Focused ultrasound-mediated suppression of chemically-induced acute epileptic EEG activity

Byoung-Kyong Min, Alexander Bystritsky, Kwang-Ik Jung, Krisztina Fischer, Yongzhi Zhang, Lee-So Maeng, Sang In Park, Yong-An Chung, Ferenc A Jolesz, Seung-Schik Yoo, Byoung-Kyong Min, Alexander Bystritsky, Kwang-Ik Jung, Krisztina Fischer, Yongzhi Zhang, Lee-So Maeng, Sang In Park, Yong-An Chung, Ferenc A Jolesz, Seung-Schik Yoo

Abstract

Background: Epilepsy is a common neurological disorder, which is attributed to uncontrollable abnormal hyper-excitability of neurons. We investigated the feasibility of using low-intensity, pulsed radiation of focused ultrasound (FUS) to non-invasively suppress epileptic activity in an animal model (rat), which was induced by the intraperitonial injection of pentylenetetrazol (PTZ).

Results: After the onset of induced seizures, FUS was transcranially administered to the brain twice for three minutes each while undergoing electroencephalographic (EEG) monitoring. An air-backed, spherical segment ultrasound transducer (diameter: 6 cm; radius-of-curvature: 7 cm) operating at a fundamental frequency of 690 KHz was used to deliver a train of 0.5 msec-long pulses of sonication at a repetitive rate of 100 Hz to the thalamic areas of the brain. The acoustic intensity (130 mW/cm2) used in the experiment was sufficiently within the range of safety guidelines for the clinical ultrasound imaging. The occurrence of epileptic EEG bursts from epilepsy-induced rats significantly decreased after sonication when it was compared to the pre-sonication epileptic state. The PTZ-induced control group that did not receive any sonication showed a sustained number of epileptic EEG signal bursts. The animals that underwent sonication also showed less severe epileptic behavior, as assessed by the Racine score. Histological analysis confirmed that the sonication did not cause any damage to the brain tissue.

Conclusions: These results revealed that low-intensity, pulsed FUS sonication suppressed the number of epileptic signal bursts using acute epilepsy model in animal. Due to its non-invasiveness and spatial selectivity, FUS may offer new perspectives for a possible non-invasive treatment of epilepsy.

Figures

Figure 1
Figure 1
A diagram of the experimental apparatus and the topographical arrangement of the sonication path and electrodes. (A) The transducer was mounted on a 3-axis stereotactic positioning system and submerged in degassed water. The coordinates of the acoustic focus were controlled under the metric guidance of the stereotactic coordinates of the rat brain, which were marked on a plastic plate. After localization of the acoustic focus, the animal was laid supine on a plastic tray that was mounted above the system. The head was partially submerged in an oval hole that opened into a bag of degassed water in order to secure an uninterrupted beam profile from the transducer to the targeted tissue. (B) Schematic diagram of the positions of bilateral subdermal EEG electrodes (marked as small blue circles) relative to the area of the sonication beam path (marked as a red circular boundary) on the rat skull as well as to the sonication focus (marked as a solid red circle).
Figure 2
Figure 2
Flowchart of the EEG acquisition and FUS sonication. Block-A represents the baseline period. The baseline EEG was recorded for ten minutes after the EEG signals stabilized following the administration of anesthesia. Block-B (named as 'Pre-FUS') indicates the time-interval after observing significant evidence of ictal behavior (e.g., bilateral forepaw-twitches) and just before the first sonication. Block-C represents the three-minute period of the first sonication (named as 'FUS1'), and Block-E represents the second three-minute sonication interval (named as 'FUS2'). Block-D represents the time-interval after the first sonication (named as 'Post1'), and Block-F represents the time-interval after the second sonication (named as 'Post2').
Figure 3
Figure 3
The sample time-courses of EEG recordings from the test groups. The representative example of EEG recordings from PTZ-induced epileptic rats (A) with sonication and (B) without sonication. In each EEG dataset, the upper signals represent unfiltered (raw) EEG data, and the lower signals show their corresponding theta-band activity. The insets of magnified windows represent EEG samples for 30 seconds in each highlighted time-window. Red boxes indicate an interval before the first sonication period, blue boxes indicate an interval between the first and the second sonication periods, and green boxes indicate an interval after the second sonication period. Note the changes in raw EEG spikes before, during, and after the sonication in the FUS-treated rat (see the upper red, blue and green boxes in (A)). The ictal activity during the pre-sonication period started to diminish along with each of the two sonication sessions (marked with gray boxes; FUS1-2) and was effectively suppressed after the second sonication (see the upper green box in (A)). Vertical scale bars indicate 100 μV in raw EEG signals and 20 μV in theta activity. Horizontal scale bars indicate a one-minute time scale (ten seconds for the insets).
Figure 4
Figure 4
Analysis of epileptic EEG signal bursts. (A) Comparison of the average number of threshold-exceeding raw EEG peaks (greater than 4.75 standard deviations from the individual baseline activity of the raw EEG data) between the FUS-treated and untreated groups. (B) Comparison of the average number of EEG theta peaks exceeding the absolute magnitude above the threshold (4.75 standard deviations from the individual baseline theta activity) between the same two groups. Red bars indicate Group 1 (the FUS-treated PTZ group: PTZ(+)/FUS(+)), and blue bars indicate Group 2 (the untreated epileptic group: PTZ(+)/FUS(-)). As shown in both graphs, there were no significant differences between the two groups before the sonication. Red brackets indicate statistically significant differences (p < 0.05) within the FUS-treated group, and green brackets indicate statistically significant differences between the two groups. Error bars indicate standard errors of the mean.
Figure 5
Figure 5
Examples of histology from the sonicated brain area. Exemplary histological data obtained from Group 3 (PTZ(-)/FUS(+)). (Left column) H&E staining results and (Right column) TUNEL staining results (DAPI in blue, apoptotic cell in green) from (A) a sonicated thalamic site, (B) the cortex above the sonicated thalamus in the FUS beam path, and (C) an unsonicated posterior cortex. Note the absence of apoptotic DNA-damaged cells in all of the examined locations.

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