Transcranial Ultrasound Stimulation Attenuates Repetitive Behavior and Improves Sociability and Communication in a BTBR Mouse Model of Autism
Rasha Noureddine, Monica Roman, Reese Hassall, Ruiqi Wang, Kartikeya Murari, Samuel Pichardo, G. Bruce Pike
2026 · 10.2139/ssrn.7110032
Abstract
Background: Autism Spectrum Disorder (ASD) is a complex neurodevelopmental condition with increasing prevalence and no approved therapies targeting its core social and behavioral deficits. Transcranial ultrasound stimulation (TUS) has emerged as a promising noninvasive, spatially precise, and deep-penetrating neuromodulatory approach for various neurological and neuropsychiatric disorders, yet its potential in ASD remains largely unexplored. <div> <br> </div> <div> Methods: This study investigated whether TUS could modulate ASD-related phenotypes in the Black and Tan BRachyury (BTBR) mouse model of autism (BTBR: n = 33; B6: n = 23). TUS was administered to the medial prefrontal cortex for five consecutive days, followed by behavioral assessments. </div> <div> <br> </div> <div> Findings: TUS successfully reduced repetitive behaviors (p < 0·001) and improved measures of sociability, specifically: a reversal of abnormal non-social preference (pre-stim p < 0·05; post-stim p > 0·05); communication with increased vocalization frequency jumps (p < 0·05), step down calls (p < 0·05) and decreased flat calls (p < 0·05). These behavioral effects were accompanied by increased cFOS-positive cell density in the targeted brain region, indicating region-specific neuronal activation, with no detectable histological damage. </div> <div> <br> </div> <div> Interpretation: These findings provide the first preclinical evidence that TUS can beneficially modulate ASD-like behavioral phenotypes in a validated mouse model through targeted engagement of a disease-relevant neural circuit. This establishes a framework for future mechanistic and translational investigations, highlighting the therapeutic potential of TUS for ASD. </div>
Abstract via crossref.
Exposures
Exposure 1: TUS to the medial prefrontal cortex (mPFC)
Target: ventromedial prefrontal cortex — “medial prefrontal cortex (mPFC), comprising the infralimbic cortex, prelimbic cortex, and cingulate cortex”
Device: other named manufacturer · FUS Instruments · RK-50 ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 476.5 | ✓✓✓ |
| Pulse duration (ms) | 0.25 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 1,500 | ✓✓✓ |
| Duty cycle (%) | 37.5pulse duration × PRF gives 37.5% | ✓✓✓ |
| Sonication duration (s) | 0.45 | ✓✓✓ |
| 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 | |
| Pressure, domain unspecified (kPa) | 640 | ✓✓✓⚑ |
| Isppa, domain unspecified (W/cm²) | 13.3 | ✓✓✓ |
| Ispta, domain unspecified (W/cm²) | 5 | ✓✓✓ |
The transducer was driven with a pulse repetition frequency (PRF) of 1.5 kHz, a pulse length (PL) of 0.25 ms, a duty cycle of 37.5%, a burst duration (BD) of 0.45 s, and an inter-stimulus interval (ISI) of 2.1 s; TUS pulses were smoothed with 30% ramping to minimize unintended auditory brainstem response. The end of the ultrasonic beam (-6 dB cutout) was used to stimulate the mPFC at two stereotactic coordinates (0.64 mm and 1.78 mm anterior to Bregma, at depths of 2.5 mm and 3.25 mm respectively) to encompass the entire mPFC. TUS was applied for a total time of 15 minutes daily over five consecutive days.
Flags from extraction
n_subjects— Paper gives TUS-treated group sizes across two separate cohorts (pilot: 3 BTBR stimulated; main cohort: 12 B6 stimulated, 18 BTBR stimulated) but never states a combined total of TUS-exposed animals; reported as a list rather than summed.sham_type— Paper states sham mice underwent identical preparatory procedures but TUS/Bregma registration was applied only to stimulated mice; it does not describe whether a transducer was present but inactive for sham mice, so mechanism is undescribed.exposures[0].unspecified_domain.pressure_kpa— Paper does not state whether the reported Ispta/Isppa/pressure values at the -6dB beam cutout are free-field (water) or in-situ (brain) measurements; recorded as unspecified domain.direction_of_effect— Effects were heterogeneous across behavioral domains (reduced grooming, increased locomotion, no anxiety effect, selective USV changes); classified as mixed_or_unclear.