FULL-LENGTH ORIGINAL RESEARCH
In vivo imaging of epileptic foci in rats using a miniature probe integrating diffuse optical tomography and electroencephalographic source localization *Hao Yang, *Tao Zhang, †Junli Zhou, *†‡§Paul R. Carney, and *Huabei Jiang Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
SUMMARY
Dr. Hao Yang is a postdoctoral associate at J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, U.S.A.
Objective: The goal of this work is to establish a new dual-modal brain-mapping technique based on diffuse optical tomography (DOT) and electroencephalographic source localization (ESL) that can chronically/intracranially record optical/electroencephalography (EEG) data to precisely map seizures and localize the seizure-onset zone and associated epileptic brain network. Methods: The dual-modal imaging system was employed to image seizures in an experimental acute bicuculline methiodide rat model of focal epilepsy. Depth information derived from DOT was used as constraint in ESL to enhance the image reconstruction. Groups of animals were compared based on localization of seizure foci, either at different positions or at different depths. Results: This novel imaging technique successfully localized the seizure-onset zone in rat induced by bicuculline methiodide injected at a depth of 1, 2, and 3 mm, respectively. The results demonstrated that the incorporation of the depth information from DOT into the ESL image reconstruction resulted in more accurate and reliable ESL images. Although the ESL images showed a horizontal shift of the source localization, the DOT identified the seizure focus accurately. In one case, when the bicuculline methiodide (BMI) was injected at a site outside the field of view (FOV) of the DOT/ ESL interface, ESL gave false-positive detection of the focus, while DOT showed negative detection. Significance: This study represents the first to identify seizure-onset zone using implantable DOT. In addition, the combination of DOT/ESL has never been documented in neuroscience and epilepsy imaging. This technology will enable us to precisely measure the neural activity and hemodynamic response at exactly the same tissue site and at both cortical and subcortical levels. KEY WORDS: Diffuse optical tomography, Electroencephalographic source localization, Epileptic foci.
Epilepsy is a common neurologic syndrome, affecting up to 3% of the worldwide population.1 Although medication is the first line of treatment, for many patients seizures cannot be adequately controlled with medication alone. Many
of these patients with intractable seizures are candidates for epilepsy surgery, which is potentially curative if the epileptogenic focus can be identified and safely removed. Surgical options for patients with partial seizures are based on identification and subsequent surgical treatment of the seizureonset zone, or the minimum cortical volume from which seizures arise.2–4 Although, in the most commonly performed surgery, medial temporal lobe epilepsy, cure rates often exceed 70% in carefully selected patients, cure rates are still much lower in patients with nonlesional neocortical epilepsy.5–10 One reason for such low cure rates is that
Accepted October 29; Early View publication December 19, 2014. Departments of *Biomedical Engineering, †Pediatrics, ‡Neurology, and §Neuroscience, University of Florida, Gainesville, Florida, U.S.A. Address correspondence to Huabei Jiang, Department of Biomedical Engineering, University of Florida, Gainesville, FL 32611, U.S.A. E-mail: [email protected] Wiley Periodicals, Inc. © 2014 International League Against Epilepsy
94
95 Imaging of Epileptic Foci in Rats localization of the seizure-onset zone in neocortical epilepsy is less well understood. Thus, there is unmet need for research focused on identifying the brain regions responsible for generating seizures in neocortical epilepsy syndromes. In an effort to overcome this limitation, several modalities, such as functional magnetic resonance imaging (fMRI), single-photon emission computed tomography (SPECT), and positron emission tomography (PET), have been employed clinically to identify areas of abnormal blood flow and metabolism associated with epileptic form events including the seizure-onset zone.11 Although each of these techniques has its merits, they all have poor temporal resolution (on the scale of several seconds) and does not offer reliable information about seizure onset or propagation patterns. These factors underscore the unmet need for more effective techniques for presurgical mapping of neocortical epilepsy. Diffuse optical tomography (DOT) has been used recently to map in vivo the onset and spread of epileptic events with excellent spatial and temporal resolution.12,13 Meanwhile, electroencephalography (EEG) source localization (ESL) is emerging as a useful technique for the study of temporal brain dynamics in animals and in humans.14–23 Furthermore, use of priori information provided by other imaging modalities to constrain ESL reconstruction has been demonstrated to be an effective approach that can improve the accuracy and reliability of the solution.24–28 By integrating DOT and ESL, a novel technique that can precisely map seizures and localize the seizure-onset zone
and associated epileptic brain network could be realized. The depth information derived from DOT can be used as a priori information about the location of the sources to enhance the inverse solution in ESL, which is a promising approach for better electrical localization of seizures. Herein we integrate DOT and ESL through a miniaturized probe, and demonstrate its ability using an acute rat model of focal epilepsy.
