWright State University’s School ofMedicine and is an AssociateProfessor in the Department ofBiochemistry and MolecularBiology. His laboratory studies the regulation of the p53 tumorsuppressor protein and is fundedby the National Cancer Institute.

GeneChip Analysis and GliomasSteven Berberich, Ph.D.

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Figure 3. Outline of a gene expression profiling experiment.

1. Small DNA molecules (oligonucleotides), specific forthe gene of interest, are identified and placed on the

GeneChip probe array. 2. RNA is isolated from tissue orcells and used to create labeled cRNA that is loaded into

the GeneChip. 3. The amount of cRNA that binds to thevarious oligonucleotides (probe features) is determined

by fluorescence. 4. Using computer programs, the fluorescence images are digitized and used in data

analysis programs to develop a profile of gene expression.

Figure 2. Cartoon of the steps to detect those genes that are expressing mRNA (active). 1. Each probe cell or feature contains approximately 1 million identical oligonucleotides of a particular sequence specific for a given gene. 2. Labeled cRNA probe generated from a given sample binds to its “matching” oligonucleotide sequence on the GeneChip.3. The amount of bound cRNA is determined by capturing the fluorescence intensity for each probe cell.

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Affymetrix GeneChips are available now for a hostof species. The human GeneChip being used in the2DG project contains over 12,000 different genes,each identified as a series of spots or “features”arranged on a grid. Each feature is approximately24 microns wide. This array of DNA molecules provides a fixed reference in an assay that beginswith isolating messenger RNA from cell or tissuesamples (Figure 2). The method is as follows. The RNA is converted to a probe, then applied tothe chip. Probe from RNA expressed by specificgenes in the sample bind to their counterparts on the chip (Figure 3). Excess probe for RNA thathasn’t matched with genes on the chip is washedoff, and the chip is placed in a laser scanner thatmeasures the fluorescent intensity of the features. Each feature’s fluorescence is proportional to the amount of probepresent, which is proportional to theamount of RNA present in the sample. Figure 4 represents the fluorescence pattern seen with aGeneChip containing over 12,000human genes.

The resulting gene expression profile indicateswhich genes were active and inactive in the sample.The gene expression profile represents a “snapshot”in time, documenting gene expression at themoment when the RNA was isolated. When examining cells stimulated with a compound like 2-deoxyglucose the snapshot depends on the doseand time of treatment. While many of the cell’sgenes that are expressed will be unaffected by2-deoxyglucose, the others that do fluctuate inresponse to 2-deoxyglucose represent new insightsinto the biological effects that lead to radiosensitizingmalignant gliomas.

Figure 4. HG_U95Av2 GeneChip image.

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Diffusion Tensor Imaging (DTI)Mehdi Adineh, Ph.D.

What Is DTI?Brownian motion describes the properties of waterdiffusion that are affected by the nature of the medium in which it occurs. Diffusion within biological tissues reflects both tissue structure andarchitecture at the microscopic level. In brain tissue,the motion of water molecules can be restricted in many ways (i.e., by interacting with tissue components). Anisotropic diffusion describes howwater diffusion in the white matter of the brain isrestricted in random directions. Observing this typeof diffusion provides a good means of characterizingthe local structure in tissues like muscle and thewhite matter of the brain.

The first attempts to detect diffusion with MR withinthe brain led to the development of diffusionweighted imaging (DWI) and, more recently, to thedevelopment of multidimensional assessment ofdiffusion data known as diffusion tensor imaging(DTI). Gray matter, white matter and CSF (cerebrospinal fluid) in the brain can easily be identified using the current, conventional MRI.However, the appearance of white matter is difficultto accurately observe and quantify. DTI can be usedto explore anisotropic diffusion, and to more precisely analyze the white matter of the brain(Figure 1).

Diffusion MR imaging was introduced in 1986, butwas plagued with technical constraints like motionsensitivity. This limitation provided a major incentive to develop faster sequences that weremore accommodating in terms of bulk motion. Thedevelopment of diffusion sensitive pulse sequencesbasically followed two directions: echo-planar imaging methods, which acquire a complete imagewithin a single shot, and navigator methods, whichacquire images in multiple shots, and use navigatorMR signals to detect and correct the bulk motion.

In the mid ’90s, a more complex technique todescribe multidirectional water diffusion, DTI, wasintroduced to the field of MR diffusion imaging.The term “tensor” was taken from physics and engineering, where it had been used to describetension forces in solid bodies. In MR diffusion tensor imaging, is a tensor that describes diffusionin all spatial directions. The importance of this technique is that it can be used to make detailed, 3-D maps of nerve pathways in the brain.

