Noninvasive acoustic vagus nerve stimulation modulates Emotion-Related brain networks

The vagus nerve, a critical bidirectional pathway linking the central nervous system and peripheral organ systems, plays a core role in emotional regulation. Here, we introduce a noninvasive acoustic vagus nerve stimulation (aVNS) paradigm and systematically characterize its modulatory effects on emotion-related brain circuitry using in vivo fiber photometry, behavioral assays, and transcriptomic profiling. The pulsed aVNS protocol was defined as follows: fundamental frequency = 1 MHz, sonication duration = 1 s, inter-stimulus interval = 9 s, pulse repetition frequency = 100 Hz, tone-burst duration = 0.5 ms, duty cycle = 5 %, and acoustic pressure = 3 MPa. Acoustic field mapping confirmed a spatially confined ultrasound beam, supporting localized stimulation of the cervical vagus nerve region. In vivo calcium imaging revealed robust aVNS-evoked neural responses across multiple emotion-associated brain regions, with the medial prefrontal cortex (mPFC) showing the strongest response magnitude, as indicated by significantly increased area under the curve values during both the stimulation period (0-1 s, p = 0.0156) and the post-stimulation period (1-10 s, p = 0.0156). Response-profile and inter-regional similarity analyses further demonstrated that aVNS organized distributed neural responses into a coordinated network-level activation pattern rather than inducing isolated regional excitation. Behaviorally, aVNS alleviated anxiety-like and depressive-like behaviors in acute lipopolysaccharide-treated mice (0.5 mg/kg), as shown by increased open-arm exploration in the elevated plus maze (EPM; p = 0.0018) and reduced immobility in the tail suspension test (TST; p = 0.0045), as well as in chronic restraint stress mice (6 h/day for 28 consecutive days), as shown by increased open-arm exploration (p = 0.0167) and reduced immobility time (p = 0.0057). Bulk RNA sequencing further showed that repeated aVNS partially reversed stress-induced transcriptional alterations in the mPFC, particularly inflammation-related signatures. Together, these findings demonstrate that noninvasive aVNS modulates emotion-related brain networks and provide neural, behavioral, and molecular evidence supporting its potential for intervention in emotional dysfunction.