Methods Animals Male Sprague-Dawley rats (Harlan Labs, Indianapolis, IN, U.S.A.) weighing 240–260 g on arrival were allowed 1 week to acclimate to the 12-h light/dark cycle and given food and water ad libitum. All procedures were approved by the University of Florida Animal Care and Use Committee and conducted in accordance with the National Institutes of Health Guide for the Care and Use of Experimental Animals. Imaging system The schematic of the integrated DOT/ESL system is shown in Figure 1. This system has been validated using extensive tissue-mimicking phantom experiments.29 As illustrated in Figure 1, the computer sends a starting signal to the LED controller to sequentially light up 12 LEDs (780 nm) (Epitex Inc., Kyoto, Japan). The light beams are delivered to the measuring interface via fiber optic bundles. Diffusing light received by 13 detection fiber bundles are
Figure 1. Schematic of the integrated DOT/ESL imaging system. Epilepsia ILAE
Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
96 H. Yang et al. converted to electrical signals and preamplified by photodetectors (high sensitivity avalanche photodiode [APD]: C5460-01, Hamamatsu, Shizuoka-ken, Japan). The amplified signals are collected through multi-channel data acquisition (DAQ) boards (NI, PXI-6358). A current driving circuit is designed to drive the LEDs, and a Field Programmable Gate Array (FPGA) core board (NI, PCI-7811) is used to control the LED timing sequence. The output power of each LED is adjustable through its independent DC power supply to achieve optimal signal-to-noise ratio (SNR). In the ESL subsystem, 16 channels of EEG signals from the electrodes are preamplified and digitalized by a multichannel preamplifier (Tucker-Davis Technologies, RA16PA, Alachua, FL, United States) and a fast digital signal processor (Tucker-Davis Technologies, RZ5), respectively. Another data acquisition board is used to collect the processed EEG signals. Figure 2A shows the photograph of the integrated DOT/ESL interface (17 mm 9 17 mm), which is made of aluminum with a weight of about 0.2 g. Sixteen homemade copper plates (2 mm92 mm) attached to the surface of the interface are used as the EEG electrodes in the ESL subsystem. The diameter of each optical fiber bundle is 1.5 mm. The corresponding pattern of source (red) and detector (green) positions for DOT and electrodes (black) positions for ESL is shown in Figure 2B. For the experiments, the scalp and skull of the rat brain were removed and the DOT/ESL interface was placed right at the surface of the cortex (see the inserted photograph in Figure 1). DOT/ESL data were recorded immediately before the electrographic seizure-onset time (resting state), and after the seizure onset.
Subsequently, rats (n = 4) received 10 ll of 1.9 mM bicuculline methiodide (BMI) and 10 ll saline as control at a depth of 1–3 mm below the surface of the right or left parietal cortex covered by DOT/ESL interface, respectively, and other rats (n = 2) were injected the same amount of BMI at a site outside the field of view (FOV) of the DOT/ ESL interface. Each animal was injected with BMI into one site of the brain. The infusion was performed at a rate of 0.3 ll/min. The infusion system consisted of a 100 ll gastight syringe (Hamilton, Reno, NV, U.S.A.) driven by a syringe pump (Cole-Parmer, Vernon Hills, IL, U.S.A.). The injector was mounted on a micromanipulator that allowed precise injections at a depth below the surface.
Animal procedures Animals were placed in a stereotaxic frame and anesthetized: induced with isoflurane (4% for induction and 1.5% for maintenance) and then maintained with urethane (1 g/kg of body weight, intraperitoneal injection). After the scalp was cut and skull was removed, one, 300-lm-diameter stainless steel screw electrode (FHC, Bowdoin, ME, U.S.A.) was implanted as a reference electrode into the occipital bone. Cortical local field potentials were obtained at 12 kHz, digitized with 16 bits of resolution, and bandpass filtered from 0.5 to 6 kHz.
After the dual-modal DOT/ESL system was validated using extensive phantom experiments,29 we first tested the system accuracy for identifying a focal ictal activity over a time window of 6 min. Whereas a significant increase (t-test, p < 0.05) of optical absorption is seen in the region of the BMI injection, no absorption contrast is observed in the resting state (Fig. 3). These results suggest that the increase in local and surrounding brain tissue absorption was mostly due to the local bicuculline-induced seizures. To make sure if this contrast was indeed caused by the seizure onset (not because of the injection of a liquid or
A
Image analysis DOT and ESL images were reconstructed by iteratively solving the photon diffusion equation and Poisson’s equation, respectively, using the nonlinear, finite elementbased reconstruction algorithms described in Jiang (2010).30 The algorithms use a regularized Newton’s method to update an initial optical property (absorption and scattering coefficients)/current source distribution iteratively to minimize an object function composed of a weighted sum of the squared difference between computed and measured optical/local field potential data at the medium surface. To obtain more reliable depth information about the location of the sources from ESL, the depth information derived from DOT was used as a hard-priori to constrain the source space of the electrical source imaging.
