Improved Anatomical Specificity of Non-invasive Neuro-stimulation by High Frequency (5 MHz) Ultrasound
Guo-Feng Li, Hui-Xia Zhao, Hui Zhou, Fei Yan, Jing-Yao Wang, Chang-Xi Xu, Cong-Zhi Wang, Li-Li Niu, Long Meng, Song Wu, Huai-Ling Zhang, Wei-Bao Qiu, Hai-Rong Zheng
Scientific Reports 2016, 6 · 10.1038/srep24738
Abstract
Low frequency ultrasound (<1 MHz) has been demonstrated to be a promising approach for non-invasive neuro-stimulation. However, the focal width is limited to be half centimeter scale. Minimizing the stimulation region with higher frequency ultrasound will provide a great opportunity to expand its application. This study first time examines the feasibility of using high frequency (5 MHz) ultrasound to achieve neuro-stimulation in brain, and verifies the anatomical specificity of neuro-stimulation in vivo. 1 MHz and 5 MHz ultrasound stimulation were evaluated in the same group of mice. Electromyography (EMG) collected from tail muscles together with the motion response videos were analyzed for evaluating the stimulation effects. Our results indicate that 5 MHz ultrasound can successfully achieve neuro-stimulation. The equivalent diameter (ED) of the stimulation region with 5 MHz ultrasound (0.29 ± 0.08 mm) is significantly smaller than that with 1 MHz (0.83 ± 0.11 mm). The response latency of 5 MHz ultrasound (45 ± 31 ms) is also shorter than that of 1 MHz ultrasound (208 ± 111 ms). Consequently, high frequency (5 MHz) ultrasound can successfully activate the brain circuits in mice. It provides a smaller stimulation region, which offers improved anatomical specificity for neuro-stimulation in a non-invasive manner.
Abstract via europepmc.
Exposures
Exposure 1: 1 MHz ultrasound stimulation of mouse brain (tail-flick evoking site)
Target: other — “3.5 mm lateral to the midline and 7.5 mm posterior to the rear corner of mouse eyes”
Device: Olympus / Panametrics · Olympus NDT · V314-1 MHz ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 1,000 | ✓✓✓ |
| Pulse duration (ms) | 0.5 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 1,000 | ✓✓✓ |
| Duty cycle (%) | 50pulse duration × PRF gives 50% | ✓✓✓ |
| Sonication duration (s) | 0.3 | ✓✓✓⚑ |
| 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 | |
| Ispta, domain unspecified (W/cm²) | 0.13, 0.23swept | ✓?⚑ |
All parameters were kept consistent between frequencies (50% duty cycle of pulse, 1 ms pulse period, 1 kHz pulse repetition frequency, 300 pulses per stimulus, and 3 second interval); for 1 MHz, cycles-per-pulse (CPP) was set to 500 to give a 0.5 ms pulse duration. Acoustic intensity was varied over six Ispta values from 130 to 230 mW/cm2, delivered for a stimulation duration of one minute per intensity/site; nine stimulation sites 0.3 mm apart were tested per mouse for the anatomical-specificity comparison.
Exposure 2: 5 MHz ultrasound stimulation of mouse brain (tail-flick evoking site)
Target: other — “3.5 mm lateral to the midline and 7.5 mm posterior to the rear corner of mouse eyes”
Device: Olympus / Panametrics · Olympus NDT · V308-5 MHz ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 5,000 | ✓✓✓ |
| Pulse duration (ms) | 0.5 | ✓✓✓ |
| Pulse repetition frequency (Hz) | 1,000 | ✓✓✓ |
| Duty cycle (%) | 50pulse duration × PRF gives 50% | ✓✓✓ |
| Sonication duration (s) | 0.3 | ✓✓✓⚑ |
| 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 | |
| Ispta, domain unspecified (W/cm²) | 0.13, 0.23swept | ✓?⚑ |
Same block structure as the 1 MHz condition (50% duty cycle, 1 ms pulse period, 1 kHz PRF, 300 pulses per stimulus, 3 s interval); for 5 MHz, CPP was set to 2500 to give the same 0.5 ms pulse duration, with driving amplitude increased to compensate for the ~4.2 dB skull attenuation at 5 MHz. Acoustic intensity was varied over six Ispta values from 130 to 230 mW/cm2, one minute per intensity/site; for the safety/HE-stain cohort, 5 MHz was also delivered continuously for 30 minutes.
Flags from extraction
n_subjects— Paper reports several different n values across analyses (n=5, n=6, n=10, n=20 for HE stain safety group); n_subjects taken from the main Animal preparation paragraph.exposures[0].target.terms— Paper never names the stimulated brain region anatomically; it is identified only by stereotaxic coordinates and by the tail-flick motor response it evokes.exposures[0].unspecified_domain.ispta_w_cm2— Paper does not state whether these Ispta values are free-field or in-brain; they are described only as 'compensated' to offset skull attenuation, so domain is ambiguous and placed in unspecified_domain.exposures[1].unspecified_domain.ispta_w_cm2— Same ambiguity as exposures[0]; both frequencies were driven to the same compensated intensity level.exposures[0].timing.sonication_duration_s— 300 ms stimulus duration is taken from the temperature-safety section (same protocol); the main neuromodulation Methods describe the train as '300 pulses per stimulus' at 1 kHz PRF without directly stating its length in ms, so this reuses the safety-section figure for the same protocol.exposures[1].timing.sonication_duration_s— Same as exposures[0]; 5 MHz was also tested at a shorter 150 ms duration for thermal safety, noted in protocol_description.