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Abstract Spatial cognition depends on an accurate representation of orientation within an environment. Head direction cells in distributed brain regions receive a range of sensory inputs, but visual input is particularly important for aligning their responses to environmental landmarks. To investigate how population-level heading responses are aligned to visual input, we recorded from retrosplenial cortex (RSC) of head-fixed mice in a moving environment using two-photon calcium imaging. We show that RSC neurons are tuned to the animal’s relative orientation in the environment, even in the absence of head movement. Next, we found that RSC receives functionally distinct projections from visual and thalamic areas and contains several functional classes of neurons. While some functional classes mirror RSC inputs, a newly discovered class coregisters visual and thalamic signals. Finally, decoding analyses reveal unique contributions to heading from each class. Our results suggest an RSC circuit for anchoring heading representations to environmental visual landmarks. 

Using the structured illumination, single pixel detection imaging technique SPatIal Frequency modulation Imaging (SPIFI), we demonstrate a cascaded Wavelength Domain and Spatial Domain (WD-SD-SPIFI) system enabling real-time, in-line, second order dispersion compensation optimization for multiphoton imaging. Enhanced resolution is demonstrated by imaging a sub-diffractive 140 nm fluorescent nanodiamond with Two Photon Excitation Fluorescence (2PEF) to measure the Point Spread Function (PSF). With a 1034 nm pulsed laser through a Numerical Aperture (NA) of 0.5, a PSF Full Width at Half Max (FWHM) of 780 nm was measured with minimal post processing analysis that only requires Fast Fourier Transforms (FFTs). 

While two-photon fluorescence microscopy is a powerful platform for the study of functional dynamics in living cells and tissues, the bulk motion inherent to these applications causes distortions. We have designed a motion tracking module based on spectral domain optical coherence tomography which compliments a laser scanning two-photon microscope with real-time corrective feedback. The module can be added to fluorescent imaging microscopes using a single dichroic and without additional contrast agents. We demonstrate that the system can track lateral displacements as large as 10 μ m at 5 Hz with latency under 14 ms and propose a scheme to extend the system to 3D correction with the addition of a remote focusing module. We also propose several ways to improve the module’s performance by reducing the feedback latency. We anticipate that this design can be adapted to other imaging modalities, enabling the study of samples subject to motion artifacts at higher resolution. 

Imaging beyond the diffraction limit barrier has attracted wide attention due to the ability to resolve previously hidden image features. Of the various super-resolution microscopy techniques available, a particularly simple method called saturated excitation microscopy (SAX) requires only simple modification of a laser scanning microscope: The illumination beam power is sinusoidally modulated and driven into saturation. SAX images are extracted from the harmonics of the modulation frequency and exhibit improved spatial resolution. Unfortunately, this elegant strategy is hindered by the incursion of shot noise that prevents high-resolution imaging in many realistic scenarios. Here, we demonstrate a technique for super-resolution imaging that we call computational saturated absorption (CSA) in which a joint deconvolution is applied to a set of images with diversity in spatial frequency support among the point spread functions (PSFs) used in the image formation with saturated laser scanning fluorescence microscopy. CSA microscopy allows access to the high spatial frequency diversity in a set of saturated effective PSFs, while avoiding image degradation from shot noise. 

Spatial frequency modulation for imaging (SPIFI) has traditionally employed a time-varying spatial modulation of the excitation beam. Here, for the first time to our knowledge, we introduce single-shot SPIFI, where the spatial frequency modulation is imposed across the entire spatial bandwidth of the optical system simultaneously enabling single-shot operation. 

Sp at i al f requency modulation i maging (SPIFI) is a structured illumination single pixel imaging technique that is most often achieved via a rotating modulation disk. This implementation produces line images with exposure times on the order of tens of milliseconds. Here, we present a new architecture for SPIFI using a polygonal scan mirror with the following advances: (1) reducing SPIFI line image exposure times by 2 orders of magnitude, (2) facet-to-facet measurement and correction for polygonal scan design, and (3) a new anamorphic magnification scheme that improves resolution for long working distance optics. 

The hippocampus consists of a stereotyped neuronal circuit repeated along the septal-temporal axis. This transverse circuit contains distinct subfields with stereotyped connectivity that support crucial cognitive processes, including episodic and spatial memory. However, comprehensive measurements across the transverse hippocampal circuit in vivo are intractable with existing techniques. Here, we developed an approach for two-photon imaging of the transverse hippocampal plane in awake mice via implanted glass microperiscopes, allowing optical access to the major hippocampal subfields and to the dendritic arbor of pyramidal neurons. Using this approach, we tracked dendritic morphological dynamics on CA1 apical dendrites and characterized spine turnover. We then used calcium imaging to quantify the prevalence of place and speed cells across subfields. Finally, we measured the anatomical distribution of spatial information, finding a non-uniform distribution of spatial selectivity along the DG-to-CA1 axis. This approach extends the existing toolbox for structural and functional measurements of hippocampal circuitry.