High-throughput ultrasound neuromodulation in awake and freely behaving rats
Tommaso Di Ianni, Kyle P. Morrison, Brenda Yu, Keith R. Murphy, Luis de Lecea, Raag D. Airan
Brain Stimulation 2023, 16, 1743-1752 · 10.1016/j.brs.2023.11.014
Abstract
Transcranial ultrasound neuromodulation is a promising potential therapeutic tool for the noninvasive treatment of neuropsychiatric disorders. However, the expansive parameter space and difficulties in controlling for peripheral auditory effects make it challenging to identify ultrasound sequences and brain targets that may provide therapeutic efficacy. Careful preclinical investigations in clinically relevant behavioral models are critically needed to identify suitable brain targets and acoustic parameters. However, there is a lack of ultrasound devices allowing for multi-target experimental investigations in awake and unrestrained rodents. We developed a miniaturized 64-element ultrasound array that enables neurointerventional investigations with within-trial active control targets in freely behaving rats. We first characterized the acoustic field with measurements in free water and with transcranial propagation. We then confirmed in vivo that the array can target multiple brain regions via electronic steering, and verified that wearing the device does not cause significant impairments to animal motility. Finally, we demonstrated the performance of our system in a high-throughput neuromodulation experiment, where we found that ultrasound stimulation of the rat central medial thalamus, but not an active control target, promotes arousal and increases locomotor activity.
Abstract via europepmc.
Exposures
Exposure 1: Blood-brain-barrier-opening validation sonication of the striatum
Target: striatum — “striatum”
Device: Sonic Concepts · Sonic Concepts, Inc. · custom 64-element hemispherical wearable array (Verasonics-driven) ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 1,000 | ✓✓✓ |
| Pulse duration (ms) | not reportedimplied by duty cycle ÷ PRF: 50 ms (not stated by the paper) | |
| Pulse repetition frequency (Hz) | 1 | ✓✓✓ |
| Duty cycle (%) | 5 | ✓✓✓ |
| Sonication duration (s) | 120 | ✓✓✓ |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | deratingsingle value | |
| In-situ pressure (kPa) | 600 | ✓✓✓ |
| In-situ Isppa (W/cm²) | not reported | |
| In-situ Ispta (W/cm²) | not reported |
In one male rat, ultrasound bursts were applied for 2 min with an estimated in-situ pressure of 0.6 MPa, 1-Hz pulse repetition frequency, and 5% duty cycle, targeted at the striatum, following intravenous bolus injection of microbubble contrast agent (DEFINITY); the acoustic pressure at the target was corrected to compensate for skull insertion losses. Post-contrast T1-weighted MRI confirmed blood-brain barrier permeabilization at the target.
Exposure 2: Ultrasound neuromodulation of central medial thalamus (CMT) with within-trial active control target
Target: thalamus — “central medial thalamus (CMT)”
Device: Sonic Concepts · Sonic Concepts, Inc. · custom 64-element hemispherical wearable array (Verasonics-driven) ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 1,000 | ✓✓✓ |
| Pulse duration (ms) | 80 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 2.08 | ✓✓✓⚑ |
| Duty cycle (%) | 17pulse duration × PRF gives 16.64% | ✓✓✓ |
| Sonication duration (s) | 5 | ✓✓✓ |
| Free-field pressure (kPa) | 240, 1,830swept | ✓? |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | simulation | |
| In-situ pressure (kPa) | 130, 970swept | ✓? |
| In-situ Isppa (W/cm²) | 0.52, 30.22swept | ✓? |
| In-situ Ispta (W/cm²) | 0.04, 2.43swept | ✓?⚑ |
Ultrasound stimuli of either 1 or 3 bursts were delivered to the CMT or a symmetric active control target (ventral end of dorsal peduncular cortex), with target and TPC excitation voltage (2-12 V in 2 V steps) randomized within each session. Each burst had a duration of 5 s made of 80-ms sinusoidal pulses with a 480-ms pulse repetition period (duty cycle 17%); an inter-burst interval of 10 s was used for the 3-burst condition. One stimulus was delivered every 2 min while locomotor activity was recorded continuously.
Flags from extraction
n_subjects— Different sub-experiments used different numbers of animals (BBB-opening validation: 1 rat; CMT neuromodulation locomotor experiment: 6 rats for 1-burst stimulation, of which 3 also received 3-burst stimulation); the paper never states a single total N for the study, so the two distinct group sizes are listed.n_sessions_per_subject— Stated only as a maximum ('some rats underwent up to 3 sessions'), not as an exact number of sessions per subject.exposures[1].in_situ.ispta_w_cm2— Table 1 lists ISPTA values of 0.04-2.43 W/cm2, but the main text states the corresponding range as 0.02-2.42 W/cm2 for the same simulated CMT stimuli; the table values are used here and the discrepancy is flagged.exposures[1].in_situ.reported_as— The pressure/intensity range at CMT is a sweep across TPC drive voltages (stimulus intensities), not a range or mean across subjects; 'reported_as' vocabulary options do not cleanly describe this case.exposures[1].timing.pulse_repetition_frequency_hz— PRF was not stated directly; converted from the stated pulse repetition period (480 ms) to ~2.08 Hz, a unit conversion rather than an inference.exposures[1].target— The paper's target 'central medial thalamus (CMT)' does not exactly match any specific thalamic nucleus in the target vocabulary (closest is centromedian_nucleus, a distinct intralaminar nucleus); mapped to the parent term 'thalamus' rather than force an inexact specific match.anaesthesia— Isoflurane anesthesia was used only briefly (<2 min) for headgear attachment before testing; ultrasound stimulation and behavioral testing were performed in awake, freely-behaving rats, consistent with the paper's title.