Fig 1.
Schematic of the experimental setup.
(A) An exemplar 3D rendering and triplanar view of the processed fMRI results (p < 0.05, z-score > 1.64; z-score map in pseudo color) overlaid on the 3D anatomical neuroimaging data that describes locations of the M1 and S1 as well as (B) the VL/VPL, marked by red arrows. The activation loci for the M1/S1 were observed anterior and posterior to the central sulcus (dotted white lines on the sagittal and axial views) whereas the VL/VPL areas did not show distinctive boundaries. (C) Acoustic intensity profiles in the longitudinal and transverse planes. The sonication direction is represented by the short white arrow. The dotted white circle and ellipse indicate the region of 90%-maximum of the intensity profile. (D) Apparatus of the image-guided transcranial FUS setup. (E) Illustration of acoustic parameters: acoustic intensity (Ia), tone-burst duration (TBD), pulse repetition frequency (PRF), duty cycle (DC), sonication duration (SD), and inter-stimulation interval (ISI). ISI is not applicable for suppressive sonication as a single, 2 min-long sonication was given per session. Scale bar = 10 mm in (A–C, C inset).
Table 1.
Combinations of sonication parameters used for examining the excitatory effects.
DC = duty cycle, TBD = tone-burst duration, PRF = pulse repetition frequency, SD = sonication duration, Ia = acoustic intensity, Isppa = spatial-peak pulse-average intensity, Ispta = spatial-peak temporal-average intensity, Ispta = Isppa × DC, MI = mechanical index.
Table 2.
Combinations of sonication parameters used for examining the suppressive effects.
Fig 2.
Results of excitatory sonication to the M1 and thalamus (Thal).
The group-averaged EMG response rates (with standard error bars) measured from bilateral hind limbs for sonication parameters EP1–EP30 (see Table 1), evaluated across ten sheep (8800 sonication trials for the M1 stimulation and 4800 trials for the thalamic stimulation). The box indicates the range of positive and negative standard error, and the black horizontal line in the box indicates the average value.
Fig 3.
The response rate in excitatory sonication to the M1 and thalamus, comparing different DCs, tone-burst durations (TBDs), and Isppa.
Averaged response rate (horizontal black line in box) with standard error (box) in contralateral EMG was plotted across the different sonication parameters in terms of (A) DC, (B) TBD, and (C) Isppa. The bracket with the asterisk indicates significant differences (one-way ANOVA with LSD post-hoc analysis in panels A and B, post-hoc test p < 0.05, and paired two-tailed t-test in panel C, p < 0.05) of the response rate between different sonication parameters.
Fig 4.
Histogram of onset latencies of contralateral hind limb EMG signals after FUS onset.
Blue bars indicate the responses from M1 stimulation sessions while green bars indicate those from thalamic stimulation sessions.
Fig 5.
Time-locked contralateral EMG measured from M1/thalamic stimulation.
The EMG signals were averaged, using the results obtained from the use of 70% DC (i.e., EP17–EP24) having an onset latency between 50 and 75 ms. (A) The averaged data from stimulation of the M1 and thalamus is displayed in the blue and green lines, respectively. The data obtained in the absence of sonication is plotted in the black line (labeled as ‘No FUS’). The baseline signal drift/offset was removed from all individual EMG data with respect to FUS onset. The colored bars indicate regions of significant differences (p < 0.01, one-tailed t-test) in the amplitude obtained from M1 (in blue) and thalamic (in green) stimulation compared to the amplitude obtained when FUS was not given (i.e., ‘No FUS’). Dashed lines indicate the onset timing of FUS sonication, and thick solid black bars represent the duration of sonication. (B–D) Each of the averaged signal traces shown in (A) is separately shown for (B) No FUS, (C) M1 and (D) thalamic stimulation, respectively. The shaded envelopes indicate the standard deviation of the averaged baseline/EMG signal traces. The zero padded temporal features of the signal were due to the application of low-pass digital filter (30 Hz).
Fig 6.
Normalized SEP magnitude (P50 –N40; averaged across the animals shown with standard error) before (B1–B3), during (F1, F2), and after (P1–P5) sonication of the contralateral S1 (left column) and thalamus (right column).
The top shows the data acquired from the control condition (i.e., sonication of the ipsilateral S1 and thalamus), with each row representing the data acquired with each sonication parameter set (SP1–SP8; refer to Table 2 for detailed parameters). The dashed line indicates the normalized signal level averaged across baseline B1–B3 segments (set to zero). The asterisks indicate statistically significant differences (one-tailed t-test, p < 0.01) in SEP magnitude when compared to the control condition.
Fig 7.
Group-averaged SEP (acquired from the use of SP1 and SP3 parameter conditions) across data acquisition segments for the sonication targets (S1, thalamus).
The time-locked averaged SEPs are shown before (B1–B3), during (F1, F2), and after (P1–P5) sonication of the S1 (left column, n = 17 across 9 sheep) and thalamus (middle column, n = 16 across 8 sheep). The SEPs acquired from the control condition (i.e., sonication of the ipsilateral S1/thalamic targets, right column, n = 17 across 5 sheep) are also shown. Gray bars in the plot indicate the time intervals that showed significant differences in amplitude compared to the control condition (one-tailed t-test, p < 0.01).
Fig 8.
Estimated temperature rise at the focus.
(A) 20 sonication repetitions with 5 s ISI using excitatory parameter set of EP30. (B) Sonication duration of 2 min using suppressive parameter set of SP6. The trace of thermal change was illustrated with the raw data of numerical simulation performed with a temporal resolution of 0.2 ms.
Fig 9.
Exemplar histology results of sheep brain tissue (‘SH4’).
The microscopic images (×40 magnification) of the respective tissue sampling location (M1/S1, Thal, and control site) are displayed according to the staining method (H&E, VAF-toluidine blue, caspase-3, and GFAP). Insets represent magnified images (×100 magnification).