Optical properties of the mouse eye

The Shack-Hartmann wavefront sensor (SHWS) spots upon which ocular aberration measurements depend have poor quality in mice due to light reflected from multiple retinal layers. We have designed and implemented a SHWS that can favor light from a specific retinal layer and measured monochromatic aberrations in 20 eyes from 10 anesthetized C57BL/6J mice. Using this instrument, we show that mice are myopic, not hyperopic as is frequently reported. We have also measured longitudinal chromatic aberration (LCA) of the mouse eye and found that it follows predictions of the water-filled schematic mouse eye. Results indicate that the optical quality of the mouse eye assessed by measurement of its aberrations is remarkably good, better for retinal imaging than the human eye. The dilated mouse eye has a much larger numerical aperture (NA) than that of the dilated human eye (0.5 NA vs. 0.2 NA), but it has a similar amount of root mean square (RMS) higher order aberrations compared to the dilated human eye. These measurements predict that adaptive optics based on this method of wavefront sensing will provide improvements in retinal image quality and potentially two times higher lateral resolution than that in the human eye.     

Contrast-based sensorless adaptive optics for retinal imaging

Conventional adaptive optics ophthalmoscopes use wavefront sensing methods to characterize ocular aberrations for real-time correction. However, there are important situations in which the wavefront sensing step is susceptible to difficulties that affect the accuracy of the correction. To circumvent these, wavefront sensorless adaptive optics (or non-wavefront sensing AO; NS-AO) imaging has recently been developed and has been applied to point-scanning based retinal imaging modalities. In this study we show, for the first time, contrast-based NS-AO ophthalmoscopy for full-frame in vivo imaging of human and animal eyes. We suggest a robust image quality metric that could be used for any imaging modality, and test its performance against other metrics using (physical) model eyes.OCIS codes: (010.1080) Active or adaptive optics, (170.3880) Medical and biological imaging, (170.4460) Ophthalmic optics and devices, (330.4875) Optics of physiological systems     

Imaging cone photoreceptors in three dimensions and in time using ultrahigh resolution optical coherence tomography with adaptive optics

Cone photoreceptors in the living human eye have recently been imaged with micron-scale resolution in all three spatial dimensions using adaptive optics optical coherence tomography. While these advances have allowed non-invasive study of the three-dimensional structure of living human cones, studies of their function and physiology are still hampered by the difficulties to monitor the same cells over time. The purpose of this study is to demonstrate the feasibility of cone monitoring using ultrahigh-resolution adaptive optics optical coherence tomography. Critical to this is incorporation of a high speed CMOS camera (125 KHz) and a novel feature-based, image registration/dewarping algorithm for reducing the deleterious effects of eye motion on volume images. Volume movies were acquired on three healthy subjects at retinal eccentricities from 0.5° to 6°. Image registration/dewarping reduced motion artifacts in the movies from 15 μm to 1.3 μm root mean square, the latter sufficient for identifying and tracking cones. Cone row-to-row spacing and outer segment lengths were consistent with that reported in the literature. Cone length analysis demonstrates that UHR-AO-OCT is sufficiently sensitive to measure real length differences between cones in the same 0.5° retinal patch, and requires no more than five measurements of OS length to achieve 95% confidence. We know of no other imaging modality that can monitor foveal or parafoveal cones over time with comparable resolution in all three dimensions.     

Improving wavefront boundary condition for in vivo high resolution adaptive optics ophthalmic imaging

An ophthalmic adaptive optics (AO) imaging system is especially affected by pupil edge effects due to the higher noise and aberration level at the edge of the human pupil as well as the impact of head and eye motions on the pupil. In this paper, a two-step approach was proposed and implemented for reducing the edge effects and improving wavefront slope boundary condition. First, given an imaging pupil, a smaller size of sampling aperture can be adopted to avoid the noisy boundary slope data. To do this, we calibrated a set of influence matrices for different aperture sizes to accommodate pupil variations within the population. In step two, the slope data was extrapolated from the less noisy slope data inside the pupil towards the outside such that we had reasonable slope data over a larger aperture to stabilize the impact of eye pupil dynamics. This technique is applicable to any Neumann boundary-based active /adaptive modality but it is especially useful in the eye for improving AO retinal image quality where the boundary positions fluctuate.     

