Low‐intensity pulsed ultrasound ameliorates depression‐like behaviors in a rat model of chronic unpredictable stress

Jinniu Zhang, Hui Zhou, Jian Yang, Jun Jia, Lili Niu, Zuoli Sun, Dandan Shi, Long Meng, Weibao Qiu, Xiaomin Wang, Hairong Zheng, Gang Wang

CNS Neuroscience & Therapeutics 2021, 27, 233-243 · 10.1111/cns.13463

rodentdepressionbehaviourhistology molecular

Abstract

Introduction There is an unmet need for better nonpharmaceutical treatments for depression. Low-intensity pulsed ultrasound (LIPUS) is a novel type of neuromodulation that could be helpful for depressed patients. Objective The goal of this study was to investigate the feasibility and potential mechanisms of LIPUS in the treatment of depression. Methods Chronic unpredictable stress (CUS) was used to generate rats with depression-like features that were treated with four weeks of LIPUS stimulation of the ventromedial prefrontal cortex. Depression-like behaviors were assessed with the sucrose preference, forced swim, and open field tests. BDNF/mTORC1 signaling was examined by Western blot to investigate this potential molecular mechanism. The safety of LIPUS was evaluated using hematoxylin-eosin and Nissl staining. Results Four weeks of LIPUS stimulation significantly increased sucrose preference and reduced forced swim immobility time in CUS rats. LIPUS also partially reversed the molecular effects of CUS that included decreased levels of BDNF, phosphorylated tyrosine receptor kinase B (TrkB), extracellular signal-regulated kinase (ERK), mammalian target of rapamycin complex 1 (mTORC1), and S6 kinase (S6K). Moreover, histological staining revealed no gross tissue damage. Conclusions Chronic LIPUS stimulation can effectively and safely improve depression-like behaviors in CUS rats. The underlying mechanisms may be related to enhancement of BDNF/ERK/mTORC1 signaling pathways in the prefrontal cortex (PFC). LIPUS is a promising noninvasive neuromodulation tool that merits further study as a potential treatment for depression.

Abstract via europepmc.

Speciesrat (Sprague-Dawley)

Subjects8 animals

Sessions per subjectnot reported

Randomisedyes

Blindingnot reported

Sham / controlinactive transducer

Auditory controlnot reported

Readout timingoffline

Anaesthesiaawake

Readoutsbehaviour, histology molecular — Sucrose preference test (SPT); forced swim test (FST); open field test (OFT); Western blot (BDNF, TrkB, ERK, Akt, mTOR, S6K); c-Fos immunohistochemistry; hematoxylin-eosin and Nissl staining

Direction of effectexcitatorySingle 200 Hz LIPUS stimulation of the vmPFC increased c-Fos-positive neuron counts (285 Hz and 500 Hz did not alter behavior); four weeks of chronic 200 Hz LIPUS improved depression-like behaviors (increased sucrose preference, decreased FST immobility) and partially restored BDNF/TrkB/ERK/mTORC1/S6K signaling in CUS rats.

Adverse eventsnone observedHE and Nissl staining revealed intact brain structure and neurons with no microhemorrhage, edema, cell necrosis, or local inflammatory response in any group; measured temperature elevation was theoretically calculated to be less than 0.4 degrees C; negative acoustic pressure (0.28 MPa in situ) was far below the cavitation threshold (40 MPa) and intensity/MI were below FDA diagnostic ultrasound limits.

Exposures

Exposure 1: LIPUS to ventromedial prefrontal cortex (vmPFC), PRF screening (200/285/500 Hz) and chronic 200 Hz stimulation

Target: ventromedial prefrontal cortex — “ventromedial prefrontal cortex (vmPFC)”
Device: custom-built

Pulse timing
Waveform	pulsed
Fundamental frequency (kHz)	800	✓✓✓
Pulse duration (ms)	0.2	✓✓✓
Pulse repetition frequency (Hz)	200, 285, 500swept	✓✓✓⚑
Duty cycle (%)	4	✓✓✓
Sonication duration (s)	1	✓✓✓

Pressure and intensity, by domain
Free-field pressure (kPa)	400	✓✓✓
Free-field Isppa (W/cm²)	6.2	✓✓✓
Free-field Ispta (W/cm²)	0.248	✓✓✓
In-situ estimate	measurementsingle value
In-situ pressure (kPa)	280	✓✓✓
In-situ Isppa (W/cm²)	3.84	✓✓✓
In-situ Ispta (W/cm²)	0.154	✓✓✓

Protocol, in the paper’s words

Parameter screening: normal rats received a single 30-minute LIPUS session of the vmPFC at one of three PRFs (200, 285, or 500 Hz); only 200 Hz significantly reduced FST immobility, so 200 Hz was used going forward. Chronic study: rats in the CUS+LIPUS group received ultrasound stimulation for twenty minutes each day for four consecutive weeks (CUS+sham LIPUS and control groups wore the same apparatus with power turned off). Chronic parameters: 200 Hz PRF, 4% duty cycle, 0.2 ms tone burst duration, 1 s sonication duration, 3 s inter-stimulus interval.

Flags from extraction

n_subjects — Reported total covers only the chronic CUS+LIPUS group (n=8); a separate parameter-screening experiment also exposed normal rats to single LIPUS sessions (n=6-8 per PRF group across 3 PRFs), an imprecise range not included in this total.
exposures[0].timing.pulse_repetition_frequency_hz — The three PRFs (200/285/500 Hz) were tested only in the single-dose screening experiment; the chronic 4-week CUS study used only 200 Hz with the other timing parameters (DC, TBD, SD, ISI) given specifically for the chronic protocol.
randomised — Initial split into control/CUS groups was explicitly randomized (stratified); the subsequent redistribution of CUS rats into CUS+sham-LIPUS and CUS+LIPUS groups was based on sucrose preference index stratification rather than stated as random.