Mitigation of Aβ 25-35 -Induced Cognitive Deficits in C57BL/6 Mice via Thermal Cycling Stimulation Employing Focused Ultrasound

Guan-Bo Lin, Hsu-Hsiang Liu, Yu-Yi Kuo, You-Ming Chen, Fang-Tzu Hsu, Yu-Wei Wang, Yi Kung, Chien Ching, Chih-Yu Chao

2025 · 10.64898/2025.12.10.693347

rodentalzheimers diseasebehaviourhistology molecular

Abstract

Hyperthermia (HT) is recognized across various medical disciplines for its capacity to modulate specific protein expressions. In efforts to improve Alzheimer’s disease (AD), HT has the potential to regulate heat shock proteins (HSPs) and antioxidant enzymes, which helps decrease the aberrant accumulation of β-amyloid (Aβ) protein and oxidative stress. Nonetheless, the precise delivery of mild hyperthermia to the brain remains a significant challenge. To apply mild hyperthermia targeted to the brain and evaluate its impact on cognitive improvement, this study used focused ultrasound (FUS) to administer localized mild hyperthermia to the brains of AD mouse induced by intracerebroventricular (i.c.v.) injection of Aβ 25-35 . For considerations of safety and therapeutic efficacy, a thermal cycling-hyperthermia (TC-HT) protocol was adapted into a focused ultrasound-mediated thermal cycling stimulation (FUS-TCS), which was compared with the continuous focused ultrasound-mediated hyperthermia stimulation (FUS-HTS). The findings revealed that the FUS-TCS treatment group exhibited a significant improvement in cognitive performance, as evidenced by enhanced outcomes in the Y-maze and novel object recognition (NOR) tests. Furthermore, this group demonstrated increased expression of Aβ-degrading enzymes and antioxidant proteins, including heat shock protein 70 (HSP70), neprilysin (NEP), insulin degrading enzyme (IDE), sirtuin 1 (SIRT1), and superoxide dismutase 2 (SOD2). These results suggest that localized mild hyperthermia targeting the brain using FUS-TCS treatment represents a promising strategy for ameliorating cognitive deficits associated with AD.

Abstract via europepmc.

Speciesmouse (C57BL/6)

Subjects8, 8, 8, 8swept animals

Sessions per subject3

Randomisedyes

Blindingnot reported

Sham / controlno treatment control

Auditory controlnot reported

Readout timingoffline

Anaesthesiaanaesthetised

Readoutsbehaviour, histology molecular — Y-maze spontaneous alternation and total arm entries; novel object recognition (NOR) test (1 h and 24 h discrimination index); Western blot for Aβ, HSP70, IDE, NEP, SIRT1, and SOD2 in hippocampus

Direction of effectnot assessedStudy assessed thermal neuromodulation (mild hyperthermia) effects on cognition and protein expression rather than direct neuronal excitability; FUS-TCS improved behavioural outcomes and upregulated neuroprotective/antioxidant proteins relative to Aβ-only and continuous FUS-HTS groups.

Adverse eventsobservedFUS-HTS (continuous 30-min heating) significantly reduced the spontaneous alternation index (51.0% vs 60.4% control) and reduced NOR discrimination indices (63.1% to 42.1% at 1 h; 61.7% to 42.6% at 24 h) relative to control, indicating impaired cognitive performance with continuous thermal stimulation; FUS-TCS (intermittent cycling) did not differ from control on these measures.

Exposures

Exposure 1: FUS-mediated thermal cycling stimulation (FUS-TCS) and continuous hyperthermia stimulation (FUS-HTS) to right cerebral hemisphere

Target: hemisphere unspecified — “right cerebral hemisphere”
Device: custom-built

Pulse timing
Waveform	pulsed, continuous
Fundamental frequency (kHz)	1,200	✓✓✓
Pulse duration (ms)	not reported
Pulse repetition frequency (Hz)	not reported
Duty cycle (%)	not reported
Sonication duration (s)	180, 1,800swept	✓✓✓

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 estimate	not reported
In-situ pressure (kPa)	not reported
In-situ Isppa (W/cm²)	not reported
In-situ Ispta (W/cm²)	not reported

Protocol, in the paper’s words

For the FUS-TCS procedure, the right cerebral hemispheres of the mice underwent a 10-cycle repeated treatment, where each cycle consisted of 3 min FUS heating activation followed by a 1 min rest interval. On the other hand, the FUS-HTS group received continuous FUS heating treatment on the right brain hemisphere for 30 min without interspersed rest intervals. Thermal treatments were administered on days 4, 8, and 12.

Flags from extraction

n_subjects — Paper does not state a single total number of animals exposed to real ultrasound; group sizes of n=8 are stated repeatedly in figure legends for FUS-TCS, FUS-HTS, Aβ+FUS-TCS groups (an Aβ+FUS-HTS group is also named in the Experimental schedule design section but not shown in main-text figures), so group sizes are listed rather than summed into an unstated total.
exposures[0].timing.waveform — The FUS-TCS 10-cycle on/off macro-structure (3 min heating / 1 min rest) and the FUS-HTS continuous 30-min heating are both described as thermal (hyperthermia) regimes at minute timescale, not conventional kHz-range pulsed neuromodulation; no PRF or duty cycle for a sub-second pulse structure is reported, so pulse_duration_ms, pulse_repetition_frequency_hz and duty_cycle_pct are left not_reported and the cycle structure is described in protocol_description instead.
sham_type — Control mice received an i.c.v. injection of PBS/saline but no FUS device or ultrasound procedure; classified as no_treatment_control (closest available term) though they did undergo an injection procedure.
direction_of_effect — This is a thermal/hyperthermia neuroprotection study assessing cognition and protein expression, not excitatory/inhibitory neurophysiology; classified as not_assessed.