What Is the Role of DTI in Neurological Assessment?In the absence of any barriers, diffusion is equallyprobable in all directions. Using DTI, a disruptedflow of water can be used to locate places wherethere might be an underlying abnormality. The DTIscan can be compared with the scans of peoplewithout neurological disorders.

Epilepsy:DTI may be a useful tool for gathering informationabout the nature and severity of epilepsy in individualswhose pathology is difficult to characterize withMRI. This technique gives further useful information about the brain and is a major step forward in the ability to pinpoint the causes ofepilepsy, which may lead to successful surgicaltreatment.

Mehdi Adineh, Ph.D.,

is a Research Associate and MRI Physicist at WKNI.

Figure 1.

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Arfanakis et. al. (2002) suggest that DTI may determine the extent of damage to white matterstructures of temporal lobe epileptics. Rugg-Gunnet. al. (2002), using DTI to examine a patient with clinical temporal lobe epilepsy not detectedusing MRI, found an area of abnormal diffusion in the right frontal lobe, which was identified and surgically resected.

Multiple Sclerosis:Tensor diffusion imaging detects changes in thewhite matter of the brain in multiple sclerosis (MS)patients that are not visible in standard MR images.Standard MR images can show plaques (the areas ofwhite matter affected by the disease), but plaquestypically become visible only during the moreadvanced stages of the disease. In addition, MRimages can appear somewhat normal while thepatient is experiencing severe symptoms of MS. Theimages are not always correlated with the currentprogression of the disease. Tensor imaging allows amore accurate detection of abnormalities in MSpatients, which may aid early diagnosis and treatment of the disease.

Werring et. al. (1999) found that DTI does in factdetect structural changes in seemingly normalwhite matter and does discriminate between focallesions of varying severity. Horsfield et. al. (1998),using DTI in patients with MS, found an increase inthe diffusion coefficient of water molecules in the plaques of patients with MS, compared tohealthy brains.

Schizophrenia:DTI can explain neural circuitry abnormalities inschizophrenia that affect brain regions that are notnecessarily structurally proximal, as seen by MRI,but are nonetheless functionally related. DTI investigates brain connectivity (how different areasof the brain interact with one another) and whitematter fiber tracts to better understand the neuropathology of schizophrenia.

When 10 men with schizophrenia and 10 healthymen were scanned using DTI and MR structuralimaging, significant differences were found,indicating widespread alterations in brain whitematter integrity, but not necessarily in white mattervolume Lim et. al. (1999). In order to verify thehypothesized disturbance in frontal-striatal-thalamiccircuitry in schizophrenia, Buchsbaum et. al. (1998)compared results obtained from diffusion tensorimages with Positron Emission Tomography (PET) scans and revealed convergent evidence fordiminished frontal-striatal connectivity.

Developmental Diseases:DTI can be used to investigate developmental diseases such as attention deficit disorders andautism. Tensor imaging can reveal neural abnormalities linked to these diseases, and aid indiagnosis and treatment planning. Cody et. al.(2002) suggest examining frontal-striatal connections with DTI to measure the integrity ofthe white matter connections between the caudate,the pre-frontal cortex, and other regions, in order toexplore the relationship of these pathways tostereotyped behaviors in autism. Based on previousfindings, Fredericksen et. al. (2002) suggest thatDTI can be used to determine if white matter connections are involved in basal ganglia anomaliesin attention deficit hyperactivity disorder (ADHD).

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Epilepsy:For many epileptics, seizures can be found to originate from a very specific area of the brain(called an epileptic focus). WKNI is testing to see ifDTI can be used to localize seizure foci when theycannot be delineated by other means (i.e., MRI orneuropsychological evaluation).

Schizophrenia:WKNI researchers are combining the use of PETand DTI to further the understanding of how thecaudate (a dopamine-rich region of the brain implicated in schizophrenia) communicates with abroader network of brain areas. DTI can be used toinvestigate the connections between the caudateand other areas, and how these connections mightbe interrupted. Silenced communication betweenthese areas of the brain would lead to topographicaldisorganization of function (Figure 3).

Using these imaging techniques in conjunctionwith each other is unique to WKNI, and will enableour researchers to study schizophrenia in detail,extending our knowledge about the neural circuitslinked to schizophrenia.

