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Transient ultrasound stimulation has lasting effects on neuronal excitability

Benjamin Clennell, Tom G.J. Steward, Meg Elley, Eunju Shin, Miles Weston, Bruce W. Drinkwater, Daniel J. Whitcomb

Brain Stimulation 2021, 14, 217-225 · 10.1016/j.brs.2021.01.003

in vitro cellhealthyinvasive electrophysiologycellular imaging

Abstract

Background Transcranial ultrasound stimulation can acutely modulate brain activity, but the lasting effects on neurons are unknown. Objective To assess the excitability profile of neurons in the hours following transient ultrasound stimulation. Methods Primary rat cortical neurons were stimulated with a 40 s, 200 kHz pulsed ultrasound stimulation or sham-stimulation. Intrinsic firing properties were investigated through whole-cell patch-clamp recording by evoking action potentials in response to somatic current injection. Recordings were taken at set timepoints following ultrasound stimulation: 0-2 h, 6-8 h, 12-14 h and 24-26 h. Transmission electron microscopy was used to assess synaptic ultrastructure at the same timepoints. Results In the 0-2 h window, neurons stimulated with ultrasound displayed an increase in the mean frequency of evoked action potentials of 32% above control cell levels (p = 0.023). After 4-6 h this increase was measured as 44% (p = 0.0043). By 12-14 h this effect was eliminated and remained absent 24-26 h post-stimulation. These changes to action potential firing occurred in conjunction with statistically significant differences between control and ultrasound-stimulated neurons in action potential half-width, depolarisation rate, and repolarisation rate, that were similarly eliminated by 24 h following stimulation. These effects occurred in the absence of alterations to intrinsic membrane properties or synaptic ultrastructure. Conclusion We report that stimulating neurons with 40 s of ultrasound enhances their excitability for up to 8 h in conjunction with modifications to action potential kinetics. This occurs in the absence of major ultrastructural change or modification of intrinsic membrane properties. These results can inform the application of transcranial ultrasound in experimental and therapeutic settings.

Abstract via europepmc.

Speciesrat (Wistar, post-natal day 0)
Subjectsnot reported cultures
Sessions per subjectnot applicable
Randomisednot reported
Blindingnot reported
Sham / controlinactive transducer
Auditory controlnot reported
Readout timingoffline
Anaesthesianot applicable
Readoutsinvasive electrophysiology, cellular imagingwhole-cell patch-clamp recording of evoked action potentials and passive membrane properties; transmission electron microscopy of pre-/post-synaptic ultrastructure
Direction of effectexcitatoryUltrasound-stimulated neurons showed a 32% increase in mean evoked spike frequency at 0-2 h and 44% at 6-8 h post-stimulation versus sham; effect was eliminated by 12-14 h and absent at 24-26 h.
Adverse eventsnot applicable

Exposures

Exposure 1: Primary rat cortical neuron culture, single 40 s pulsed ultrasound stimulation

Target: cultured neurons — “primary rat cortical neurons in culture
Device: other named manufacturer · Farnell, UK · MCUSD19A200B11RS

Pulse timing
Waveformpulsed
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✓✓
Pressure and intensity, by domain
Free-field pressure (kPa)not reported
Free-field Isppa (W/cm²)not reported
Free-field Ispta (W/cm²)not reported
In-situ estimatenot applicable
In-situ pressure (kPa)not applicable
In-situ Isppa (W/cm²)not applicable
In-situ Ispta (W/cm²)not applicable
Protocol, in the paper’s words

200 kHz sinusoidal wave, 50 V peak-to-peak transducer drive amplitude, delivered in 100 ms pulses at 100 ms intervals for 40 s total (equating to a 5 Hz modulation/pulse-repetition frequency). Sham stimulation used the same setup and duration with all equipment powered on but no excitation signal generated.

Flags from extraction

  • exposures[0].timing.duty_cycle_pct100 ms pulses at 100 ms intervals imply a 50% duty cycle, but the paper never states a duty-cycle percentage directly, so it is left not_reported rather than computed.
  • n_subjectsN (number of ultrasound-stimulated neurons) differs across the four post-stimulation timepoints (15, 14, 12, 13); no single total is given.
  • device.manufacturerFarnell is the distributor named alongside the transducer part number; the original manufacturer is not stated.