Retinal adaptive optics imaging with a pyramid wavefront sensor

The pyramid wavefront sensor (P-WFS) has replaced the Shack-Hartmann (SH-) WFS as the sensor of choice for high-performance adaptive optics (AO) systems in astronomy. Many advantages of the P-WFS, such as its adjustable pupil sampling and superior sensitivity, are potentially of great benefit for AO-supported imaging in ophthalmology as well. However, so far no high quality ophthalmic AO imaging was achieved using this novel sensor. Usually, a P-WFS requires modulation and high precision optics that lead to high complexity and costs of the sensor. These factors limit the competitiveness of the P-WFS with respect to other WFS devices for AO correction in visual science. Here, we present a cost-effective realization of AO correction with a non-modulated P-WFS based on standard components and apply this technique to human retinal in vivo imaging using optical coherence tomography (OCT). P-WFS based high quality AO imaging was successfully performed in 5 healthy subjects and smallest retinal cells such as central foveal cone photoreceptors are visualized. The robustness and versatility of the sensor is demonstrated in the model eye under various conditions and in vivo by high-resolution imaging of other structures in the retina using standard and extended fields of view. As a quality benchmark, the performance of conventional SH-WFS based AO was used and successfully met. This work may trigger a paradigm shift with respect to the wavefront sensor of choice for AO in ophthalmic imaging.     

Woofer-tweeter adaptive optics scanning laser ophthalmoscopic imaging based on Lagrange-multiplier damped least-squares algorithm

We implemented a Lagrange-multiplier (LM)-based damped least-squares (DLS) control algorithm in a woofer-tweeter dual deformable-mirror (DM) adaptive optics scanning laser ophthalmoscope (AOSLO). The algorithm uses data from a single Shack-Hartmann wavefront sensor to simultaneously correct large-amplitude low-order aberrations by a woofer DM and small-amplitude higher-order aberrations by a tweeter DM. We measured the in vivo performance of high resolution retinal imaging with the dual DM AOSLO. We compared the simultaneous LM-based DLS dual DM controller with both single DM controller, and a successive dual DM controller. We evaluated performance using both wavefront (RMS) and image quality metrics including brightness and power spectrum. The simultaneous LM-based dual DM AO can consistently provide near diffraction-limited in vivo routine imaging of human retina.     

Non-common path aberration correction in an adaptive optics scanning ophthalmoscope

The correction of non-common path aberrations (NCPAs) between the imaging and wavefront sensing channel in a confocal scanning adaptive optics ophthalmoscope is demonstrated. NCPA correction is achieved by maximizing an image sharpness metric while the confocal detection aperture is temporarily removed, effectively minimizing the monochromatic aberrations in the illumination path of the imaging channel. Comparison of NCPA estimated using zonal and modal orthogonal wavefront corrector bases provided wavefronts that differ by ~λ/20 in root-mean-squared (~λ/30 standard deviation). Sequential insertion of a cylindrical lens in the illumination and light collection paths of the imaging channel was used to compare image resolution after changing the wavefront correction to maximize image sharpness and intensity metrics. Finally, the NCPA correction was incorporated into the closed-loop adaptive optics control by biasing the wavefront sensor signals without reducing its bandwidth.OCIS codes: (110.1080) Active or adaptive optics, (170.4460) Ophthalmic optics and devices     

Wavefront sensorless adaptive optics optical coherence tomography for in vivo retinal imaging in mice

We present wavefront sensorless adaptive optics (WSAO) Fourier domain optical coherence tomography (FD-OCT) for in vivo small animal retinal imaging. WSAO is attractive especially for mouse retinal imaging because it simplifies optical design and eliminates the need for wavefront sensing, which is difficult in the small animal eye. GPU accelerated processing of the OCT data permitted real-time extraction of image quality metrics (intensity) for arbitrarily selected retinal layers to be optimized. Modal control of a commercially available segmented deformable mirror (IrisAO Inc.) provided rapid convergence using a sequential search algorithm. Image quality improvements with WSAO OCT are presented for both pigmented and albino mouse retinal data, acquired in vivo.OCIS codes: (170.4460) Ophthalmic optics and devices, (110.1080) Active or adaptive optics, (110.4500) Optical coherence tomography     

