1. Focused ultrasound reduces epileptic EEG bursts Department of Radiology Brigham and Women's Hospital Harvard Medical School Byoung-Kyong Min

2. Introduction Method Result Discussion Treatment on Neurological Disorders Representative non-invasive treatment on the brain: Medication However, it has side-effect and non-spatially specificity

3. Introduction Method Result Discussion Non-pharmacological neuro-modulation Invasive tools (e.g. EpCS, DBS) Non-invasive tools (e.g. TMS, tDCS) Adapted from Hoy and Fitzgerald, Nature Review/Neurology, 2010 tDCS DBS TMS EpCS Adapted from Hoy and Fitzgerald, Nature Review/Neurology, 2010

4. Image-guided, non-invasive, spatially-accurate focused- ultrasound (FUS) could be a potent tool for neuro-modulation. Introduction Method Result Discussion Focused-Ultrasound Sonication (FUS) L1L1 L2L2 IR Marker Transducer Laser guide Motion camera Exablate (Insightec and GE): Array of small 1000 transducers

5. Introduction Method Result Discussion Fig. TBD: tone-burst-duration, PRF: pulse repetition frequency, AI: acoustic intensity Idea: Pulsed application of FUS To avoid heating the tissue, pulsed sonication is used rather than continuous sonication. FUS suppresses VEP in LGN-sonicated cats (Fry, et al. 1958) FUS affects the neurophysiology of in vitro local neural circuitry (Bachtold, et al. 1998, Rinaldi, et al. 1991) FUS can temporarily modify the excitability of the neuronal tissue (Gavrilov et al. 1996) Previous observations by FUS

6. Introduction Method Result Discussion A B FUS transducer B Superior Inferior A Caudal Rostral Right Lef t Right Lef t C * Minutes B Pre Sonication Post Sonication Recovery A Fig. Visual evoked potentials (A) and normalized amplitudes of the p30 components (B) Fig. FUS-mediated fMRI activation maps of the motor cortex (A & B) and FUS-mediated BOLD signal time course (C: gray bar: sonication) 1. Excitation (Yoo et al., 2008; 2009) 2. Suppression (Yoo et al., 2008; 2009)

7. We were motivated to examine if the FUS could suppress hyper-excitability of neural tissue based on a chemical kindling model of acute-stage epilepsy. Epilepsy is a chronic neurological disorder (~50 million), and is characterized by seizures (abnormal hyper-excitability of neurons). Since PTZ (pentylenetetrazol) was used to induce epileptic activity and progressive increments of theta activity has been reported during PTZ-induced epilepsy, not only raw EEG but also its theta band was assessed. Introduction Method Result Discussion Application to Epilepsy suppression

8. Sprague–Dawley rats (275±30g) Group 1 (PTZ(+)/FUS(+); n=9), Group 2 (PTZ(+)/FUS(-) ; n=9), Group 3 (PTZ(-)/FUS(+) ; n=9) PTZ :GABA A receptor antagonist  45 mg/kg in 0.4 mL saline Introduction Method Result Discussion Fig. A diagram of the experimental apparatus Experimental Setup & Design

9. Introduction Method Result Discussion Transducer Characterization Time Space peakaverage peakI SPTP I SATP averageI SPTA I SATA Mechanical Index (MI): the maximum peak negative pressure (P r,α ) of an ultrasound longitudinal wave divided by the square root of its center frequency (C MI ) FUS transducer: 690 KHz, 7cm ROC, 6cm OD, 0.5 ms TBD, 100Hz PRF, 130 mW/cm 2 (I spta )=2.6 W/cm 2 (I sppa ) Hydrophone Fig. Transducer characterization Transducer

10. EEG measures: sub-dermal electrodes (5 mm lateral to the midline & 7 mm anterior to the lambda), 1KHz sampling rate Counting the number of raw EEG and theta bursts (4-8 Hz) exceeding the determined threshold (baseline σ × 4.75) in each session. Racine scoring & Histological analysis Statistics: Independent t-test (one-tailed) between the two groups, and paired t-test (one-tailed) within each animal. In order to compare body weights, a repeated-measures ANOVA with a covariance of the individual body weight before the experiment was applied. Baseline (10 min) Post1 (10 min) Pre-FUS (10 min) PTZ Full ictal Anesthesia FUS1 (3min) FUS2 (3min) Block-A Block-B Block-CBlock-DBlock-E Block-F Post2 (10 min) Introduction Method Result Discussion Fig. Flowchart of the EEG acquisition and FUS sonication