Results
B Figure 2. (A) Photograph of the DOT/ESL interface prototype. (B) The pattern of source (red) and detector (green) positions for DOT and electrode (black) positions for ESL. Epilepsia ILAE
Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
97 Imaging of Epileptic Foci in Rats
Figure 3. Detection of seizure focus. Top panel: EEG recordings at resting state (2 min window) and after BMI injection (6 min window). Bottom panel: time series DOT and ESL images overlaid on a structural MRI in the horizontal plane at resting state (t = 0 min) and after the BMI injection (t = 0.65, 2.25, 3.85, and 5.45 min). Epilepsia ILAE
physical contractions), we have performed controlled measurements where the rat was injected 10 ll of saline: no significant contrast (t-test, p > 0.05) in absorption or scattering was observed at the location of the injection. The seizure focus was also clearly identified by the ESL images. The size of the focus detected by ESL, however, was overestimated, and the ESL-identified position of the focus was shifted as well compared to the BMI injection site. In this case, the depth information derived from DOT was used as a priori information to segment the regions in ESL reconstruction. To test the sensitivity of detecting seizure focus at different depths, we injected BMI at depths of 1, 2, and 3 mm below the surface of the right or left sensory cortex (S1) (n = 3),31 and the resulting DOT/ESL images are shown in Figure 4. In ESL, the depth information derived from DOT was also used as a priori information to constrain the ESL inverse solution. The results demonstrated that the constraint derived from DOT effective improved the accuracy and reliability of the ESL solution, especially the depth information of the source. We also see that the position of the focus imaged by ESL is shifted horizontally and its size is overestimated (top panel, Fig. 4), whereas DOT identified the seizure focus more accurately. The volumetric imaging ability of DOT is better appreciated from the coronal section images shown by the bottom panel of Figure 4. We also evaluated the specificity of seizure detection for DOT/ESL. In this study, we injected BMI at a site outside the FOV of the DOT/ESL interface at the left primary motor cortex (M1 [AP: 3.2 mm; ML: 2.5 mm; DV: 1.5 mm]) (Fig. 5A) and the right caudate putamen (CPu [AP: 3.2 mm; ML: 2.5 mm; DV: 1.5 mm]) (Fig. 5B) (n = 2).26 In both cases, from the sagittal section DOT/ESL images shown in Figure 5 we see that ESL gives false-positive detection of the focus, whereas DOT shows negative detection for both cases. These results show that, in this specific case, DOT can correctly detect negative seizure focus for
cases ESL provides false positive detection. We need to point out that we could not use the depth information derived from DOT as the constraint to ESL here because DOT shows negative detection in this case. This case demonstrates the unique advantage of this dual-modal technique for providing the surgeons with complementary and complete information for correctly localizing the seizure-onset zone.
Discussion The main finding of this study is that implantable DOT/ ESL imaging is a novel tool for precisely mapping seizures and localizing the seizure-onset zone. This study investigated the localization of seizure-onset zone at different locations and different depths during focal seizure onset in an acute model of focal epilepsy. This technology will enable us to precisely measure the neural activity and hemodynamic response at exactly the same tissue site and at both cortical and subcortical levels, providing a new tool for unprecedented investigation of neurovascular coupling in epilepsy. To the best of our knowledge, this is the first to identify seizure-onset zone using implantable DOT, and also the first to combine DOT and ESL in a miniature probe. This technique would have a large impact in several areas. The first is that it could complement existing gold standard subdural electrodes techniques. In fact, the multichannel intracranial EEG data recorded by the dual-modal device coupled with a finite element-based reconstruction algorithm allow us to realize EEG-source localization for spatial localization of the seizure-onset zone.18,22,32,33 However, as we pointed out earlier, some works need to be done to improve the reliability of the ESL results. Therefore, this simultaneous DOT/ESL brain mapping provides not only for more precise mapping of the seizure-onset zone and the brain tissue, which is the key to successful epilepsy surgery, but also provides a unique technique for unprecedented Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
98 H. Yang et al. A
B
C
Figure 4. Depth sensitivity of seizure detection. Top panel: DOT and ESL images overlaid on a photograph of the cortex at 1 mm (A), 2 mm (B), and 3 mm (C) depth of BMI injection. Bottom panel: Coronal section DOT and ESL images overlaid on coronal section MRI at 1 mm (A), 2 mm (B), and 3 mm (C) depth of BMI injection. Epilepsia ILAE
A
B
Figure 5. Specificity of seizure detection. Left of (A) and (B): DOT and ESL images overlaid on a photograph of the cortex at 3 mm (A), and 4 mm (B) depth of BMI injection. Right of (A) and (B): Sagittal section DOT and ESL images overlaid on sagittal section MRI at 3 mm (A), and 4 mm (B) depth of BMI injection. The dashed square indicates the field-of-view of the DOT/ESL interface. Epilepsia ILAE
investigation of neurovascular coupling in epilepsy, since DOT images hemodynamics and ESL measures neural activity. In particular, the results demonstrated that it is an effective approach to improve the accuracy of ESL by Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
imposing the constraints derived from DOT. Second, because our previous DOT study detected early hemodynamic responses with heterogeneous patterns several minutes before onset of the electroencephalographic seizure,
99 Imaging of Epileptic Foci in Rats supporting the presence of a “preseizure” state, the device could be used for early seizure detection and prediction.34 Optical signal measures could be combined with closedloop seizure prevention strategies and local treatment. The possibility of seizure prediction has given hope for new warning and therapeutic devices for individuals who cannot be treated successfully with current therapies. Finally, the device could be modified to drive light-sensitive channels (optogenetics) while measuring optical responses in near real-time.35 As a result, an optrode optical mapping device could be used to optically guide plasticity. Although these studies are beyond the scope of this work, they are directly dependent on this initial step, which is to design, build, and test a device for subdural chronic monitoring of optical signals in a realistic animal model of neocortical epilepsy. We found that the shift of the focus position imaged by ESL occurred even the depth information from DOT was imposed to constrain the space of the source localization in ESL. Although the development of ESL reconstruction algorithm is not the focus of this work, we still note the potentials to improve the EEG source localization accuracy with other reconstruction algorithms. In recent years, several algorithms such as sLORETA and MUSCIS, have been developed and compared through extensive simulation studies.19–23 These studies suggest that some key factors, including the number and distribution of EEG electrodes, and the SNR of recording signal, significantly affect the localization accuracy. Similar to the discussion by Michel et al.,19 we believe that the shift of the localization of seizure activity seen in this study was derived mainly from the limited number of EEG electrodes and the plane structure of the probe used in our DOT/ESL interface. One of the limitations of this technique in this study is the probe size, although still smaller than in many other reports of DOT system used in the study of epilepsy. To truly realize an implantable technique, a more miniaturized design is required. Fortunately, with the rapid development of the electronic engineering technology, more and more miniature electronic components, especially the ultra-micro-sized version of the surface mount LEDs and Photodiode, appeared and was widely applied in the field of biomedical imaging.36,37 Based on these new techniques, a miniature probe more suitable for implantation in animal and human brain can be achieved. The inflexible configuration of the interface is also one of the limitations of this design, which may affect the accuracy of ESL imaging. In future, a more flexible material such as medical grade silastic films can be used as the substrate of the sensors. Another limitation is the number of EEG electrodes used in ESL. Theoretical and experimental studies have demonstrated that increased spatial sampling of the interictal EEG will considerably increase the output of interictal EEG recordings in patients with epilepsy.38 Prior studies also indicate that a minimum of 100 electrodes is needed to properly sample the electric
field from the full head surface.39 However, this problem could be easily resolved by decreasing the size of electrode and increasing the electrode number. Despite these limitations, a more sophisticated and implantable dual-model imaging technique is expected in the near future.