Active Tensor Imaging Program at theWallace-Kettering Neuroscience Institute Here at WKNI, we have added this powerful newtool to our growing collection of imaging techniques. We boast a 25-direction DTI with automated processing, ensuring timely data analysis.We are using DTI to investigate diseases such asschizophrenia, MS and epilepsy. By using this technology, we are ensuring better patient care forthe Dayton area.

Multiple Sclerosis:Scientists from WKNI and Yale University are currently using DTI to more precisely characterizewhite matter abnormalities caused by MS, whichcould lead to more accurate diagnosis and treatment of specific subtypes of MS (Figure 2).

Figure 2.

Figure 3.

Ongoing WKNI Research Projects

Boonshoft Schizophrenia SymposiumThe Wallace-Kettering Neuroscience Institute’s BoonshoftSchizophrenia Center – in collaboration with the WrightState University School of Medicine’s Department ofPsychiatry – will present the Fall SchizophreniaSymposium on October 28, 2003, from 8:30 a.m. to 5 p.m. The event will be held at Sinclair CommunityCollege’s David H. Ponitz Conference Center, located at 444 West Third Street in downtown Dayton. Foradditional information, please call Aaron Murray at (937) 395-8227.

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Acknowledgment: The support of the United States Air Force, Air ForceResearch Laboratory (AFRL/HEOP), Air Force Material Command, undercooperative agreement F33615-98-2-6002 is gratefully acknowledged.

Figure 4. Fractional Anisotropy

Future Directions of DTICritical evaluation of the diffusion of water in neural tissues is already being investigated in manydiseases, such as MS, autism, and others. DTI canproduce intricate images of soft-tissue structuresthat can aid doctors in studying development,degeneration, disease and aging in soft tissue. The implementation of this technique at very highfield strength (3 Tesla MR) and characterization ofimaging features will extend diffusion tensor capabilities in the future (Figure 4).

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Neuroscience Summer Intern Program at WKNIIn order to provide students an opportunity to gainexperience in the field of medical research, Wallace-Kettering Neuroscience Institute created a summerinternship program, which is in its third year ofexistence. The program allows students to gainhands-on experience in many aspects of medicalresearch by working with medical professionals.Through the summer of 2003, 13 students haveparticipated in this program and expressed overallsatisfaction with their time spent here. The studentshave come from a diverse background of varyingareas of study at different institutions. Some of the interns from the past and present are discussed here:

Katie Kimble is currently a graduatestudent in the field of human factors psychology at ClemsonUniversity. She was an intern atWKNI in the summer of 2001 andspent her time looking at fMRI studies that focused on mappinglanguage areas of the brain, includingBroca’s area and Wernicke’s area.

“I learned a great deal at WKNI – about how to doresearch and conduct studies. It was a very interestingand fun experience!”

John Maffie, currently attendingCornell University to complete adegree in biology and chemistrywith a concentration in psychology,is applying to medical schools andM.D./Ph.D. programs. Primarily, during his time at WKNI, he worked with Bingzhi Shi adapting a synthesis method forF18-flurocholine, a PET tracer to

use at the nuclear chemistry lab with a dramaticallylonger half-life, enabling it to be transported to nearby hospitals for use in tumor detection. In

addition, John assisted Mike Kent, looking at therole of antiphospholipid in multiple sclerosis, andhelped Mehdi Adineh, researching, writing andediting grant applications while observing fMRIand MRI work.

“Working at WKNI gave me the unique opportunity tosee and participate in a diverse cross-section of scientificresearch in a medical setting.”

Robert Nauman (Tom)graduated from the Universityof Dayton in May of 2003with a psychology major and a philosophy minor. He plans to apply to neuroscience Ph.D. programsto attend in the fall of 2004.He began working for WKNIin the summer of 2002, and

continued his internship this summer. He hasworked on developing a battery of language paradigm presentations to be used with functionalMRI (fMRI). He also had the opportunity to workwith Mike Kent doing immunohistochemistry.

“Mehdi and Mike and all of the MRI techs are greatpeople to work with. I’ve been interested in functionalimaging and I’m glad I’ve had the opportunity to learnabout it firsthand.”

Leslie Mathewson will graduatefrom Antioch College in Mayof 2004 with a major in biochemistry and plans toapply to medical schools.She recently began interningat WKNI. She is workingwith Mike Kent, performingscientific literature searchesand investigating multiple

sclerosis and antiphospholipid antibodies usingneuroimaging and neuroimmunological methodsin detecting phospholipid changes in the hippocampus of patients with epilepsy.