11. Raw EEG EEG theta 100 μV 20 μV 10 sec 20 μV 1 min 100 μV FUS2 FUS1 10 sec Introduction Method Result Discussion Fig. The sample time-courses of EEG recordings from PTZ- induced epileptic rats with sonication. A. Sample EEGs from Group 1 (PTZ(+)/FUS(+))

12. Raw EEG EEG theta 100 μV 20 μV 10 sec 20 μV 1 min 100 μV 10 sec Introduction Method Result Discussion Fig. The sample time-courses of EEG recordings from PTZ- induced epileptic rats without sonication. B. Sample EEGs from Group 2 (PTZ(+)/FUS(-))

13. After sonication, the number of epileptic EEG bursts decreased. (‘Post1’: t(16)= -1.74; ‘FUS2’: t(16)= -2.03; ‘Post2’: t(16)= -1.72). After 2 nd sonication, the number of theta EEG bursts decreased. (‘Post2’: t(16)= -1.98) A B Introduction Method Result Discussion Group Analysis Fig. Comparison of the average number of threshold-exceeding raw (upper) and theta (lower) EEG peaks between the FUS-treated and untreated groups.

14. The number of epileptic EEG bursts within the FUS-treated group was significantly reduced after the sonication period (‘Post1’: t(8)= 2.26; ‘FUS2’: t(8)= 1.91; ‘Post2’: t(8)= 2.58). The number of EEG theta peaks was significantly reduced during (63.0% reduction) and after (up to 68.5% reduction) the second sonication (‘FUS2’: t(8)= 2.81; ‘Post2’: t(8)= 3.14). Racine scores of the FUS-treated group during a day after the experiment were significantly lower than those of the control group (t(15)= -2.41; FUS-treated group: 0.33; Control group: 1.13). Introduction Method Result Discussion Summary

15. Introduction Method Result Discussion Fig. Exemplary histological data obtained from Group 3 (left) H&E staining (right) TUNEL staining (DAPI in blue, apoptotic cell in green) Histological Analysis

16. The low-power, pulsed FUS sonication suppressed the number of epileptic EEG signal bursts without any significant tissue damages. –Stretch-sensitive ion channels (e.g. the novel chloride channels) may be involved in modifying the excitability of neural tissue. –Local hyperpolarization of the cell membrane would eventually raise the threshold for eliciting the epileptogenic activity. –Synaptic contacts could be disrupted by ultrasound, reducing the propagation of the epileptic activity across the brain. –Regulation of thalamic GABAergic inhibitory interneurons; PTZ  a GABA A receptor antagonist Therefore, FUS could provide a new non-invasive treatment of epileptic seizure. Introduction Method Result Discussion Discussion

17. Intra-brain injection of KA (e.g. Hippocampus or amygdala) and evaluation of FUS on suppression of chronic focal epilepsy. Assessment of neurotransmitter modulation associated with sonication (Microdialysis). Stereotactic guidance: MRI-compatible stereotactic positioning system Acoustic radiation force impulse (ARFI) imaging Introduction Method Result Discussion Future Works

18. Seung-Schik Yoo, Krisztina Fischer, Yongzhi Zhang, Ferenc A. Jolesz: Department of Radiology, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA, USA Alexander Bystritsky: The Semel Institute for Neuroscience and Human Behavior, UCLA, LA, CA, USA Kwang-Ik Jung: Department of Physical Medicine & Rehabilitation, Hallym University Sacred Heart Hospital, Korea Lee-So Maeng, Sang In Park, Yong-An Chung: Institute of Catholic Integrative Medicine (ICIM), Incheon Saint Mary’s Hospital, Korea Introduction Method Result Discussion Acknowledgements