Non-genetic photoacoustic stimulation of single neurons by a tapered fiber optoacoustic emitter
Linli Shi, Ying Jiang, Fernando R. Fernandez, Guo Chen, Lu Lan, Heng-Ye Man, John A. White, Ji-Xin Cheng, Chen Yang
Light: Science & Applications 2021, 10 · 10.1038/s41377-021-00580-z
Abstract
Neuromodulation at high spatial resolution poses great significance in advancing fundamental knowledge in the field of neuroscience and offering novel clinical treatments. Here, we developed a tapered fiber optoacoustic emitter (TFOE) generating an ultrasound field with a high spatial precision of 39.6 µm, enabling optoacoustic activation of single neurons or subcellular structures, such as axons and dendrites. Temporally, a single acoustic pulse of sub-microsecond converted by the TFOE from a single laser pulse of 3 ns is shown as the shortest acoustic stimuli so far for successful neuron activation. The precise ultrasound generated by the TFOE enabled the integration of the optoacoustic stimulation with highly stable patch-clamp recording on single neurons. Direct measurements of the electrical response of single neurons to acoustic stimulation, which is difficult for conventional ultrasound stimulation, have been demonstrated. By coupling TFOE with ex vivo brain slice electrophysiology, we unveil cell-type-specific responses of excitatory and inhibitory neurons to acoustic stimulation. These results demonstrate that TFOE is a non-genetic single-cell and sub-cellular modulation technology, which could shed new insights into the mechanism of ultrasound neurostimulation.
Abstract via europepmc.
Exposures
Exposure 1: Single-cell / subcellular TFOE stimulation of cultured primary cortical neurons (calcium imaging)
Target: cultured neurons — “primary rat cortical neurons”
Device: custom-built · tapered fiber optoacoustic emitter (TFOE)
| Waveform | pulsed, other_patterned | |
|---|---|---|
| Fundamental frequency (kHz) | 6,600 | ✓✓✓ |
| Pulse duration (ms) | not reported | |
| Pulse repetition frequency (Hz) | 1,700 | ✓✓✓ |
| Duty cycle (%) | not reported | |
| Sonication duration (s) | 0.05, 0.001swept | ✓✓✓ |
| Free-field pressure (kPa) | 2,700 | ✓✓✓⚑ |
|---|---|---|
| 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 |
A TFOE was placed ~5 um from a targeted neuron. Laser pulse trains at 1030 nm, 1.7 kHz repetition rate, of either 50 ms duration (7.8 mW, 85 pulses) or 1 ms duration (two pulses, 11.4 mW) were delivered to evoke calcium transients. Separately, single-laser-pulse (3 ns) stimulation was used to test threshold pulse energy (1-8 pulses) and to demonstrate sub-microsecond acoustic pulses as the shortest stimuli reported for neuron activation.
Exposure 2: TFOE stimulation of neurons in acute mouse cortical brain slices with whole-cell patch-clamp recording
Target: brain slice — “mouse cortical slices”
Device: custom-built · tapered fiber optoacoustic emitter (TFOE)
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 6,600 | ✓✓✓ |
| Pulse duration (ms) | not reported | |
| Pulse repetition frequency (Hz) | 1,700 | ✓✓✓⚑ |
| Duty cycle (%) | not reported | |
| Sonication duration (s) | 0.005 | ✓✓✓⚑ |
| Free-field pressure (kPa) | 2,700 | ✓✓✓ |
|---|---|---|
| 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 |
TFOE was integrated with a patch pipette and placed ~5-10 um from GAD2-tdTomato-negative pyramidal neurons or GAD2-tdTomato-positive inhibitory interneurons in mouse cortical slices; laser pulse trains of 1030 nm, 11.4 mW, 1.7 kHz repetition rate and 5 ms duration were used while recording membrane voltage under current-clamp.
Flags from extraction
exposures[1].timing.pulse_repetition_frequency_hz— Value (Laser: 11.4 mW, 1.7 kHz, 5 ms duration) is stated only in a figure caption (Fig. 5), not in the main running text describing the patch-clamp experiment.exposures[1].timing.sonication_duration_s— Same figure-caption source as above.exposures[0].free_field.pressure_kpa— 2.7 MPa is the headline device characterization (measured in water at the emitter tip, ~0 um distance); actual pressure delivered to cells stimulated at ~5-10 um distance is lower (paper separately states 1.9 MPa at 10 um) and was not applied uniformly to every reported exposure condition.sham_type— Controls used (TTX pharmacology; uncoated bare-fiber 'laser only' control) do not map cleanly onto the closed sham_type vocabulary; coded as 'other'.model_system— This is a photoacoustic (optically-driven) ultrasound emitter applied at single-cell/tissue-slice range rather than a conventional transcranial/piezoelectric transducer study; included per batch listing.n_subjects— Paper reports per-experiment sample sizes (e.g. N=6 from 3 cultures; N=8 cells from three cultures; 10 neurons tested) but never a single aggregate count of cultures/preparations used across the whole study.