Pupil tracking optical coherence tomography for precise control of pupil entry position

To maximize the collection efficiency of back-scattered light, and to minimize aberrations and vignetting, the lateral position of the scan pivot of an optical coherence tomography (OCT) retinal scanner should be imaged to the center of the ocular pupil. Additionally, several retinal structures including Henle’s Fiber Layer (HFL) exhibit reflectivities that depend on illumination angle, which can be controlled by varying the pupil entry position of the OCT beam. In this work, we describe an automated method for controlling the lateral pupil entry position in retinal OCT by utilizing pupil tracking in conjunction with a 2D fast steering mirror placed conjugate to the retinal plane. We demonstrate that pupil tracking prevents lateral motion artifacts from impeding desired pupil entry locations, and enables precise pupil entry positioning and therefore control of the illumination angle of incidence at the retinal plane. We use our prototype pupil tracking OCT system to directly visualize the obliquely oriented HFL.OCIS codes: (110.4500) Optical coherence tomography, (170.4460) Ophthalmic optics and devices     

Understanding the changes of cone reflectance in adaptive optics flood illumination retinal images over three years

Although there is increasing interest in the investigation of cone reflectance variability, little is understood about its characteristics over long time scales. Cone detection and its automation is now becoming a fundamental step in the assessment and monitoring of the health of the retina and in the understanding of the photoreceptor physiology. In this work we provide an insight into the cone reflectance variability over time scales ranging from minutes to three years on the same eye, and for large areas of the retina (≥ 2.0 × 2.0 degrees) at two different retinal eccentricities using a commercial adaptive optics (AO) flood illumination retinal camera. We observed that the difference in reflectance observed in the cones increases with the time separation between the data acquisitions and this may have a negative impact on algorithms attempting to track cones over time. In addition, we determined that displacements of the light source within 0.35 mm of the pupil center, which is the farthest location from the pupil center used by operators of the AO camera to acquire high-quality images of the cone mosaic in clinical studies, does not significantly affect the cone detection and density estimation.OCIS codes: (110.1080) Active or adaptive optics, (330.7331) Visual optics, receptor optics, (170.3880) Medical and biological imaging     

Imaging of retinal vasculature using adaptive optics SLO/OCT

We use our previously developed adaptive optics (AO) scanning laser ophthalmoscope (SLO)/ optical coherence tomography (OCT) instrument to investigate its capability for imaging retinal vasculature. The system records SLO and OCT images simultaneously with a pixel to pixel correspondence which allows a direct comparison between those imaging modalities. Different field of views ranging from 0.8°x0.8° up to 4°x4° are supported by the instrument. In addition a dynamic focus scheme was developed for the AO-SLO/OCT system in order to maintain the high transverse resolution throughout imaging depth. The active axial eye tracking that is implemented in the OCT channel allows time resolved measurements of the retinal vasculature in the en-face imaging plane. Vessel walls and structures that we believe correspond to individual erythrocytes could be visualized with the system.OCIS codes: (170.3890) Medical optics instrumentation, (110.1080) Active or adaptive optics, (170.4470) Ophthalmology, (330.5310) Vision - photoreceptors, (110.4500) Optical coherence tomography     

In vivo imaging of human photoreceptor mosaic with wavefront sensorless adaptive optics optical coherence tomography

Wavefront sensorless adaptive optics optical coherence tomography (WSAO-OCT) is a novel imaging technique for in vivo high-resolution depth-resolved imaging that mitigates some of the challenges encountered with the use of sensor-based adaptive optics designs. This technique replaces the Hartmann Shack wavefront sensor used to measure aberrations with a depth-resolved image-driven optimization algorithm, with the metric based on the OCT volumes acquired in real-time. The custom-built ultrahigh-speed GPU processing platform and fast modal optimization algorithm presented in this paper was essential in enabling real-time, in vivo imaging of human retinas with wavefront sensorless AO correction. WSAO-OCT is especially advantageous for developing a clinical high-resolution retinal imaging system as it enables the use of a compact, low-cost and robust lens-based adaptive optics design. In this report, we describe our WSAO-OCT system for imaging the human photoreceptor mosaic in vivo. We validated our system performance by imaging the retina at several eccentricities, and demonstrated the improvement in photoreceptor visibility with WSAO compensation.OCIS codes: (110.4500) Optical coherence tomography, (010.1080) Active or adaptive optics, (220.1000) Aberration compensation, (170.0110) Imaging systems, (170.4470) Ophthalmology, (120.3890) Medical optics instrumentation     

