Super-resolution imaging of brain structures at large depths
Super-resolution microscopy now enables to visualize fine neuronal structures, such as e.g. axons and dendritic spines, at unprecedent resolution. Visualization of the shape of dendritic spines at significant depths is key to decipher synaptic computation and better understand synaptic plasticity.
The development of advanced multiphoton microscopy techniques, such as two-photon multifocal structured illumination microscopy (2P-MSIM), now enables deep imaging in biological samples beyond the diffraction limit, in particular in scattering tissue such as brain.
However, accessing such visualization at large depths is impaired by optical aberrations arising from tissue inhomogeneity, which limits both achievable image contrast and depth.
Adaptive Optics (AO) for improved image contrast, resolution and depth in 2P-MSIM
In their paper recently published in Photonix, scientists from research groups Shenzhen University implemented AO at both excitation and detection paths. To enable wavefront (WF) sensing, a non-linear guide star was generated using a Spatial Light Modulator (SLM), the corresponding fluorescence signal from a tissue structure such as e.g. a neuron body was diverted to a Shack-Hartmann wavefront sensor.
Based on the WF measurement, both the multifocal excitation pattern generated by the SLM and the shape of a Deformable Mirror (DM) in the fluorescence imaging path were adjusted to compensate for measured aberrations. When applied e.g. to deep imaging of dendritic structures in fixed mouse brain slices, the approach enabled a x5 increase in fluorescence signal a significant depths >300 µm, with a maintained 150nm resolution.
Key components for AO implementation
AO was implemented based on selected components from our AOKit BIO, namely Mirao 52e as the DM and as a Shack-Hartmann WF sensor. This combination, together with our software solutions, provide an efficient combination of WF sensing accuracy and WF modulation linearity and stroke. Alternatives providing even better linearity, more actuators, or an integrated solution such as an AO module can be found in our product range.