Acknowledgments This research was funded in part by a National Institutes of Health (NIH) grant (R01 NS069848), and by the J. Crayton Pruitt Family and the B.J. and Eve Wilder Endowment Funds.
Disclosure None of the authors have any conflicts of interest to disclose. We confirm that we have read the journal’s position on issues involved in ethical publication and affirm that this report is consistent with those guidelines.
References 1. Hauser WA, Annegers JF, Kurland LT. Incidence of epilepsy and unprovoked seizures in Rochester, Minnesota: 1935–1984. Epilepsia 1993;34:453–458. 2. Awad IA, Rosenfeld J, Ahl J, et al. Intractable epilepsy and structural lesions of the brain: mapping, resection strategies, and seizure outcome. Epilepsia 1991;32:179–186. 3. Penfield W, Jasper H. Epilepsy and the functional anatomy of the human brain 1954. 4. Rosenow F, L€uders H. Presurgical evaluation of epilepsy. Brain 2001;124:1683–1700. 5. Cohen-Gadol AA, Wilhelmi BG, Collignon F, et al. Long-term outcome of epilepsy surgery among 399 patients with nonlesional seizure foci including mesial temporal lobe sclerosis. J Neurosurg 2006;104:513–524. 6. Hong K-S, Lee SK, Kim J-Y, et al. Pre-surgical evaluation and surgical outcome of 41 patients with non-lesional neocortical epilepsy. Seizure 2002;11:184–192. 7. Dunlea O, Doherty CP, Farrell M, et al. The Irish epilepsy surgery experience: long-term follow-up. Seizure 2010;19:247–252. 8. Ludvig N, Kuzniecky RI, Baptiste SL, et al. Epidural pentobarbital delivery can prevent locally induced neocortical seizures in rats: the prospect of transmeningeal pharmacotherapy for intractable focal epilepsy. Epilepsia 2006;47:1792–1802. 9. Stacey WC, Litt B. Technology insight: neuroengineering and epilepsy – designing devices for seizure control. Nat Clin Pract Neurol 2008;4:190–201. 10. Theodore WH, Fisher RS. Brain stimulation for epilepsy. Lancet Neurol 2004;3:111–118. 11. Salamon N, Kung J, Shaw S, et al. FDG-PET/MRI coregistration improves detection of cortical dysplasia in patients with epilepsy. Neurology 2008;71:1594–1601. 12. Wang Q, Liang X, Liu Z, et al. Visualizing localized dynamic changes during epileptic seizure onset in vivo with diffuse optical tomography. Med Phys 2008;35:216. 13. Yang J, Zhang T, Yang H, et al. Fast multispectral diffuse optical tomography system for in vivo three-dimensional imaging of seizure dynamics. Appl Opt 2012;51:3461–3469. 14. Nakasatp N, Levesque MF, Barth DS, et al. Comparisons of MEG, EEG, and ECoG source localization in neocortical partial epilepsy in humans. Electroencephalogr Clin Neurophysiol 1994;91:171–178. 15. Lantz G, Grave de Peralta R, Spinelli L, et al. Epileptic source localization with high density EEG: how many electrodes are needed? Clin Neurophysiol 2003;114:63–69. 16. Michel CM, Lantz G, Spinelli L, et al. 128-channel EEG source imaging in epilepsy: clinical yield and localization precision. J Clin Neurophysiol 2004;21:71–83.