Longitudinal chromatic aberration of the human eye in the visible and near infrared from wavefront sensing,double-pass and psychophysics

Longitudinal Chromatic Aberration (LCA) influences the optical quality of the eye. However, the reported LCA varies across studies, likely associated to differences in the measurement techniques. We present LCA measured in subjects using wavefront sensing, double-pass retinal images, and psychophysical methods with a custom-developed polychromatic Adaptive Optics system in a wide spectral range (450-950 nm), with control of subjects’ natural aberrations. LCA measured psychophysically was significantly higher than that from reflectometric techniques (1.51 D vs 1.00 D in the 488-700 nm range). Ours results indicate that the presence of natural aberrations is not the cause for the discrepancies across techniques.OCIS codes: (260.0260) Physical optics, (330.0330) Vision, color, and visual optics, (330.4875) Optics of physiological systems, (330.5370) Physiological optics, (220.1010) Aberrations (global), (220.1080) Active or adaptive optics     

Wavefront sensor-less adaptive optics using deep reinforcement learning

Image degradation due to wavefront aberrations can be corrected with adaptive optics (AO). In a typical AO configuration, the aberrations are measured directly using a Shack-Hartmann wavefront sensor and corrected with a deformable mirror in order to attain diffraction limited performance for the main imaging system. Wavefront sensor-less adaptive optics (SAO) uses the image information directly to determine the aberrations and provide guidance for shaping the deformable mirror, often iteratively. In this report, we present a Deep Reinforcement Learning (DRL) approach for SAO correction using a custom-built fluorescence confocal scanning laser microscope. The experimental results demonstrate the improved performance of the DRL approach relative to a Zernike Mode Hill Climbing algorithm for SAO.     

Measurement and correction of transverse chromatic offsets for multi-wavelength retinal microscopy in the living eye

A special challenge arises when pursuing multi-wavelength imaging of retinal tissue in vivo, because the eye’s optics must be used as the main focusing elements, and they introduce significant chromatic dispersion. Here we present an image-based method to measure and correct for the eye’s transverse chromatic aberrations rapidly, non-invasively, and with high precision. We validate the technique against hyperacute psychophysical performance and the standard chromatic human eye model. In vivo correction of chromatic dispersion will enable confocal multi-wavelength images of the living retina to be aligned, and allow targeted chromatic stimulation of the photoreceptor mosaic to be performed accurately with sub-cellular resolution.OCIS codes: (110.1080) Active or adaptive optics, (130.2035) Dispersion compensation devices, (170.5810) Scanning microscopy, (330.5510) Psychophysics, (330.7327) Visual optics, ophthalmic instrumentation     

Digital adaptive optics based on digital lateral shearing of the computed pupil field for point scanning retinal swept source OCT

A novel non-iterative digital adaptive optics technique is presented in which the wavefront error is calculated using the phase difference between the pupil field and its digital copies translated by a pixel along the horizontal and vertical direction in the pupil plane. This method provides slope data per pixel, thus can generate > 50k local slope data samples for a circular pupil of diameter 256 pixels with high accuracy and dynamic range. It offers more than 12x faster computational speed in comparison to the sub-aperture based digital adaptive optics method. Furthermore, it is independent of any system parameters, the light distribution in the pupil plane, or the intensity of the image. The technique is useful in applications such as interferometric or digital holography based microscopy, metrology, and as digital wavefront sensor in adaptive optics, where focusing of light in the sample is involved that creates a guide star or where the sample itself exhibits guide star-like structures. This technique is implemented in a point scanning swept-source OCT at 1060 nm, and a large wavefront error with a peak to valley of 20 radians and root mean square error of 0.71 waves is detected and corrected in case of a micro-beads phantom sample. Also, human photoreceptor images are recovered from aberrated retinal OCT volumes acquired at eccentricities of 2 and 2.5 degrees from the fovea in vivo.     

