Medial prefrontal cortex-targeted low-intensity focused ultrasound alleviates migraine-related allodynia and affective symptoms via modulating glutamate/GABA balance in mice
Jiayi Liu, Mingjie Zhang, Qiuquan Cai, Chunxiao Yang, Yajin Wu, Weinan Na, Deqi Zhai, Huijuan Yuan, Ya Cao, Xiaoya Chen, Yingji Li, Wenjing Tang, Dengfa Zhao, Ruozhuo Liu, Zhao Dong, Jiajia Yang, Dong Ming, Shengyuan Yu
The Journal of Headache and Pain 2026 · 10.1186/s10194-026-02377-0
Abstract
Background Migraine affects nearly one billion people worldwide and remains a major global health burden with inadequate treatment options. In this study, transcranial low-intensity focused ultrasound (LIFU) targeting the medial prefrontal cortex (mPFC) was assessed as a novel, non-invasive neuromodulatory strategy for migraine abortive treatment. The therapeutic potential and safety of LIFU were evaluated in animal models to support its future clinical translation. Methods Male C57BL/6 mice were administered intraperitoneally with nitroglycerin (NTG) to induce an acute migraine model, whereas control animals received vehicle (VEH). Following model induction, mice were subjected to either mPFC-targeted LIFU or sham stimulation, which underwent an identical protocol without ultrasonic output. A series of behavioral, histological, and molecular assays was performed to evaluate the therapeutic effects and neuromodulatory mechanisms of LIFU. Mechanical allodynia was assessed using the von Frey test; anxiety-like behavior was evaluated in the elevated plus maze. Neuronal activation was examined via c-Fos immunofluorescence and GABA/glutamate co-staining. Tissue safety was assessed by HE staining and TUNEL assay. All quantitative analyses were conducted under blinded conditions. Results NTG injection induced significant allodynia and anxiety-like behaviors. LIFU stimulation significantly attenuated cephalic and plantar allodynia and ameliorated anxiety-like behaviors compared to the sham group. The NTG-induced migraine model exhibited significant mPFC c-Fos hyperactivation, which LIFU stimulation effectively suppressed. NTG injection significantly increased the proportion of activated glutamatergic neurons (GLU-N) and decreased activated GABAergic neurons (GABA-Ns) among total c-Fos-positive cells in the mPFC. This imbalance was reversed by LIFU, i.e., it was characterized by a decrease in activated GLU-Ns and an increase in activated GABA-Ns. No significant histopathological damage or apoptosis was detected following LIFU exposure. Conclusions Aberrant activation and excitatory and inhibitory (E/I) imbalance of neurons in mPFC were involved in acute NTG-induced episodes. The LIFU targeting mPFC could alleviate NTG-induced mechanical allodynia and anxiety-like behavior by restoring E/I balance. LIFU is a novel, safe, non-invasive neuromodulatory strategy offering a potential migraine treatment.
Abstract via europepmc.
Exposures
Exposure 1: mPFC (PL) LIFU
Target: ventromedial prefrontal cortex — “prelimbic (PL) subdivision of the mPFC”
Device: Olympus / Panametrics · Olympus Corporation of the Americas · V318 ✓
| Waveform | pulsed | |
|---|---|---|
| Fundamental frequency (kHz) | 500 | ✓✓✓ |
| Pulse duration (ms) | 0.3 | ✓✓✓⚑ |
| Pulse repetition frequency (Hz) | 2,000 | ✓✓✓ |
| Duty cycle (%) | 60pulse duration × PRF gives 60% | ✓✓✓ |
| Sonication duration (s) | 0.05 | ✓✓✓ |
| 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) | 0.6 | ✓✓✓⚑ |
Isppa not given; Ispta 600 mW/cm2, PRF 2 kHz, sonication duration 0.05 s, inter-stimulation interval 2 s, duty cycle 60%, tone-burst duration 0.3 ms. LIFU began 20 min after NTG injection under 0.5% isoflurane; the number or total duration of stimulations is not stated.
Flags from extraction
n_subjects— 9 mice per group in the behavioural cohort (NTG+LIFU); separate cohorts of 6 per group (immunofluorescence) and 3 per group (histology) were also used, total exposed not statedexposures[0].unspecified_domain.ispta_w_cm2— Ispta 600 mW/cm2 stated without saying whether free field or in situexposures[0].timing.pulse_duration_ms— Tone-burst duration 0.3 ms is stated; it is consistent with duty cycle / PRFprovenance.auditory_control— No auditory control used; authors argue 0.5 MHz lies outside the mouse hearing rangeblinding— Paper states quantitative analyses were blinded and describes the histology as double-blind; blinding level of the stimulation itself is unclearexposures[0].timing— Sham is anaesthesia and positioning without ultrasonic output; no VEH+LIFU group