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100 H. Yang et al. 17. Benar C-G, Grova C, Kobayashi E, et al. EEG–fMRI of epileptic spikes: concordance with EEG source localization and intracranial EEG. NeuroImage 2006;30:1161–1170. 18. Plummer C, Harvey AS, Cook M. EEG source localization in focal epilepsy: where are we now? Epilepsia 2008;49:201–218. 19. Michel CM, Murray MM, Lantz G, et al. EEG source imaging. Clin Neurophysiol 2004;115:2195–2222. 20. D€ umpelmann M, Fell J, Wellmer J, et al. 3D source localization derived from subdural strip and grid electrodes: a simulation study. Clin Neurophysiol 2009;120:1061–1069. 21. D€ umpelmann M, Ball T, Schulze-Bonhage A. sLORETA allows reliable distributed source reconstruction based on subdural strip and grid recordings. Hum Brain Mapp 2012;33:1172–1188. 22. Zhang Y, van Drongelen W, Kohrman M, et al. Three-dimensional brain current source reconstruction from intra-cranial ECoG recordings. NeuroImage 2008;42:683–695. 23. Keeser D, Padberg F, Reisinger E, et al. Prefrontal direct current stimulation modulates resting EEG and event-related potentials in healthy subjects: a standardized low resolution tomography (sLORETA) study. NeuroImage 2011;55:644–657. 24. Van Veen BD, van Drongelen W, Yuchtman M, et al. Localization of brain electrical activity via linearly constrained minimum variance spatial filtering. IEEE Trans Biomed Eng 1997;44:867–880. 25. Phillips C, Rugg MD, Friston KJ. Anatomically informed basis functions for EEG source localization: combining functional and anatomical constraints. NeuroImage 2002;16:678–695. 26. Mattout J, Phillips C, Penny WD, et al. MEG source localization under multiple constraints: an extended Bayesian framework. NeuroImage 2006;30:753–767. 27. Friston K, Harrison L, Daunizeau J, et al. Multiple sparse priors for the M/EEG inverse problem. NeuroImage 2008;39:1104–1120. 28. Henson RN, Flandin G, Friston KJ, et al. A Parametric Empirical Bayesian framework for fMRI-constrained MEG/EEG source reconstruction. Hum Brain Mapp 2010;31:1512–1531.
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29. Yang H, Jiang H. Design and evaluation of a miniature probe integrating diffuse optical tomography and electroencephalographic source localization. Appl Opt 2013;52:5036–5041. 30. Jiang H. Diffuse optical tomography: principles and applications. New York: Taylor & Francis US; 2011. 31. Paxinos G, Watson C. The rat brain in stereotaxic coordinates. San Diego, CA: Academic Press; 1998. 32. Darvas F, Pantazis D, Kucukaltun-Yildirim E, et al. Mapping human brain function with MEG and EEG: methods and validation. NeuroImage 2004;23:S289–S299. 33. Rullmann M, Anwander A, Dannhauer M, et al. EEG source analysis of epileptiform activity using a 1 mm anisotropic hexahedra finite element head model. NeuroImage 2009;44:399–410. 34. Zhang T, Zhou J, Jiang R, et al. Pre-seizure state identified by diffuse optical tomography. Scientific reports 2014:4. 35. Ji L, Zhou J, Zafar R, et al. Cortical neurovascular coupling driven by stimulation of channelrhodopsin-2. PLoS One 2012;7:e46607. 36. Tokuda T, Miyatani T, Maezawa Y, et al. A CMOS-based on-chip neural interface device equipped with integrated LED array for optogenetics. Engineering in Medicine and Biology Society (EMBC), 2012. Annual International Conference of the IEEE: San Diego, CA; 2012:5146–5149. 37. Kwon KY, Khomenko A, Haq M, et al. Integrated slanted microneedle-LED array for optogenetics. Engineering in Medicine and Biology Society (EMBC), 2013. 35th Annual International Conference of the IEEE: San Diego, CA; 2013:249–252. 38. Koles ZJ. Trends in EEG source localization. Electroencephalogr Clin Neurophysiol 1998;106:127–137. 39. Srinivasan R, Tucker DM, Murias M. Estimating the spatial Nyquist of the human EEG. Behav Res Methods Instrum Comput 1998;30: 8–19.
In vivo imaging of epileptic foci in rats using a miniature probe integrating diffuse optical tomography and electroencephalographic source localization *Hao Yang, *Tao Zhang, †Junli Zhou, *†‡§Paul R. Carney, and *Huabei Jiang Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
SUMMARY
Dr. Hao Yang is a postdoctoral associate at J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, U.S.A.
Objective: The goal of this work is to establish a new dual-modal brain-mapping technique based on diffuse optical tomography (DOT) and electroencephalographic source localization (ESL) that can chronically/intracranially record optical/electroencephalography (EEG) data to precisely map seizures and localize the seizure-onset zone and associated epileptic brain network. Methods: The dual-modal imaging system was employed to image seizures in an experimental acute bicuculline methiodide rat model of focal epilepsy. Depth information derived from DOT was used as constraint in ESL to enhance the image reconstruction. Groups of animals were compared based on localization of seizure foci, either at different positions or at different depths. Results: This novel imaging technique successfully localized the seizure-onset zone in rat induced by bicuculline methiodide injected at a depth of 1, 2, and 3 mm, respectively. The results demonstrated that the incorporation of the depth information from DOT into the ESL image reconstruction resulted in more accurate and reliable ESL images. Although the ESL images showed a horizontal shift of the source localization, the DOT identified the seizure focus accurately. In one case, when the bicuculline methiodide (BMI) was injected at a site outside the field of view (FOV) of the DOT/ ESL interface, ESL gave false-positive detection of the focus, while DOT showed negative detection. Significance: This study represents the first to identify seizure-onset zone using implantable DOT. In addition, the combination of DOT/ESL has never been documented in neuroscience and epilepsy imaging. This technology will enable us to precisely measure the neural activity and hemodynamic response at exactly the same tissue site and at both cortical and subcortical levels. KEY WORDS: Diffuse optical tomography, Electroencephalographic source localization, Epileptic foci.
