Ultrasound enhances glymphatic-associated solute transport via Piezo1-related mechanotransduction in 5xFAD mice

Glymphatic dysfunction contributes to impaired clearance of neurotoxic proteins (e.g. Aβ) and AD progression. Low-intensity transcranial focused ultrasound (LITFUS) is a noninvasive mechanomodulation strategy that can enhance glymphatic CSF transport via acoustic radiation force. However, its therapeutic efficacy in promoting glymphatic Aβ clearance to alleviate cerebral amyloid burden and cognitive decline remains to be established. 5xFAD mice underwent bilateral hippocampal LITFUS for 4 weeks. Glymphatic transport was assessed using fluorescent tracers administered via intracisternal or intrahippocampal injection, followed by ex vivo imaging. Soluble Aβ levels in the deep cervical lymph nodes (dCLNs) were also measured to evaluate downstream drainage. Immunohistochemistry was used to assess Aβ_1-42 deposition and Iba1 immunoreactivity, and cognitive function was evaluated using behavioral tests. Piezo1 involvement was examined using pharmacological inhibition with GsMTx4 together with qPCR and Western blot analyses of isolated brain vascular fractions. Safety was evaluated by blood-brain barrier permeability assays and H&E staining. Compared with sham treatment, LITFUS significantly increased brain-wide cerebrospinal fluid tracer influx and enhanced hippocampal interstitial solute transport in 5xFAD mice. LITFUS also increased soluble Aβ levels in the dCLNs, reduced Aβ deposition in the hippocampus and prefrontal cortex, decreased Iba1 immunoreactivity, and improved learning and memory performance. These effects were accompanied by increased Piezo1 expression in isolated brain vascular fractions and enhanced CaMKII/eNOS signaling. Pharmacological inhibition with GsMTx4 attenuated the LITFUS-induced improvements in glymphatic transport, reduction of Aβ burden, and downstream signaling. LITFUS did not induce detectable blood-brain barrier disruption or histological injury. LITFUS noninvasively enhances glymphatic transport, reduces cerebral Aβ burden, and improves cognitive function in 5xFAD mice. The findings support a role for Piezo1-related mechanotransduction in mediating these effects and highlight the translational potential of LITFUS as a noninvasive therapeutic approach for AD.