Multi-target transcranial alternating current stimulation (tACS) enhances motor learning and brain network connection in middle-aged adults

Background 40 Hz transcranial alternating current stimulation (tACS) enhances motor learning, but single-target effects and mechanisms remain unclear. We proposed multi-target tACS to improve efficacy. Methods Twenty-five healthy adults (>45 years) were randomized into sham (A), single-target (B), double-target (C), and multi-target (D) tACS groups. Outcomes included sequence reaction time task (SRTT), transcranial magnetic stimulation (TMS), MRI (gray matter density, activation and functional connectivity (FC)), and RNA sequencing. Results SRTT showed that group D significantly shortened the reaction time and error rate compared to baseline. TMS results indicate increased cortical excitability before and after tACS intervention, but no significant difference exists. MRI results showed that the gray matter density in the right middle frontal gyrus (MFG), including the dorsolateral prefrontal cortex (DLPFC) of group D, significantly increased. The activation value of group D in the frontal lobe (left) and cerebellum (left) is substantially higher than that of the other three groups. The functional connection (FC) of motor-cognitive-related brain networks, including primary motor cortex (M1) and frontal lobe and supplementary motor area (SMA), was significantly improved in group D. RNA sequencing analysis revealed a significant increase in oxygen metabolism of group D when compared to group C. Conclusion Multi-target tACS enhances motor learning, likely by activating left frontal and cerebellar regions, strengthening M1-frontal-SMA connectivity, and boosting oxygen metabolism.