Brief transcranial focused ultrasound stimulation causes lasting modifications to the synaptic circuitry of the hippocampus
William W. Watts, Benjamin Clennell, James K. Jiang, Konoha Izaki-Lee, Abhishek Binodh, Robyn Cuthell, Deniz Tonyali, Jon Crompton, Rosie Taaffe, Alanoud Alqahtani, Anna Andrieu, Igino Rafael Besinga, Kate Heesom, Tom G.J. Steward, Kwangwook Cho, Bruce W. Drinkwater, Elek Molnár, Daniel J. Whitcomb
Brain Stimulation 2025, 18, 1587-1599 · 10.1016/j.brs.2025.08.014
Abstract
Background Brief transcranial focused ultrasound stimulation (tFUS) is used in cognitive mapping, where it is assumed that the intervention itself does not cause lasting modifications to the underlying networks being targeted. However, how so-called 'offline' effects impact the dynamic function of neural circuits is largely unknown. Objectives To determine the persistent effects of ultrasound stimulation on hippocampal circuit function. Methods Acute rat hippocampal slices in vitro, and rat hippocampi in vivo, were exposed to 40 s, 5 Hz pulsed ultrasound or sham stimulation. The effects of ultrasound on the dynamic synaptic and circuit function of the hippocampus were assessed through quantitative proteomics and extracellular field electrophysiology. Results We find that ultrasound stimulation induces persistent and differential changes in protein expression and kinase activity in the hippocampus. This occurs concurrently with an enhancement of basal synaptic transmission and modifications to the susceptibility for the hippocampal circuit to undergo synaptic plasticity. These effects occur via a canonical Akt-dependent metaplastic process. Conclusion The results indicate that tFUS can fundamentally modulate key signalling mechanisms that are responsible for determining the synaptic efficacy in a neural circuit. Importantly, these effects last beyond the duration of the stimulus. These findings provide a mechanistic insight into the sustained impact of tFUS on network function, and emphasise the importance of considering such effects in animal and human studies.
Abstract via europepmc.
Exposures
Exposure 1: In vitro hippocampal slice stimulation, 200 kHz unfocused transducer
Target: hippocampus — “acute hippocampal slice (CA1 stratum radiatum / Schaffer collateral-CA1 circuit)”
Device: other named manufacturer · Farnell · MCUSD25P200B10.7RS-30C
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 200 | ✓✓✓ |
| Pulse duration (ms) | 100 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 5 | ✓✓✓ |
| Duty cycle (%) | not reportedpulse duration × PRF gives 50% | ⚑ |
| Sonication duration (s) | 40 | ✓✓✓ |
| Free-field pressure (kPa) | not reported | |
|---|---|---|
| Free-field Isppa (W/cm²) | 0.34 | ✓✓✓ |
| 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 |
Acute hippocampal slices (400 µm) were placed directly beneath a 200 kHz centre-frequency planar unfocused transducer submerged in a Petri dish of aCSF. Sham stimulation followed the same procedure but the function generator was not activated. A 5 Hz PRF stimulus was chosen based on previous evidence that this pulsing frequency induces sustained off-line neuromodulatory effects.
Exposure 2: 1 MHz focused transducer stimulation of hippocampus (in vivo transcranial and later in vitro slice alignment experiments)
Target: hippocampus — “dorsal hippocampus (in vivo, transcranial) / hippocampal slice (in vitro, aligned with in vivo paradigm)”
Device: other named manufacturer · Precision Acoustics Ltd. · TXH-1-75 ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 1,000 | ✓✓✓ |
| Pulse duration (ms) | 100 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 5 | ✓✓✓ |
| Duty cycle (%) | not reportedpulse duration × PRF gives 50% | |
| Sonication duration (s) | 40 | ✓✓✓ |
| Free-field pressure (kPa) | 140 | ✓✓✓ |
|---|---|---|
| Free-field Isppa (W/cm²) | 0.44, 1.35swept | ✓✓✓⚑ |
| Free-field Ispta (W/cm²) | not reported | |
| In-situ estimate | measurementsingle value | |
| In-situ pressure (kPa) | 45 | ✓✓✓ |
| In-situ Isppa (W/cm²) | 0.14 | ✓✓✓⚑ |
| In-situ Ispta (W/cm²) | not reported |
A 1 MHz focused transducer was used for later in vivo and in vitro experiments, delivering the same 5 Hz PRF, 100 ms pulse length, 40 s total duration protocol. In vivo, ultrasound was delivered transcranially to the left and then right dorsal hippocampus of anaesthetised rats in the same session; free-field Isppa was 0.44 W/cm2, estimated as 0.14 W/cm2 at the rat hippocampus, with an estimated in situ pressure of ~45 kPa determined by quantifying ultrasound transmission through excised rat skull ex vivo. Later in vitro slice experiments used the same 1 MHz transducer at a free-field Isppa of 1.35 W/cm2, with a predicted acoustic pressure at the slice of ~140 kPa measured in a water tank (no skull).
Consistency checks: intensity pressure inconsistent in situ.
Flags from extraction
n_subjects— Multiple sub-experiments used different animal cohorts (N=6-8 animals per assay: proteomics, kinome, electrophysiology, LTP, in vivo, pharmacology); no single overall total number of subjects is stated.exposures[1].free_field.isppa_w_cm2— This exposure merges two different experiment types at the same target and frequency (in vivo transcranial 0.44 W/cm2 and later in vitro slice 1.35 W/cm2) into a list, per the target x frequency exposure rule; domains and measurement contexts (skull vs water tank) differ.exposures[1].in_situ.isppa_w_cm2— In situ Isppa of 0.14 W/cm2 is described as 'estimated'; method of estimation (derating vs ex-vivo skull transmission measurement) is not fully disambiguated in the text.exposures[1].free_field.isppa_w_cm2— The 140 kPa pressure value for the later in vitro slice experiment (Fig. 6B) was reported as a pressure rather than intensity and is described as 'predicted'; placed in free_field.pressure_kpa is omitted here since no matching Isppa was reported for that same measurement, described only in protocol_description.exposures[0].timing.duty_cycle_pct— Duty cycle is not explicitly stated as a percentage; only pulse length (100 ms) and pulse repetition interval (200 ms) are given.