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Current Biology 23, 24302433, December 2, 2013 2013 Elsevier Ltd All rights reserved

http://dx.doi.org/10.1016/j.cub.2013.10.029

Report
Low-Intensity Focused Ultrasound
Modulates Monkey Visuomotor Behavior
Thomas Deffieux,1,3 Youliana Younan,1,3 Nicolas Wattiez,2
Mickael Tanter,1 Pierre Pouget,2,4,*
and Jean-Francois Aubry1,4,*
1Institut Langevin Ondes et Images, ESPCI ParisTech,
CNRS UMR 7587, INSERM U979, Paris 75005, France
2Institute of Brain and Spinal Cord, INSERM UMRS 975,
CNRS 7225, UPMC, Paris 75013, France

Summary
In vivo feasibility of using low-intensity focused ultrasound
(FUS) to transiently modulate the function of regional brain
tissue has been recently tested in anesthetized lagomorphs
[1] and rodents [24]. Hypothetically, ultrasonic stimulation
of the brain possesses several advantages [5]: it does not
necessitate surgery or genetic alteration but could ultimately confer spatial resolutions superior to other noninvasive methods. Here, we gauged the ability of noninvasive
FUS to causally modulate high-level cognitive behavior.
Therefore, we examined how FUS might interfere with prefrontal activity in two awake macaque rhesus monkeys that
had been trained to perform an antisaccade (AS) task. We
show that ultrasound significantly modulated AS latencies.
Such effects proved to be dependent on FUS hemifield of
stimulation (relative latency increases most for ipsilateral
AS). These results are interpreted in terms of a modulation
of saccade inhibition to the contralateral visual field due to
the disruption of processing across the frontal eye fields.
Our study demonstrates for the first time the feasibility of
using FUS stimulation to causally modulate behavior in the
awake nonhuman primate brain. This result supports the
use of this approach to study brain function. Neurostimulation with ultrasound could be used for exploratory
and therapeutic purposes noninvasively, with potentially
unprecedented spatial resolution.

in Figure 2C, FUS stimulation applied within a control premotor


cortex did not significantly affect ipsilateral AS latencies (t test:
monkey Y, p > 0.69; monkey L, p > 0.1) or contralateral
latencies (monkey Y, p > 0.11; monkey L, p > 0.74). The effect
of stimulation was transient (no significant effects were
observed on the consecutive control trial following the stimulation, p > 0.5 for both animals). Stimulation in the left FEF while
the animals were performing a prosaccade (PS) task only
marginally affected the latencies of contralateral saccades
for one animal (p < 0.001 and p > 0.52 respectively; Figure 2B).
In both monkeys, sham FUS did not interfere with ipsilateral or
contralateral saccade latency (p > 0.5).
FUS Effect on AS Error Rate
The impact of FUS stimulation over FEF on AS error rate was
statistically tested separately for each condition. Ipsilateral
error rate AS with ultrasound stimulation was not different
from the nonstimulation condition (p > 0.5, across conditions
and animals). Only for monkey Y did the contralateral error
rate AS marginally increase with ultrasound stimulation (FUS,
9% 6 5.7% error; no FUS, 5.1% 6 3.1% error, p = 0.05).
FUS Effect on Saccade Amplitude, Peak Velocity, and
Skewness
The impact of FUS stimulation over FEF on saccade amplitude,
peak velocity, and skewness was statistically tested separately for each condition. The FUS pulses did not affect eye
movement metrics. As compared to sham nonstimulated trials, the amplitude of trials with FUS did not significantly vary
in any of the experimental conditions (p > 0.5 for all conditions
in both animals). Ipsilateral peak velocity AS with ultrasound
stimulation was not different from the nonstimulation condition (p > 0.5 across conditions and animals). Finally, the US
pulses did not affect the skewness of saccades (p > 0.5 for
all conditions in both animals).
Discussion

Results
Effects of Focused Ultrasound over Left FEF 100
Milliseconds after Target Onset
Focused ultrasound (FUS) stimulations were delivered in the
left frontal eye field (FEF) (Figure 1). As shown in Figure 2A,
antisaccade (AS) latencies were affected by FUS. More specifically, across the experimental sessions, ipsilateral mean AS
latencies with ultrasound stimulation were significantly slowed
(monkey Y, p = 0.0018; monkey L, p < 0.001) compared to
the nonstimulated condition (monkey Y, no FUS= 221 ms,
FUS = 235 ms; monkey L, no FUS = 239 ms, FUS = 269 ms).
For the two animals, contralateral mean AS latencies were
not significantly slowed (t test: monkey Y, p > 0.8; monkey L,
p > 0.6) compared to the nonstimulated condition. As shown

3These

authors contributed equally to this work and are co-first authors


authors contributed equally to this work and are co-senior authors
*Correspondence: pierre.pouget@upmc.fr (P.P.), jean-francois.aubry@
espci.fr (J.-F.A.)

