Biology subjects

Natan, R.

Publications and source records attributed to Natan, R..

3 recordsLinked to original sources

bioRxiv2026

In vivo aberration measurement and correction for ultrafast FACED two-photon fluorescence microscopy of the brain

Ultrafast two-photon fluorescence microscopy (2PFM) based on free-space angular-chirp-enhanced delay (FACED) enables megahertz line scanning and kilohertz frame rates for in vivo brain imaging. However, optical aberrations from the imaging system and brain tissue degrade spatial resolution, signal, and contrast at depth. Here we integrate adaptive optics (AO) with FACED 2PFM to achieve synapse-resolving ultrafast imaging in the living mouse brain. Because FACED generates a one-dimensional array of temporally delayed, spatially separated excitation foci at 1 gigahertz, we developed a focus-averaging, frequency-multiplexed aberration measurement method that simultaneously measures and corrects the average aberration across all FACED foci using a segmented deformable mirror. We validated the accuracy of our method in correcting both system and artificial aberrations. When applied to in vivo morphological imaging of the mouse brain, AO enhances resolution, signal, contrast of dendritic shafts, spines, and boutons. Functionally, AO improves cerebral blood flow imaging by increasing plasma signal and kymograph contrast over large fields of view; when used for glutamate imaging, it amplifies transient amplitudes and reveals visually evoked glutamate release that were undetectable without correction. Together, these results establish AO-FACED 2PFM as a powerful approach that combines ultrafast imaging with high spatial resolution for structural and functional imaging in the living mouse brain.

neuroscience↗

bioRxiv2025

Parallel frequency-multiplexed aberration measurement for widefield fluorescence microscopy

Widefield fluorescence microscopy is widely used for imaging at subcellular resolution, but its performance in complex samples is degraded by optical aberrations. Because aberrations can vary spatially across the field of view (FOV), accurate aberration measurement and correction at multiple FOV locations are essential for achieving high-quality imaging over large areas. Here, we introduce parallel frequency-multiplexed aberration measurement (PFAM) to perform massively parallel aberration measurements across an extended FOV. We validated PFAM using fluorescent beads and demonstrated simultaneous measurement and effective correction of spatially varying aberrations at 125 FOV locations. To address the challenges of wavefront sensing in complex samples, we further developed PFAM-SIFT by integrating structured illumination, thereby achieving robust aberration measurement in both brain slices and the mouse brain in vivo. Together, PFAM and PFAM-SIFT provide accurate and scalable wavefront sensing solutions for widefield imaging, enabling simultaneous aberration measurement of spatially varying aberrations in complex biological samples.

biophysics↗

bioRxiv2024

Adaptive optical correction in in vivo two-photon fluorescence microscopy with neural fields

Adaptive optics (AO) restore ideal imaging performance in complex samples by measuring and correcting optical aberrations, but often require custom-built microscopes with carefully aligned wavefront sensing/shaping devices and can be susceptible to sample motion. Here we describe NeAT, a computational framework using neural fields for AO two-photon fluorescence microscopy. NeAT estimates wavefront aberration and recovers sample structure from a 3D image stack without requiring external datasets for training. Incorporating motion correction in learning and correcting conjugation errors commonly found in commercial microscopes, NeAT is designed for deployment in biological laboratories for in vivo imaging. We validate NeATs performance using a custom-built microscope with a wavefront sensor under varying signal-to-noise ratios, aberration, and motion conditions. With a commercial microscope, we demonstrate real-time aberration correction for in vivo morphological and functional imaging in the living mouse brain, with NeAT improving signal and accuracy of glutamate and calcium imaging of synapses and neurons.

neuroscience↗