Transcranial Ultrasound Stimulation Pulsed at 40 Hz Improves Cognition and Neuroinflammation in Female Mice with Alzheimer’s Disease
Shasha Yi, Junjie Zou, Xiaofei He, Houminji Chen, Zhengrong Lin, Liying Zhang, Yiyue Zhu, Zejie Zuo, Zehao Chen, Xiquan Hu, Lili Niu
Research 2026 · 10.34133/research.1244
Abstract
Recent advances in transcranial ultrasound stimulation (TUS) pulsed at 40 Hz have demonstrated the potential to ameliorate cognitive deficits in mouse models of Alzheimer's disease. However, technical barriers remain as general anesthesia is required for mice, which restricts the accurate elucidation of biological mechanisms and behavioral effects under awake physiological conditions. Here, we report a wearable, free-moving ultrasound stimulation system that delivers TUS pulsed at 40 Hz to female 5xFAD transgenic mice to systematically evaluate the behavioral outcomes and underlying mechanistic pathways. Among the treatment groups, a 14-d regimen at an acoustic intensity of 2.14 W/cm 2 yielded the optimal cognitive outcome in Alzheimer's disease mice, which was consistently verified across Y-maze and Morris water maze tests. Additionally, this group showed reduced Aβ plaque deposition and increased plaque-associated microglial activity. Furthermore, enhanced gamma oscillations in the hippocampus were detected following treatment. RNA sequencing revealed modulation of innate immune and inflammatory pathways. Corresponding molecular analysis demonstrated a marked down-regulation in RIPK1 , phosphorylated NF-κB , and necroptosis markers, alongside reductions in key pro-inflammatory cytokines ( IL-6 , IL-1β , and TNF-α ). Collectively, our findings suggest that the cognitive improvement observed after treatment with TUS pulsed at 40 Hz may be linked to the modulation of neuroinflammatory and necroptotic pathways, possibly involving RIPK1 / NF-κB signaling.
Abstract via europepmc.
Exposures
Exposure 1: TUS to bilateral hippocampus and cortex (TUS1-TUS4 groups)
Target: hippocampus, cerebral cortex — “bilateral hippocampal and cortical regions”
Device: custom-built · head-mounted planar PZT transducer
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 500 | ✓✓✓ |
| Pulse duration (ms) | 1.25 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 40 | ✓✓✓ |
| Duty cycle (%) | not reportedpulse duration × PRF gives 5% | ⚑ |
| Sonication duration (s) | 1 | ✓✓✓ |
| 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) | 42.6 | ✓✓✓⚑ |
| Ispta, domain unspecified (W/cm²) | 0.33, 2.14swept | ✓✓✓ |
0.5 MHz, PRF 40 Hz, 1.25 ms tone burst, 1 s sonication, 10 s interstimulus interval; 20 min twice daily for 7 or 14 d.
Exposure 2: BV2 microglial cells, ultrasound from beneath culture plate
Target: cultured glia — “BV2 cells”
Device: custom-built · head-mounted planar PZT transducer
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 500 | ✓✓✓⚑ |
| Pulse duration (ms) | 1.25 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 40 | ✓✓✓ |
| Duty cycle (%) | not reportedpulse duration × PRF gives 5% | |
| Sonication duration (s) | 1 | ✓✓✓ |
| Free-field pressure (kPa) | not reported | ⚑ |
|---|---|---|
| Free-field Isppa (W/cm²) | not reported | |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | not applicable | |
| In-situ pressure (kPa) | not applicable | |
| In-situ Isppa (W/cm²) | not applicable | |
| In-situ Ispta (W/cm²) | not applicable | |
| Ispta, domain unspecified (W/cm²) | 0.33 | ✓✓✓ |
In vitro: ultrasound applied from beneath the culture plate for 10 min; the remaining parameters same as in vivo (0.5 MHz, PRF 40 Hz, 1.25 ms tone burst, 1 s sonication, 10 s interstimulus interval).
Flags from extraction
n_subjects— No total or group sizes found in the text.exposures[0].unspecified_domain.isppa_w_cm2— Isppa 42.6 W/cm2 stated to correspond to the selected Ispta; domain not stated; applies to the 2.14 W/cm2 setting.exposures[0].timing.duty_cycle_pct— Duty cycle not stated.exposures[1].free_field.pressure_kpa— In vitro 'peak negative acoustic pressure ... 0.33 W/cm' has mismatched units; not recorded.exposures[1].fundamental_frequency_khz— In vitro frequency assumed same as in vivo only because paper says remaining parameters were the same.