“I’ve expanded my knowledge of multiple sclerosis andepilepsy, and I’ve really gotten a chance to apply myclassroom lab experience to an actual project.”

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Brittany Cleveland workedas a student intern duringthe summer of 2001. She edited the final reportfor WKNI’s CooperativeAgreement with theDepartment of Defense. Shecurrently attends BaylorCollege as a pre-med major.

Heather Singleton is a graduate student in the field of human factors and experimental psychologyat the University of Dayton. After completing herMaster’s in May of 2004, she plans to continueresearch work, and may move to the field of sensoryresearch and usability testing. Heather has workedon scientific literature reviews and on IRB proposalsfor research projects. She is creating paradigms (ortasks) to use with fMRI for mapping variousregions of the brain.

“Working on many diverse projects while I’ve been herehas really helped me develop as a professional. I trulyenjoy the work that I do, and the people that I workwith during my time here.”

Ryan Mace is currently attendingthe University of Maryland with a focus in business and pre-law. After college, he plans on applying to law schools in the Washington D.C. area. He is interning this summer,working on the NeuroscienceDirections publications forWKNI, and updating the currentWKNI website.

Christina Hochwalt is currentlystudying zoology and neuroscience at MiamiUniversity. During her time here,she learned how the grantprocess worked. Her major project was a grant for a smokingcessation program for teens athigh risk. Under the assistanceof Dr. Kent, she performed

immunohistochemistry and histology and observedstaining of slides in research studies of patientswith epilepsy and MS.

Jeremy Olson is currently a student at Harvard University,and is considering physics orbiophysics studies in graduateschool. As a WKNI intern, heresearched and worked on astudy to optimize techniques forassaying the quantities of phospholipids in the brain,which would eventually be used

on hippocampal tissue biopsies.

WKNI Scientist ProfileBingzhi Shi, Ph.D., has been a Radiochemist in thePET/Nuclear Medicine Department at KetteringMedical Center Network since 1998. He is currentlyconducting research in the development of radiopharmaceuticals for the study of dopaminereceptor and transporter systems. He received his Masters in Organic Chemistry from BeijingPolytechnic University and a Ph.D. in OrganicChemistry at Iowa State University. After receivinghis Ph.D., he stayed at Iowa State as a research associate in PET until 1994. He then went on tocomplete a postdoctoral fellowship in PET at EmoryUniversity School of Medicine. After finishing his fellowship, Dr. Shi stayed on as a faculty member for the Department of Radiology untilDecember 1997.

Along with his responsibilities to produce routine clinical radiopharmaceuticals, Dr. Shi has implemented an improved method for the production of F-18 Fdopa, gold standard PET tracerfor Parkinson’s disease. He has contributed to various WKNI research projects by adopting automated production of C-11 methionine and C-11

choline for tumor diagnosis, C-11 flumazenil forseizure focus of epilepsy sponsored by the U.S.Department of Defense, and F-18 fallypride for schizophrenia research sponsored by the BoonshoftSchizophrenia Center.

Dr. Shi’s research interests center on applying synthetic radiochemistry technique to develop PETradiopharmaceuticals. He is currently involved inthe design and synthesis of PET biological probesfor the investigation of neurotransmission systems,especially dopamine neurotransmission systems.Recent efforts include the development of PET radioligands of dopamine receptor agonists and dopamine transporter, and their potential application to the studies of neuropsychiatric illnesses, including schizophrenia. Dr. Shi has published a dozen peer-reviewed journal articlesand numerous meeting abstracts and symposiumproceedings. He serves as a member of the KMCNradioactive drug research committee.

Dr. Shi lives in Mason with his wife, Yili Wang, andtheir children, James and Chelsea.

3533 Southern Boulevard, Kettering, Ohio 45429 • 937-395-8002 • www.wkni.org • © 2003

Coming in future issues ...• Spinal Disorder Surgery Techniques• Vertebroplasty• Highlights of the Boonshoft Schizophrenia Symposium• Frameless Stereotactic Surgery• Gamma-Knife Surgery and MR Spectroscopy• Neurotoxins• Neuroscience Specialty Nursing• Software for Image-Guided Spine Surgery

References and Bibliographies for all articles can be found on our website at www.wkni.org

Bingzhi Shi, Ph.D.,

Radiochemist, PET/NuclearMedicine Department,

Kettering Medical Center Network.

For Life.SM

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