Low‐Intensity Pulsed Ultrasound Alleviation of LPS‐Induced Depression‐Like Behavior via Microglial P2X4R Inhibition and BDNF/TrkB Pathway Activation

Abstract

Objective Low-intensity pulsed ultrasound (LIPUS) shows promising anti-inflammatory and neuroprotective effects for different types of neurological disorders. This study aims to investigate the therapeutic effects of LIPUS on LPS-induced depression-like behavior and neuroinflammation and to elucidate the underlying molecular mechanisms. Methods A depressive mouse model is established by intraperitoneal injection of LPS (1.0 mg/kg/day for 7 days). LIPUS is applied to the hippocampal region (30 min/day). Behavioral assessments include the open field test (OFT), forced swim test (FST), and tail suspension test (TST). Molecular analyses, including Western blotting, immunofluorescence, and qPCR, are performed to evaluate the expression of P2X4R, IBA1, inflammatory cytokines (IL-1β, IL-6, TNF-α), BDNF/TrkB signaling pathway, and apoptosis-related proteins (Bax, Bcl-2). The involvement of P2X4R is further examined using ivermectin (IVM), a selective P2X4R agonist. Results LIPUS significantly alleviates the LPS-induced depression-like behavior, suppresses hippocampal pro-inflammatory cytokine expression, inhibits microglial activation, and reduces neuronal apoptosis. Mechanistically, LIPUS downregulates P2X4R and IBA1, upregulates BDNF protein levels and TrkB phosphorylation, and modulates the Bax and Bcl-2 expression. Co-localization studies confirm that P2X4R is predominantly expressed in microglia, and LIPUS markedly reduces the overlap. Notably, the anti-inflammatory, neuroprotective, and antidepressant effects of LIPUS are significantly attenuated by IVM, highlighting the critical role of P2X4R suppression in mediating therapeutic effects. Conclusion LIPUS mitigates LPS-induced neuroinflammation, neuronal apoptosis, and depression-like behavior by targeting microglial P2X4R and activating the BDNF/TrkB pathway. The findings provide mechanistic insights and demonstrate that LIPUS is a promising non-pharmacological intervention for depression, underscoring the translational potential of P2X4R as a therapeutic target.

Abstract via europepmc.

Flags from extraction

• n_subjects — the paper states each experimental group (including Control, LPS, LIPUS, IVM, KN-93 across two experiment parts) contains n=8, but never gives a single total number of animals exposed to ultrasound; group sizes for the LIPUS-receiving arms (part-1 LIPUS group and part-2 LIPUS, IVM, KN-93 groups) are listed rather than summed, and it is unclear whether the 'LIPUS group' in part 2 is the same cohort reused from part 1 or a separate n=8 cohort.
• n_sessions_per_subject — LIPUS is stated to be applied '30 min/day' but the total number of treatment days is not explicitly stated in the Methods; the co-administered LPS/IVM/KN-93 dosing regimens ran for 7 days, which may or may not match the LIPUS treatment schedule.
• anaesthesia — the paper does not state whether animals were awake or anaesthetised during LIPUS delivery (anaesthesia is mentioned only for terminal tissue collection procedures).
• exposures[0].device.family — the ultrasound transducer manufacturer/model is not named in the main text (only 'an ultrasound transducer operating at 800 kHz' with a custom-fabricated collimator).
• exposures[0].timing.pulse_duration_ms — pulse duration is not explicitly stated; only PRF (100 Hz) and duty cycle (10%) are given, from which pulse duration could be computed (1 ms) but this was not done per instructions.
• sham_type — the LPS-only group serves as the disease comparator but there is no explicit sham-ultrasound (inactive transducer) arm described, so sham_type is set to 'none'.