4These

This study demonstrates the feasibility of using FUS brain


neurostimulation to causally modulate high-level cognitive
behavior and opens the door for further parametric studies.
Compared with the versatile modulations of response times
reported with transcranial magnetic stimulation (TMS) [6], our
results with FUS appear to be less affected by the baseline
variability of response times, with no discomfort imposed on
the animals during ultrasonic neurostimulation. Optogenetic
manipulation has been achieved recently on two monkeys during PS tasks [7] (as opposed to the AS tasks presented here).
A mean latency reduction of 14 ms (monkey 1) and 20 ms
(monkey 2) was reported for ipsilateral targets in a visually
guided PS task with multiple possible go cues. Our results
appear to be as effective (lengthening ipsilateral mean latency)
and as strong (14 ms and 30 ms for monkeys Y and L, respectively), in spite of the use of very conservative ultrasound
parameters below the limitations outlined by the US Food
and Drug Administration for transcranial ultrasound imaging
safety guidelines. We also observed a marginal increase in
the contralateral error rate and a tendency for the ipsilateral

Ultrasound Modulates Monkey Visuomotor Behavior


2431

Baseline

Mixed 400

Post

100 Trials

(360 Trials + 40 Trials with ultrasound )

100 Trials

a block of PSs might not recruit FEF as much as ASs. In a more


reflexive oculomotor task, the loop including the lateral intraparietal, the superior colliculus and the brain stem is preferentially recruited. Second, the timing of the FUS (100 ms long
pulse, 100 ms after target onset) might have been too late to
disturb the fast PS motor preparation.
Although FUS underlying mechanisms are not yet fully
understood, ultrasonic neurostimulation is believed to rely on
the opening of voltage-gated sodium channels due to the
ultrasonic pressure wave, as observed on rodent hippocampal
slices [8]. Although the stimulation spatial resolution was out of
the scope of this report, the 320 kHz transducer used here concentrates ultrasound in a 5 3 5 3 33 mm cigar-shaped focal
spot (at w3 dB size, cf. Figure 3), 63 mm away from the surface
of the transducer. Since frequencies up to 1 MHz (1.5 3 1.5 3
5 mm focal spot) can be focused transcranially anywhere deep
in the human brain using MR-guided multielement arrays [9],
ultrasonic neurostimulation is regarded as a potentially deep
and millimetric noninvasive neurostimulation tool. For comparison, the best lateral resolution for TMS was evaluated to
be around 1 cm using a small and thus highly superficial coil
in cats [10].
The potential of modulating small and deep brain structures with this noninvasive and spatially specific tool opens
up exciting possibilities in brain circuitry exploration in
primates and in the treatment of neurological disorders
e.g., Parkinsons disease, essential tremor, and disorders of
consciousness.

Anti saccade task + ( Ultrasound )


Fixation
0 ms

Target

Antis
acca
de la
tenc
y (m
s)

500 -1000 ms

( Ultrasound )
Target +100 ms

Anti Saccade

Figure 1. Experimental Session


(A) One session consists of three blocks with left and right (50%) antisaccade (AS) trials (100 trials for the baseline, 400 trials including 40 trials
with ultrasound during the central block, and 100 trials as a post block).
(B) The AS paradigm. Monkeys were required to maintain eye fixation on a
central stimulus. After initial fixation onset, a red square (target) appeared
on the right or left. Monkeys were trained to initiate a saccade toward the
opposite direction. During ultrasound trials, 320 kHz ultrasound was applied
for 100 ms, 100 ms after the target appearance.

latencies of the PSs. Altogether, we have interpreted these results in terms of a modulation of saccade inhibition to the
contralateral visual field due to disruption of processing
across the FEFs. At least two main factors may explain why
in our experiments only marginal effects of FUS over the FEF
were observed in the PS task. First, in highly trained monkeys,

ipsilateral
NoStim
Stim

300

Focused Ultrasound
A single-element ultrasound transducer (H115, diameter 64 mm, Sonic
Concept), geometrically focused to 63 mm, was used with a coupling
cone (C103, Sonic Concepts) filled with degassed water. The frequency
was set to 320 kHz and the pulse duration to 100 ms using a first function
generator, with rise and fall times set to 1 ms using a second generator
(AFG3101, Tektronix) connected to the amplitude modulation entry of the
first one. A 75 W amplifier (75A250A, Amplifier Research) was used to deliver
the required power to the transducer, and the input voltage of the transducer was monitored with an oscilloscope (TDS2022B, Tektronix). The pressure amplitude at focus was set to 0.6 MPa, as measured in free water with a
heterodyne interferometer.
Skull transmission was estimated on a clean and degassed primate skull
specimen (Macaca mulatta skull) at seven different locations and was found
to be 58% 6 8%. This allowed us to estimate the derated pressure at
Figure 2. Results

Prosaccades / FEF region


350

contralateral
NoStim
Stim

***

Latency (ms)

350

NS

NS

300

NS
250
200

***

NS
150
100

**

monkey Y

250

C
NS

monkey L

Antisaccades / Premotor cortex


350

Latency (ms)

Latency (ms)

Antisaccades / FEF region

Experimental Procedures

200

NS
NS

300
250

NS

NS

200
150

150

monkey Y

monkey L

monkey Y

monkey L

(A) Ipsilateral and contralateral mean AS latencies


with and without ultrasound on both monkeys.
Ipsilateral AS latencies are significantly slowed
(p < 0.05) with ultrasound.
(B) Same convention, for prosaccade (PS) tasks.
(C) AS latencies when the transducer is positioned over the premotor cortex. No significant
changes in latencies were observed.
Error bars represent SEM over all trials.