Epilepsy is a common neurologic syndrome, affecting up to 3% of the worldwide population.1 Although medication is the first line of treatment, for many patients seizures cannot be adequately controlled with medication alone. Many
of these patients with intractable seizures are candidates for epilepsy surgery, which is potentially curative if the epileptogenic focus can be identified and safely removed. Surgical options for patients with partial seizures are based on identification and subsequent surgical treatment of the seizureonset zone, or the minimum cortical volume from which seizures arise.2–4 Although, in the most commonly performed surgery, medial temporal lobe epilepsy, cure rates often exceed 70% in carefully selected patients, cure rates are still much lower in patients with nonlesional neocortical epilepsy.5–10 One reason for such low cure rates is that
Accepted October 29; Early View publication December 19, 2014. Departments of *Biomedical Engineering, †Pediatrics, ‡Neurology, and §Neuroscience, University of Florida, Gainesville, Florida, U.S.A. Address correspondence to Huabei Jiang, Department of Biomedical Engineering, University of Florida, Gainesville, FL 32611, U.S.A. E-mail: [email protected] Wiley Periodicals, Inc. © 2014 International League Against Epilepsy
94
95 Imaging of Epileptic Foci in Rats localization of the seizure-onset zone in neocortical epilepsy is less well understood. Thus, there is unmet need for research focused on identifying the brain regions responsible for generating seizures in neocortical epilepsy syndromes. In an effort to overcome this limitation, several modalities, such as functional magnetic resonance imaging (fMRI), single-photon emission computed tomography (SPECT), and positron emission tomography (PET), have been employed clinically to identify areas of abnormal blood flow and metabolism associated with epileptic form events including the seizure-onset zone.11 Although each of these techniques has its merits, they all have poor temporal resolution (on the scale of several seconds) and does not offer reliable information about seizure onset or propagation patterns. These factors underscore the unmet need for more effective techniques for presurgical mapping of neocortical epilepsy. Diffuse optical tomography (DOT) has been used recently to map in vivo the onset and spread of epileptic events with excellent spatial and temporal resolution.12,13 Meanwhile, electroencephalography (EEG) source localization (ESL) is emerging as a useful technique for the study of temporal brain dynamics in animals and in humans.14–23 Furthermore, use of priori information provided by other imaging modalities to constrain ESL reconstruction has been demonstrated to be an effective approach that can improve the accuracy and reliability of the solution.24–28 By integrating DOT and ESL, a novel technique that can precisely map seizures and localize the seizure-onset zone
and associated epileptic brain network could be realized. The depth information derived from DOT can be used as a priori information about the location of the sources to enhance the inverse solution in ESL, which is a promising approach for better electrical localization of seizures. Herein we integrate DOT and ESL through a miniaturized probe, and demonstrate its ability using an acute rat model of focal epilepsy.
Methods Animals Male Sprague-Dawley rats (Harlan Labs, Indianapolis, IN, U.S.A.) weighing 240–260 g on arrival were allowed 1 week to acclimate to the 12-h light/dark cycle and given food and water ad libitum. All procedures were approved by the University of Florida Animal Care and Use Committee and conducted in accordance with the National Institutes of Health Guide for the Care and Use of Experimental Animals. Imaging system The schematic of the integrated DOT/ESL system is shown in Figure 1. This system has been validated using extensive tissue-mimicking phantom experiments.29 As illustrated in Figure 1, the computer sends a starting signal to the LED controller to sequentially light up 12 LEDs (780 nm) (Epitex Inc., Kyoto, Japan). The light beams are delivered to the measuring interface via fiber optic bundles. Diffusing light received by 13 detection fiber bundles are
Figure 1. Schematic of the integrated DOT/ESL imaging system. Epilepsia ILAE
Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
96 H. Yang et al. converted to electrical signals and preamplified by photodetectors (high sensitivity avalanche photodiode [APD]: C5460-01, Hamamatsu, Shizuoka-ken, Japan). The amplified signals are collected through multi-channel data acquisition (DAQ) boards (NI, PXI-6358). A current driving circuit is designed to drive the LEDs, and a Field Programmable Gate Array (FPGA) core board (NI, PCI-7811) is used to control the LED timing sequence. The output power of each LED is adjustable through its independent DC power supply to achieve optimal signal-to-noise ratio (SNR). In the ESL subsystem, 16 channels of EEG signals from the electrodes are preamplified and digitalized by a multichannel preamplifier (Tucker-Davis Technologies, RA16PA, Alachua, FL, United States) and a fast digital signal processor (Tucker-Davis Technologies, RZ5), respectively. Another data acquisition board is used to collect the processed EEG signals. Figure 2A shows the photograph of the integrated DOT/ESL interface (17 mm 9 17 mm), which is made of aluminum with a weight of about 0.2 g. Sixteen homemade copper plates (2 mm92 mm) attached to the surface of the interface are used as the EEG electrodes in the ESL subsystem. The diameter of each optical fiber bundle is 1.5 mm. The corresponding pattern of source (red) and detector (green) positions for DOT and electrodes (black) positions for ESL is shown in Figure 2B. For the experiments, the scalp and skull of the rat brain were removed and the DOT/ESL interface was placed right at the surface of the cortex (see the inserted photograph in Figure 1). DOT/ESL data were recorded immediately before the electrographic seizure-onset time (resting state), and after the seizure onset.
