Optimized ultrasound neuromodulation for non-invasive control of behavior and physiology
Keith R. Murphy, Jordan S. Farrell, Jonas Bendig, Anish Mitra, Charlotte Luff, Ina A. Stelzer, Hiroshi Yamaguchi, Christopher C. Angelakos, Mihyun Choi, Wenjie Bian, Tommaso DiIanni, Esther Martinez Pujol, Noa Matosevich, Raag Airan, Brice Gaudillière, Elisa E. Konofagou, Kim Butts-Pauly, Ivan Soltesz, Luis de Lecea
Neuron 2024, 112, 3252-3266.e5 · 10.1016/j.neuron.2024.07.002
Abstract
Focused ultrasound can non-invasively modulate neural activity, but whether effective stimulation parameters generalize across brain regions and cell types remains unknown. We used focused ultrasound coupled with fiber photometry to identify optimal neuromodulation parameters for four different arousal centers of the brain in an effort to yield overt changes in behavior. Applying coordinate descent, we found that optimal parameters for excitation or inhibition are highly distinct, the effects of which are generally conserved across brain regions and cell types. Optimized stimulations induced clear, target-specific behavioral effects, whereas non-optimized protocols of equivalent energy resulted in substantially less or no change in behavior. These outcomes were independent of auditory confounds and, contrary to expectation, accompanied by a cyclooxygenase-dependent and prolonged reduction in local blood flow and temperature with brain-region-specific scaling. These findings demonstrate that carefully tuned and targeted ultrasound can exhibit powerful effects on complex behavior and physiology.
Abstract via europepmc.
Exposures
Exposure 1: CMT (CAMKII+) PRF / temporal-compression / intensity coordinate-descent optimization
Target: thalamus — “central medial thalamus (CMT)”
Device: custom-built · PhoCUS mountable ring transducer
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 550 | ✓✓✓ |
| Pulse duration (ms) | not reported | ⚑ |
| Pulse repetition frequency (Hz) | 2.5, 40swept | ✓✓✓ |
| Duty cycle (%) | 20, 2.5swept | ✓✓✓⚑ |
| Sonication duration (s) | 5, 40swept | ✓✓✓⚑ |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not reported | |
| In-situ pressure (kPa) | not reported | |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported | |
| Isppa, domain unspecified (W/cm²) | 0.3, 7.4swept | ✓✓✓⚑ |
Ultrasound was pulsed with 5 different PRFs ranging from 2.5 to 40 Hz while maintaining a fixed pulse train duty cycle of 20%, an intensity of 3.7 W/cm2, and a pulse train of 5 s, with largest excitation at 2.5 Hz and significant post-stimulus inhibition at 20 Hz. With a PRF of 2.5 Hz selected for excitation and 20 Hz for inhibition, the duration of pulsing was incrementally increased while duty cycle decreased so total power delivery was held constant (maximal excitation at 5 s, 20% duty cycle; greatest inhibition at 40 s, 2.5% duty cycle). Spatial-peak intensity was then incrementally increased from 0.3 to 7.4 W/cm2 (peak excitation at 5.4 W/cm2; significant post-stimulus inhibition only at 3.7 W/cm2). Each parameter combination was trialed 7 times in randomized order for each animal with a 3-min interval between the beginning of each trial.
Exposure 2: DMH (DLX+/GABAergic) PRF / compression optimization and suboptimal-protocol comparison
Target: hypothalamus — “dorsomedial hypothalamus (DMH)”
Device: custom-built · PhoCUS mountable ring transducer
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 550 | ✓✓✓ |
| Pulse duration (ms) | not reported | |
| Pulse repetition frequency (Hz) | 2.5, 40swept | ✓✓✓ |
| Duty cycle (%) | 2.5, 20swept | ✓✓✓ |
| Sonication duration (s) | 5, 40swept | ✓✓✓ |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not reported | |
| In-situ pressure (kPa) | not reported | |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported |
In examining compression, the DMH responded maximally at 10-s, 10% D.C. compression. To examine whether the optimizations had bearing on behavior, the DMH was also stimulated with the same intensity and duty cycle but with a suboptimal PRF of 20 Hz, which produced less activation and eliminated significant walking induction. Blood volume was also measured following optimized stimulation (2.5 Hz, 10 s, 10% D.C.) of the DMH, LC, and BNST.
Exposure 3: LC (TH+/noradrenergic) compression optimization
Target: locus coeruleus — “locus coeruleus (LC)”
Device: custom-built · PhoCUS mountable ring transducer
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 550 | ✓✓✓ |
| Pulse duration (ms) | not reported | |
| Pulse repetition frequency (Hz) | 2.5, 40swept | ✓✓✓ |
| Duty cycle (%) | 2.5, 20swept | ✓✓✓⚑ |
| Sonication duration (s) | 5, 40swept | ✓✓✓ |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not reported | |
| In-situ pressure (kPa) | not reported | |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported |
In examining compression, LC neurons showed greatest excitation at 5-s 20% duty cycle (D.C.), matching the optimal CMT waveform. For the separate blood-volume assay, LC was instead stimulated with the 'optimized' protocol shared across regions (2.5 Hz, 10 s, 10% D.C.) used for DMH, LC and BNST.
