• • Aβ42 (1 μM, 48 h) significantly decreased cell viability on stiff substrates (P < 0.05) but not on soft substrates, indicating that ECM stiffness above a threshold exacerbates Aβ42 toxicity; this has clinical implications for AD patients with altered brain ECM mechanics.
• • The percentage of neurons with spontaneous Ca2+ oscillations was significantly greater on stiff substrates than on soft substrates following Aβ42 exposure (P < 0.05), suggesting that stiff ECM promotes aberrant neuronal excitability, a potential early biomarker for AD progression.
• • Aβ42 exposure reduced synaptic formation in a substrate stiffness-dependent manner (P < 0.05), with stiff substrates showing greater loss, highlighting the role of mechanical cues in synaptic integrity and cognitive decline.
• • Electrophysiological analysis of 58–62 neurons revealed that Aβ42 altered sEPSC and sIPSC amplitudes and frequencies differentially on soft versus stiff substrates (P < 0.05), demonstrating that ECM stiffness modulates both excitatory and inhibitory synaptic transmission, which may underlie network dysfunction in AD.