Subsequently, rats (n = 4) received 10 ll of 1.9 mM bicuculline methiodide (BMI) and 10 ll saline as control at a depth of 1–3 mm below the surface of the right or left parietal cortex covered by DOT/ESL interface, respectively, and other rats (n = 2) were injected the same amount of BMI at a site outside the field of view (FOV) of the DOT/ ESL interface. Each animal was injected with BMI into one site of the brain. The infusion was performed at a rate of 0.3 ll/min. The infusion system consisted of a 100 ll gastight syringe (Hamilton, Reno, NV, U.S.A.) driven by a syringe pump (Cole-Parmer, Vernon Hills, IL, U.S.A.). The injector was mounted on a micromanipulator that allowed precise injections at a depth below the surface.
Animal procedures Animals were placed in a stereotaxic frame and anesthetized: induced with isoflurane (4% for induction and 1.5% for maintenance) and then maintained with urethane (1 g/kg of body weight, intraperitoneal injection). After the scalp was cut and skull was removed, one, 300-lm-diameter stainless steel screw electrode (FHC, Bowdoin, ME, U.S.A.) was implanted as a reference electrode into the occipital bone. Cortical local field potentials were obtained at 12 kHz, digitized with 16 bits of resolution, and bandpass filtered from 0.5 to 6 kHz.
After the dual-modal DOT/ESL system was validated using extensive phantom experiments,29 we first tested the system accuracy for identifying a focal ictal activity over a time window of 6 min. Whereas a significant increase (t-test, p < 0.05) of optical absorption is seen in the region of the BMI injection, no absorption contrast is observed in the resting state (Fig. 3). These results suggest that the increase in local and surrounding brain tissue absorption was mostly due to the local bicuculline-induced seizures. To make sure if this contrast was indeed caused by the seizure onset (not because of the injection of a liquid or
A
Image analysis DOT and ESL images were reconstructed by iteratively solving the photon diffusion equation and Poisson’s equation, respectively, using the nonlinear, finite elementbased reconstruction algorithms described in Jiang (2010).30 The algorithms use a regularized Newton’s method to update an initial optical property (absorption and scattering coefficients)/current source distribution iteratively to minimize an object function composed of a weighted sum of the squared difference between computed and measured optical/local field potential data at the medium surface. To obtain more reliable depth information about the location of the sources from ESL, the depth information derived from DOT was used as a hard-priori to constrain the source space of the electrical source imaging.
Results
B Figure 2. (A) Photograph of the DOT/ESL interface prototype. (B) The pattern of source (red) and detector (green) positions for DOT and electrode (black) positions for ESL. Epilepsia ILAE
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97 Imaging of Epileptic Foci in Rats
Figure 3. Detection of seizure focus. Top panel: EEG recordings at resting state (2 min window) and after BMI injection (6 min window). Bottom panel: time series DOT and ESL images overlaid on a structural MRI in the horizontal plane at resting state (t = 0 min) and after the BMI injection (t = 0.65, 2.25, 3.85, and 5.45 min). Epilepsia ILAE
physical contractions), we have performed controlled measurements where the rat was injected 10 ll of saline: no significant contrast (t-test, p > 0.05) in absorption or scattering was observed at the location of the injection. The seizure focus was also clearly identified by the ESL images. The size of the focus detected by ESL, however, was overestimated, and the ESL-identified position of the focus was shifted as well compared to the BMI injection site. In this case, the depth information derived from DOT was used as a priori information to segment the regions in ESL reconstruction. To test the sensitivity of detecting seizure focus at different depths, we injected BMI at depths of 1, 2, and 3 mm below the surface of the right or left sensory cortex (S1) (n = 3),31 and the resulting DOT/ESL images are shown in Figure 4. In ESL, the depth information derived from DOT was also used as a priori information to constrain the ESL inverse solution. The results demonstrated that the constraint derived from DOT effective improved the accuracy and reliability of the ESL solution, especially the depth information of the source. We also see that the position of the focus imaged by ESL is shifted horizontally and its size is overestimated (top panel, Fig. 4), whereas DOT identified the seizure focus more accurately. The volumetric imaging ability of DOT is better appreciated from the coronal section images shown by the bottom panel of Figure 4. We also evaluated the specificity of seizure detection for DOT/ESL. In this study, we injected BMI at a site outside the FOV of the DOT/ESL interface at the left primary motor cortex (M1 [AP: 3.2 mm; ML: 2.5 mm; DV: 1.5 mm]) (Fig. 5A) and the right caudate putamen (CPu [AP: 3.2 mm; ML: 2.5 mm; DV: 1.5 mm]) (Fig. 5B) (n = 2).26 In both cases, from the sagittal section DOT/ESL images shown in Figure 5 we see that ESL gives false-positive detection of the focus, whereas DOT shows negative detection for both cases. These results show that, in this specific case, DOT can correctly detect negative seizure focus for
cases ESL provides false positive detection. We need to point out that we could not use the depth information derived from DOT as the constraint to ESL here because DOT shows negative detection in this case. This case demonstrates the unique advantage of this dual-modal technique for providing the surgeons with complementary and complete information for correctly localizing the seizure-onset zone.
