Focal neurostimulation of propagating calcium waves in human neural cells using megahertz-range single-pulse focused ultrasound
Tom Aubier, Ivan M. Suarez-Castellanos, Magali Perier, Gilles Huberfeld, Alexandre Carpentier, W. Apoutou N'Djin
2025 · 10.1101/2025.07.02.662731
Abstract
Focused ultrasound (FUS) offers a promising neurostimulation technique that addresses the trade-off between invasiveness and spatial selectivity found in conventional electromagnetic-based methods. Evidence supporting its theoretical selectivity and comprehensive descriptions of the underlying biophysical mechanisms remain insufficient to ensure adequate control and safety. Based on observations of causal , focal , and propagating calcium waves in human neural cells in vitro , evoked using single-pulse megahertz-range FUS, a transdural FUS approach relying on an intracranial implant is proposed to overcome frequency limitations associated with transcranial ultrasound. Using an extended parameter space (0.7-8 MHz), calcium waves were shown to be selectively elicited by cavitation or acoustic radiation force. Cavitation, predominant at frequencies under 5 MHz, induced dispersed and unpredictable responses that could lead to cell damage. FUS radiation force however, predominant at frequencies over 5 MHz, elicited focal and predictable responses, without affecting cell viability. FUS-evoked intercellular calcium waves were shown to propagate across the surrounding neural network via intra cellular and extra cellular pathways driven by calcium amplification mechanisms. Collectively, these findings lay the foundation for developing wearable neurostimulation approaches to manage chronic neurological conditions.
Abstract via europepmc.
Exposures
Exposure 1: Single 400 us pulse FUS at 0.73-8.14 MHz on human neural cell culture
Target: cultured neurons — “in vitro human neural cells (2D network)”
Device: custom-built · homemade single-element spherical ceramic transducer (C-21, Fuji Ceramics Corp.), 15 mm diameter and focal length ✓
| Waveform | continuous | |
|---|---|---|
| Fundamental frequency (kHz) | 730, 2,230, 5,190, 8,140swept | ✓✓✓⚑ |
| Pulse duration (ms) | not applicable | |
| Pulse repetition frequency (Hz) | not applicable | |
| Duty cycle (%) | not applicable | |
| Sonication duration (s) | 0.0004 | ✓✓✓⚑ |
| 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 | |
| Pressure, domain unspecified (kPa) | 780, 7,780, 8,790, 9,300swept | ✓✓✓⚑ |
| Isppa, domain unspecified (W/cm²) | 180, 4,210, 6,890, 7,170swept | ✓✓✓⚑ |
Single 400 us pulse (amplitude-modulated continuous sinusoid) at 0.73, 2.23, 5.19 or 8.14 MHz, triggered 10 s after the start of fluorescence recording; pressure escalated to find the minimum threshold at each frequency.
Flags from extraction
exposures[0].unspecified_domain.pressure_kpa— Spatial-peak RMS pressure p_spr from numerical simulation of the dish (Table 1), MPa converted to kPa; RMS rather than peak amplitude; values are medians with IQR/2 across trials at threshold-level drive.exposures[0].unspecified_domain.isppa_w_cm2— Table 1 gives Isppa in kW/cm2 (0.18, 4.21, 6.89, 7.17), converted to W/cm2; simulated, linear model, not measured.exposures[0].timing.sonication_duration_s— 400 us single pulse, recorded as 0.0004 s.exposures[0].fundamental_frequency_khz— One transducer driven at fundamental and harmonics; frequencies from the transducer impedance measurement.n_subjects— Number of cultures or dishes not stated; only 78% success over 18 trials at 8.14 MHz.sham_type— Sham stimulation is mentioned for the viability controls but its mechanism is not described.exposures[0].target.terms— Neural progenitor-derived culture containing neurons and glia; cultured_neurons chosen.