Current Biology Vol 23 No 23


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10 mm

5 mm
AS

PS
x2

Monkey
head
Ultrasound
transducer

Coupling cone

-3dB focal spot

4.7 mm
33 mm

Supporting
arm

x4

pulses were delivered to the left FEF at 100 ms stimulus onset asynchrony
interval (SOA), after the visual target appearance, which was selected based
on preliminary monkey saccade latency measurements.
In each experiment session, animals performed a total of three blocks of
AS training per session. First, monkeys performed a 100-trial block of AS (50
for each side) as a baseline. A second block of 400 trials was then performed: 360 trials without FUS (180 for each side) and 40 trials with FUS
(20 for each side). Trials with FUS were pseudorandomly interleaved with
trials without FUS. A final block of 100 trials was performed as a posttest.
Monkey Y performed ten sessions and monkey L twelve sessions.
Control sessions were performed using identical procedures, with the
transducer positioned over the premotor cortex 1012 mm away (Figure 3)
from left FEF (monkey Y, eight sessions; monkey L, seven sessions). PS
modulation was also investigated using identical procedures but with the
PS paradigm (monkey Y, ten sessions; monkey L, seven sessions). Finally,
in additional sham FUS sessions directly inspired by TMS sham experiments, the ultrasound transducer was moved 4 cm away from the animals
head so that ultrasound could not reach the target (monkey Y, five sessions;
monkey L, two sessions).
Data Analysis and Presentation
Trials with blinking responses interfering with eye recordings were eliminated from the data set. Saccades were detected using homemade
MATLAB scripts that searched first for significantly elevated velocity
(>30 /s) and then for monotonic change in eye position lasting 12 ms before
and after the high-velocity gaze shift. Individual latency values were averaged for trials under FUS and compared to those without stimulation for
each session and experimental condition explored in the study on each
monkey. All statistical comparisons were based on Students t test.

Figure 3. Transducer Positioning


The ultrasonic transducer is manually positioned so that the cigar-shaped
focal spot targets the left frontal eye field or region of premotor cortex
(red and blue spot, respectively, with an estimated distance between 10
and maximum 12 mm). A coupling cone filled with water ensures the ultrasonic coupling between the transducer and the animals head.

0.35 MPa in the brain of the monkeys. The corresponding mechanical index
(MI) is 0.6 6 0.1 W/cm2 and intensity spatial peak pulse average (ISPPA) is
4 6 1.1 W/cm2 behind the skull. By taking into account a minimum 30 s
pause between each ultrasonic pulse, we also estimated the corresponding
spatial peak time average intensity (ISPTA) at less than 13.5 6 3.8 mW/cm2.
All of those values are far below the US Food and Drug Administration
limitations for ultrasound imaging safety guidelines (MI = 1.9, ISPPA =
190 W/cm2, ISPTA = 720 mW/cm2), ensuring that the parameters used
here are safe.
Tasks
Prior to the first experimental session, animals were trained in the AS and PS
paradigms. Between 500 and 1,000 ms after initial fixation onset on a brown
central fixation stimulus (green for PS) and simultaneously with its disappearance (no gap), a red square appeared for 1,000 ms at a 16 right or
left location. For AS, monkeys were trained not to look at this peripheral
target but instead to initiate a saccade in the opposite direction (Figure 1).
Conversely, for PSs, they had to look at the target as soon as possible.
After the saccade, the monkey received a reward if the saccade fell within
a 5 3 5 window centered on the correct location. Failure to trigger a
saccade within 2,000 ms after target onset cancelled the trial. The main
focus was placed on ASs, since prior human and monkey TMS experiments
have revealed PS paradigms to be much less sensitive to single-pulse TMS
interference than ASs [11].
Eye movements were recorded with an infrared eye tracker at 1,000 Hz
(EyeLink 1000, SR Research) and a real-time data acquisition system
(Rexeno) (see [6, 12]).
Experimental Protocols
Two macaques (Macaca mulatta Y and L) participated in this study. The
monkeys were paired housed and handled in strict accordance with the
recommendations of the Weatherall Report about good animal practice
(last renewals DTPP 2010-424). The surgical procedures for titanium headpost implant were the same as described previously [6, 11].
The left FEF field was identified according to stereotaxic coordinates for
this location, and its site was labeled with a color tattoo on the monkeys
skin, which lasted for several weeks and was renewed when faded. FUS

Acknowledgments
This work was supported by LABEX WIFI (Laboratory of Excellence within
the French Program Investments for the Future) under references ANR10-LABX-24 and ANR-10-IDEX-0001-02 PSL and by the Agence Nationale
de la Recherche under the program Future Investments with the reference
ANR-10-EQPX-15.
Received: May 29, 2013
Revised: September 18, 2013
Accepted: October 10, 2013
Published: November 14, 2013
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