Exposure 4: BNST (GABAergic) PRF / compression examination
Target: other — “bed nucleus of the stria terminalis (BNST)”
Device: custom-built · PhoCUS mountable ring transducer
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 550 | ✓✓✓ |
| Pulse duration (ms) | not reported | |
| Pulse repetition frequency (Hz) | 2.5, 40swept | ✓✓✓ |
| Duty cycle (%) | 2.5, 20swept | ✓✓✓ |
| Sonication duration (s) | 5, 40swept | ✓✓✓ |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not reported | |
| In-situ pressure (kPa) | not reported | |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported |
BNST neurons did not show activation at any PRF but did undergo a significant decrease during the post-stimulus period at 20 Hz stimulation, similar to the CMT. BNST neurons did not show significant suppression at any compression of the 20 Hz protocol and were not passed onto the intensity step of examination. Blood volume was measured following optimized stimulation (2.5 Hz, 10 s, 10% D.C.) of the DMH, LC, and BNST.
Exposure 5: LH (hypocretin) stimulation for walking/SAP comparison with DMH
Target: lateral hypothalamus — “lateral hypothalamus (LH)”
Device: custom-built · PhoCUS mountable ring transducer
| Waveform | not reported | |
|---|---|---|
| Fundamental frequency (kHz) | 550 | ✓✓✓ |
| Pulse duration (ms) | not reported | |
| Pulse repetition frequency (Hz) | not reported | |
| Duty cycle (%) | not reported | |
| Sonication duration (s) | not reported |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not reported | |
| In-situ pressure (kPa) | not reported | |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported |
The focus was positioned onto the LH while recording hypocretin neuron activity and behavior in Hcrt-cre animals; LH stimulation increased walking only during FUS stimulation and produced extended stretch-attend posture, in contrast to DMH stimulation. No PRF, duty cycle, duration or intensity values specific to LH stimulation are given in the main text.
Exposure 6: CMT targeting during ultrafast power-Doppler cerebral blood volume imaging
Target: thalamus — “central medial thalamus (CMT)”
Device: Sonic Concepts · Sonic Concepts · H204 ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 500 | ✓✓✓ |
| Pulse duration (ms) | 80 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 2.5 | ✓✓✓ |
| Duty cycle (%) | 20pulse duration × PRF gives 20% | ✓✓✓ |
| Sonication duration (s) | 5 | ✓✓✓ |
| Free-field pressure (kPa) | 767 | ✓✓✓ |
|---|---|---|
| Free-field Isppa (W/cm²) | 18.89 | ✓✓✓ |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not reported | |
| In-situ pressure (kPa) | not reported | |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported |
Experiments (n = 5 mice) consisted of 4 trials with a 25 s baseline, 5 s sonication and a 155 s cooldown period. FUS (pressure = 0.767 MPa, intensity = 18.89 W/cm2, pulse duration = 80 ms, duty cycle = 20%) was delivered to the CMT at 3 mm posterior to the center of the natural focus, confocally imaged with power-Doppler; the same stimulus is described elsewhere as 2.5 Hz, 5 s, 20% D.C., 18.9 W/cm2.
Flags from extraction
exposures[0].timing.pulse_duration_ms— Paper gives PRF and duty cycle for the main PhoCUS experiments but never states pulse duration numerically; burst rule cannot be resolved from the main text alone.exposures[0].timing.duty_cycle_pct— List combines the 20% value used for the 2.5 Hz excitatory optimum and the 2.5% value used for the 20 Hz inhibitory optimum; these are not simultaneous settings.exposures[0].timing.sonication_duration_s— List combines the 5 s excitatory optimum and 40 s inhibitory optimum durations, which apply to different (2.5 Hz vs 20 Hz) protocols, not a single simultaneous sweep endpoint.exposures[0].unspecified_domain.isppa_w_cm2— Paper does not state whether the 0.3-7.4 W/cm2 intensity range is a free-field or in-situ value; domain is unspecified.exposures[2].timing.duty_cycle_pct— LC's own compression optimum (5 s, 20% D.C.) differs from the shared 'optimized' protocol (2.5 Hz, 10 s, 10% D.C.) later applied to LC in the blood-volume assay; both values are listed.exposures[4].timing.waveform— No numeric stimulation parameters (PRF, duty cycle, duration, intensity) for LH stimulation are stated in the main text; only that the DMH paradigm/apparatus was repositioned to the LH.n_subjects— The paper reports many overlapping, experiment-specific group sizes (e.g., n = 9, 7, 8, 6, 7, 5, 5-6, 3-5 mice across different assays and targets) but never a total number of animals used in the study.n_sessions_per_subject— Parameter combinations were each trialed 7 times per animal with a 3-min inter-trial interval, but the paper does not clearly state whether this constitutes one session or multiple sessions per animal.exposures[0].target.terms— The central medial thalamus (CMT) does not map exactly to any listed thalamic nucleus; classified at the parent 'thalamus' level.exposures[1].target.terms— The dorsomedial hypothalamus (DMH) is not among the listed hypothalamic subnuclei; classified at the parent 'hypothalamus' level.exposures[3].target.terms— The bed nucleus of the stria terminalis (BNST) has no matching target id; classified as 'other'.