Adaptive optics retinal imaging in the living mouse eye

Correction of the eye's monochromatic aberrations using adaptive optics (AO) can improve the resolution of in vivo mouse retinal images [Biss et al., Opt. Lett. 32(6), 659 (2007) and Alt et al., Proc. SPIE 7550, 755019 (2010)], but previous attempts have been limited by poor spot quality in the Shack-Hartmann wavefront sensor (SHWS). Recent advances in mouse eye wavefront sensing using an adjustable focus beacon with an annular beam profile have improved the wavefront sensor spot quality [Geng et al., Biomed. Opt. Express 2(4), 717 (2011)], and we have incorporated them into a fluorescence adaptive optics scanning laser ophthalmoscope (AOSLO). The performance of the instrument was tested on the living mouse eye, and images of multiple retinal structures, including the photoreceptor mosaic, nerve fiber bundles, fine capillaries and fluorescently labeled ganglion cells were obtained. The in vivo transverse and axial resolutions of the fluorescence channel of the AOSLO were estimated from the full width half maximum (FWHM) of the line and point spread functions (LSF and PSF), and were found to be better than 0.79 μm ± 0.03 μm (STD)(45% wider than the diffraction limit) and 10.8 μm ± 0.7 μm (STD)(two times the diffraction limit), respectively. The axial positional accuracy was estimated to be 0.36 μm. This resolution and positional accuracy has allowed us to classify many ganglion cell types, such as bistratified ganglion cells, in vivo.     

Closed-loop wavefront sensing and correction in the mouse brain with computed optical coherence microscopy

Optical coherence microscopy (OCM) uses interferometric detection to capture the complex optical field with high sensitivity, which enables computational wavefront retrieval using back-scattered light from the sample. Compared to a conventional wavefront sensor, aberration sensing with OCM via computational adaptive optics (CAO) leverages coherence and confocal gating to obtain signals from the focus with less cross-talk from other depths or transverse locations within the field-of-view. Here, we present an investigation of the performance of CAO-based aberration sensing in simulation, bead phantoms, and ex vivo mouse brain tissue. We demonstrate that, due to the influence of the double-pass confocal OCM imaging geometry on the shape of computed pupil functions, computational sensing of high-order aberrations can suffer from signal attenuation in certain spatial-frequency bands and shape similarity with lower order counterparts. However, by sensing and correcting only low-order aberrations (astigmatism, coma, and trefoil), we still successfully corrected tissue-induced aberrations, leading to 3× increase in OCM signal intensity at a depth of ∼0.9 mm in a freshly dissected ex vivo mouse brain.     

Investigation of the isoplanatic patch and wavefront aberration along the pupillary axis compared to the line of sight in the eye

Conventional optical systems usually provide best image quality on axis, while showing unavoidable gradual decrease in image quality towards the periphery of the field. The optical system of the human eye is not an exception. Within a limiting boundary the image quality can be considered invariant with field angle, and this region is known as the isoplanatic patch. We investigate the isoplanatic patch of eight healthy eyes and measure the wavefront aberration along the pupillary axis compared to the line of sight. The results are used to discuss methods of ocular aberration correction in wide-field retinal imaging with particular application to multi-conjugate adaptive optics systems.     

Adaptive optics optical coherence tomography with dynamic retinal tracking

Adaptive optics optical coherence tomography (AO-OCT) is a highly sensitive and noninvasive method for three dimensional imaging of the microscopic retina. Like all in vivo retinal imaging techniques, however, it suffers the effects of involuntary eye movements that occur even under normal fixation. In this study we investigated dynamic retinal tracking to measure and correct eye motion at KHz rates for AO-OCT imaging. A customized retina tracking module was integrated into the sample arm of the 2nd-generation Indiana AO-OCT system and images were acquired on three subjects. Analyses were developed based on temporal amplitude and spatial power spectra in conjunction with strip-wise registration to independently measure AO-OCT tracking performance. After optimization of the tracker parameters, the system was found to correct eye movements up to 100 Hz and reduce residual motion to 10 µm root mean square. Between session precision was 33 µm. Performance was limited by tracker-generated noise at high temporal frequencies.OCIS codes: (110.1080) Active or adaptive optics, (170.4500) Optical coherence tomography, (120.3890) Medical optics instrumentation, (170.0110) Imaging systems, (170.4470) Ophthalmology, (330.5310) Vision - photoreceptors