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Low-Intensity Pulsed Ultrasound Attenuates Postoperative Neurocognitive Impairment and Salvages Hippocampal Synaptogenesis in Aged Mice

Qian Wang, Taotao Liu, Huixian Chang, Zhengqian Li, Lei Chen, Xinning Mi, Huayi Xing, Xiaoxiao Wang, Jingshu Hong, Kaixi Liu, Yitong Li, Dengyang Han, Yue Li, Ning Yang, Xiaoli Li, Yingwei Li, Xiangyang Guo

Brain Sciences 2023 · 10.3390/brainsci13040657

rodentotherbehaviourhistology molecular

Abstract

Postoperative neurocognitive impairment is an urgent problem with global aging accelerating. The prevention and treatment of postoperative neurocognitive impairment have been widely investigated but lack effective strategies. Low-intensity pulsed ultrasound (LIPUS), a non-invasive tool, has shown an effect on neuroprotection, but whether it could attenuate the postoperative neurocognitive impairment and the underlying mechanisms remains unknown. An experimental setup for LIPUS stimulation of the hippocampus was well established. A laparotomy model in aged mice was applied, and a Morris water maze was used to assess cognitive function. RT-qPCR and western blotting were used to detect levels of Piezo1, synapse-associated proteins in the hippocampus, respectively. Immunofluorescent staining was also used to determine the neural activation and Piezo1 expression. The results showed that LIPUS increased synapse-related proteins of the hippocampus and attenuated cognitive impairment in aged mice. Meanwhile, LIPUS suppressed the overexpression of Piezo1 in the hippocampus. We further found that LIPUS promoted Calpain1 activity and increased extracellular regulated protein kinases (Erk) phosphorylation. Our results suggested that LIPUS could improve cognitive impairment and increase hippocampal synaptogenesis through the Piezo1-mediated Calpain1/ Erk pathway. LIPUS could be used as an effective physical intervention to alleviate postoperative cognitive dysfunction in the aged population.

Abstract via europepmc.

SpeciesC57BL/6 mouse (aged, 16-18 months)
Subjects12, 12swept animals
Sessions per subject4
Randomisedyes
Blindingnot reported
Sham / controlno treatment control
Auditory controlnot reported
Readout timingoffline
Anaesthesiaanaesthetised
Readoutsbehaviour, histology molecularMorris water maze (escape latency, platform crossings, swimming speed); RT-qPCR and Western blot for Piezo1, synaptic proteins (BDNF, synaptophysin, PSD95), Calpain1/2, PHLPP, Erk/p-Erk and inflammatory cytokines (TNF-alpha, IL-1beta, IL-6); immunofluorescence for c-Fos and Piezo1; hematoxylin-eosin staining
Direction of effectexcitatoryLIPUS applied to the dorsal hippocampus increased c-Fos expression (neuronal activation), attenuated anaesthesia/surgery-induced increases in Morris water maze escape latency, reduced hippocampal neuroinflammatory cytokines, restored synapse-related proteins (BDNF, synaptophysin, PSD95), suppressed Piezo1 overexpression, and reversed Calpain1/PHLPP/Erk pathway changes in aged mice after anaesthesia/surgery.
Adverse eventsnone observedHematoxylin-eosin staining showed no obvious histological changes in the hippocampus after LIPUS exposure compared with control, and LIPUS alone did not increase hippocampal neuroinflammatory cytokines compared with the control group.

Exposures

Exposure 1: LIPUS to dorsal hippocampus

Target: hippocampus — “dorsal hippocampus
Device: not reported

Pulse timing
Waveformpulsed
Fundamental frequency (kHz)1,000✓✓
Pulse duration (ms)50✓✓
Pulse repetition frequency (Hz)1✓✓
Duty cycle (%)5pulse duration × PRF gives 5%✓✓
Sonication duration (s)150✓✓
Pressure and intensity, by domain
Free-field pressure (kPa)not reported
Free-field Isppa (W/cm²)not reported
Free-field Ispta (W/cm²)not reported
In-situ estimatederatingsingle value
In-situ pressure (kPa)not reported
In-situ Isppa (W/cm²)not reported
In-situ Ispta (W/cm²)0.177✓✓
Protocol, in the paper’s words

The duration of sonication was alternated bilaterally for 2.5 min and repeated three times, for a total stimulation time of 15 min, delivered at four time points around the anesthesia/surgery: 8 h before, 4 h before, 4 h after and 8 h after surgery. A calibrating cone was used so the focal point fell on the dorsal hippocampus (bregma +-1.40 mm, caudal -1.94 mm).

Flags from extraction

  • exposures[0].timing.pulse_duration_ms,exposures[0].timing.sonication_duration_sFigure 1D explicitly labels the 50 ms value 'sonication duration (SD)'. Duty cycle (5%) divided by PRF (1 Hz, i.e. a 1 s period) arithmetically equals 50 ms, which would normally identify this as the per-pulse ON duration (pulse duration) rather than a train duration under the burst rule; the paper's own label was followed (sonication_duration_s=0.05) and pulse_duration_ms left not_reported rather than resolved by arithmetic.
  • exposures[0].in_situ.methodThe paper states the intensity value is 'after transcranial attenuation' but does not specify whether this was measured through an excised skull or computed using an assumed attenuation coefficient; classified as 'derating' with this ambiguity noted.
  • deviceNo manufacturer or model is given for the ultrasound transducer itself; only the unrelated stereotaxic apparatus (RWD Life Science) is named.
  • n_subjectsDifferent subsets of mice were used for different assays (Morris water maze n=12/group, RT-qPCR n=6/group, Western blot n=4/group, immunofluorescence n=3/group across four groups); no single total number of ultrasound-exposed animals is stated.