Discussion The main finding of this study is that implantable DOT/ ESL imaging is a novel tool for precisely mapping seizures and localizing the seizure-onset zone. This study investigated the localization of seizure-onset zone at different locations and different depths during focal seizure onset in an acute model of focal epilepsy. This technology will enable us to precisely measure the neural activity and hemodynamic response at exactly the same tissue site and at both cortical and subcortical levels, providing a new tool for unprecedented investigation of neurovascular coupling in epilepsy. To the best of our knowledge, this is the first to identify seizure-onset zone using implantable DOT, and also the first to combine DOT and ESL in a miniature probe. This technique would have a large impact in several areas. The first is that it could complement existing gold standard subdural electrodes techniques. In fact, the multichannel intracranial EEG data recorded by the dual-modal device coupled with a finite element-based reconstruction algorithm allow us to realize EEG-source localization for spatial localization of the seizure-onset zone.18,22,32,33 However, as we pointed out earlier, some works need to be done to improve the reliability of the ESL results. Therefore, this simultaneous DOT/ESL brain mapping provides not only for more precise mapping of the seizure-onset zone and the brain tissue, which is the key to successful epilepsy surgery, but also provides a unique technique for unprecedented Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
98 H. Yang et al. A
B
C
Figure 4. Depth sensitivity of seizure detection. Top panel: DOT and ESL images overlaid on a photograph of the cortex at 1 mm (A), 2 mm (B), and 3 mm (C) depth of BMI injection. Bottom panel: Coronal section DOT and ESL images overlaid on coronal section MRI at 1 mm (A), 2 mm (B), and 3 mm (C) depth of BMI injection. Epilepsia ILAE
A
B
Figure 5. Specificity of seizure detection. Left of (A) and (B): DOT and ESL images overlaid on a photograph of the cortex at 3 mm (A), and 4 mm (B) depth of BMI injection. Right of (A) and (B): Sagittal section DOT and ESL images overlaid on sagittal section MRI at 3 mm (A), and 4 mm (B) depth of BMI injection. The dashed square indicates the field-of-view of the DOT/ESL interface. Epilepsia ILAE
investigation of neurovascular coupling in epilepsy, since DOT images hemodynamics and ESL measures neural activity. In particular, the results demonstrated that it is an effective approach to improve the accuracy of ESL by Epilepsia, 56(1):94–100, 2015 doi: 10.1111/epi.12880
imposing the constraints derived from DOT. Second, because our previous DOT study detected early hemodynamic responses with heterogeneous patterns several minutes before onset of the electroencephalographic seizure,
99 Imaging of Epileptic Foci in Rats supporting the presence of a “preseizure” state, the device could be used for early seizure detection and prediction.34 Optical signal measures could be combined with closedloop seizure prevention strategies and local treatment. The possibility of seizure prediction has given hope for new warning and therapeutic devices for individuals who cannot be treated successfully with current therapies. Finally, the device could be modified to drive light-sensitive channels (optogenetics) while measuring optical responses in near real-time.35 As a result, an optrode optical mapping device could be used to optically guide plasticity. Although these studies are beyond the scope of this work, they are directly dependent on this initial step, which is to design, build, and test a device for subdural chronic monitoring of optical signals in a realistic animal model of neocortical epilepsy. We found that the shift of the focus position imaged by ESL occurred even the depth information from DOT was imposed to constrain the space of the source localization in ESL. Although the development of ESL reconstruction algorithm is not the focus of this work, we still note the potentials to improve the EEG source localization accuracy with other reconstruction algorithms. In recent years, several algorithms such as sLORETA and MUSCIS, have been developed and compared through extensive simulation studies.19–23 These studies suggest that some key factors, including the number and distribution of EEG electrodes, and the SNR of recording signal, significantly affect the localization accuracy. Similar to the discussion by Michel et al.,19 we believe that the shift of the localization of seizure activity seen in this study was derived mainly from the limited number of EEG electrodes and the plane structure of the probe used in our DOT/ESL interface. One of the limitations of this technique in this study is the probe size, although still smaller than in many other reports of DOT system used in the study of epilepsy. To truly realize an implantable technique, a more miniaturized design is required. Fortunately, with the rapid development of the electronic engineering technology, more and more miniature electronic components, especially the ultra-micro-sized version of the surface mount LEDs and Photodiode, appeared and was widely applied in the field of biomedical imaging.36,37 Based on these new techniques, a miniature probe more suitable for implantation in animal and human brain can be achieved. The inflexible configuration of the interface is also one of the limitations of this design, which may affect the accuracy of ESL imaging. In future, a more flexible material such as medical grade silastic films can be used as the substrate of the sensors. Another limitation is the number of EEG electrodes used in ESL. Theoretical and experimental studies have demonstrated that increased spatial sampling of the interictal EEG will considerably increase the output of interictal EEG recordings in patients with epilepsy.38 Prior studies also indicate that a minimum of 100 electrodes is needed to properly sample the electric
field from the full head surface.39 However, this problem could be easily resolved by decreasing the size of electrode and increasing the electrode number. Despite these limitations, a more sophisticated and implantable dual-model imaging technique is expected in the near future.
Acknowledgments This research was funded in part by a National Institutes of Health (NIH) grant (R01 NS069848), and by the J. Crayton Pruitt Family and the B.J. and Eve Wilder Endowment Funds.
Disclosure None of the authors have any conflicts of interest to disclose. We confirm that we have read the journal’s position on issues involved in ethical publication and affirm that this report is consistent with those guidelines.
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