Technical development of two-photon optogenetic stimulation and its potential application to brain–machine interfaces

Over the past decade, techniques enabling bidirectional modulation of neuronal activity with single-cell precision have rapidly advanced in the form of two-photon optogenetic stimulation. Unlike conventional electrophysiological approaches or one-photon optogenetics, which inevitably activate many neurons surrounding the target, two-photon optogenetics can drive hundreds of specifically targeted neurons simultaneously, with stimulation patterns that can be flexibly and rapidly reconfigured. In this review, we trace the development of two-photon optogenetic stimulation, focusing on its progression toward implementations in large field of view two-photon microscopes capable of targeted multi-neuron control. We highlight three principal strategies: spiral scanning, temporal focusing, and three-dimensional computer-generated holography, along with their combinations, which together provide powerful tools for causal interrogation of neural circuits and behavior. Finally, we discuss the integration of these optical technologies into brain-machine interfaces, emphasizing both their transformative potential and the technical challenges that must be addressed to realize their broader impact.