Farnaz Sharif

Reduced-phase dual-illumination interferometer for measuring large stepped objects

We present a reduced-phase dual-illumination interferometer (RPDII) that measures the topography of a sample with large step height variation. We experimentally demonstrate the basic principle and the feasibility of this novel single-shot quantitative phase imaging. Two beams of this interferometer illuminate a sample at different incident angles, and two phases of the different incident angles and their phase difference are simultaneously recorded using three spatial frequencies. The relative phase difference between two beams of an RPDII can be controlled by adjusting the angle such that the maximum phase difference is smaller than 2π, and thus there is no phase wrapping ambiguity in the reconstructed phase. One 4f optical system with a transmission grating is used to illuminate the sample with two collimated beams incident at different angles. The feasibility of this technique is demonstrated by measuring the thicknesses of two stepped metal layers with heights of 150 and 660 μm. Although the change in stepped height is more than 1000 times the wavelength of the laser used in our interferometer, the thicknesses of these two metal layers are successfully obtained without the use of an unwrapping algorithm.

Large step-phase measurement by a reduced-phase triple-illumination interferometer.

We present a reduced-phase triple-illumination interferometer (RPTII) as a novel single-shot technique to increase the precision of dual-illumination optical phase unwrapping techniques. The technique employs two measurement ranges to record both low-precision unwrapped and high-precision wrapped phases. To unwrap the high-precision phase, a hierarchical optical phase unwrapping algorithm is used with the low-precision unwrapped phase. The feasibility of this technique is demonstrated by measuring a stepped object with a height 2100 times greater than the wavelength of the source. The phase is reconstructed without applying any numerical unwrapping algorithms, and its noise level is decreased by a factor of ten.

Double-field-of-view, quasi-common-path interferometer using Fourier domain multiplexing

We present a quasi-common-path interferometer with a double field of view (FOV). The laser beam of an imaging system is separated into three parts using three mirrors; the first and second beams are used to image two different areas of a sample, while the third beam functions as a reference beam. The reference beam is prepared by making clear area in a sample and projecting it on an image sensor. A double FOV is obtained by Fourier domain multiplexing, whereby two interferometric images corresponding to two different areas of a sample are modulated with two different spatial carrier frequencies. The feasibility of this technique is experimentally demonstrated by imaging two different areas of a test target with a single image sensor.

Transparent stepped phase measurement using two illuminating beams

We propose a single shot and single wavelength phase imaging technique for measuring phase of the transparent objects without using unwrapping process. A grating between a laser and the object is used to make beams with different angle, which determines the measurement range of the microscope. The grating pitch and magnification of the lens system before the sample affect the angle. The angle inside the object is changed according to Snell’s law; therefore, final angle is related to the refractive index of the object. Magnification of the lens system after sample will control the modulation frequency of microscope. The interference pattern is constructed at CCD plane and convey information of the sample. For a phase below the measurement range of the microscope, the reconstructed phase is not wrapped. By increasing the measurement range accuracy of the system will drop; therefore the magnification of the lenses must choose carefully to obtain optimal phase. The ability of this technique is demonstrated by reconstructing phases of two transparent step objects with 150 and 510 μm height. Their refractive indexes for red light are 1.515 and 1.508 , respectively. Therefore, total optical path length difference is 336 micrometers that is 500 times more than the laser wavelength. The phase is successfully reconstructed without using unwrapping algorithms.

Compact single-shot four-wavelength quantitative phase microscopy with polarization- and frequency-division demultiplexing

We present a novel single-shot four-wavelength quantitative phase microscopy (FW-QPM). Four lasers operating at different wavelengths are multiplexed with a pair of dichroic mirrors and a polarization beam splitter in a three-mirror quasi-common-path interferometer. After a single-shot interference pattern is obtained with a monochrome camera, four holograms of different wavelengths were demultiplexed from it in the frequency domain with polarization- and frequency-division multiplexing. Polarization-division demultiplexing scheme uses polarization dependent visibility changes in an interference pattern, and it plays a critical role in making only two interference patterns exist within a single quadrant in the frequency domain. We have used a single-mode optical fiber as a phase object sample and demonstrated that a measured single-shot interference pattern can be successfully demultiplexed into four different interferograms of different wavelengths with our proposed scheme.

Micro-drive and headgear for chronic implant and recovery of optoelectronic probes

Silicon probes are multisite electrodes used for the electrophysiological recording of large neuronal ensembles. Optoelectronic probes (OEPs) are recent upgrades that allow, in parallel, the delivery of local optical stimuli. The procedures to use these delicate electrodes for chronic experiments in mice are still underdeveloped and typically assume one-time uses. Here, we developed a micro-drive, a support for OEPs optical fibers, and a hat enclosure, which fabrications consist in fitting and fastening together plastic parts made with 3D printers. Excluding two parts, all components and electrodes are relatively simple to recover after the experiments, via the loosening of screws. To prevent the plugging of OEPs laser sources from altering the stability of recordings, the OEPs fibers can be transiently anchored to the hat via the tightening of screws. We test the stability of recordings in the mouse hippocampus under three different conditions: acute head-fixed, chronic head-fixed, and chronic freely moving. Drift in spike waveforms is significantly smaller in chronic compared to acute conditions, with the plugging/unplugging of head-stage and fiber connectors not affecting much the recording stability. Overall, these tools generate stable recordings of place cell in chronic conditions, and make the recovery and reuse of electrode packages relatively simple.

Optical unwrapping by triple illumination interferometer

Here, we present a triple illumination phase interferometer to have a more flexible unwrapping measurement range for single shot optical unwrapping. Three beams of this interferometer illuminate a sample at different incident angles, three phases of the different incident angles are simultaneously recorded using three spatial frequencies. The different direction phases can be used for dual illumination optical unwrapping; as a result, the phase can be unwrapped by more than one measurement range. The feasibility of this technique is demonstrated by measuring a stepped object with heights of 150 and 660 μm. The smaller stepped phase is unwrapped by two measurement range; however, the phase of larger step is wrapped for only one of the measurement range.

Effect of the angle accuracy in reduced-phase dual-illumination interferometer

Reduced-phase dual-illumination interferometer (RPDII) is an off axis, single shot and single wavelength phase imaging technique to measure large objects without using unwrapping algorithms. Two beams of this interferometer illuminate a sample at different incident angles, two phases of the different incident angles and their phase difference are recorded. The phase difference between two beams can be controlled by adjusting the incident angles. The angle accuracy that decrease the RPDII accuracy have been studied. We have shown, the groove spacing of the grating and magnification of the lens system before sample, determine the incident angle accuracy. The ability of RPDII to unwrap large phases is shown by reconstructing phase of a step object without using unwrapping algorithms. The reconstructed image shows that the total inaccuracy is much more than the inaccuracy caused by incident angles.

Differential representation of landmark and self-motion information along the CA1 radial axis: self-motion generated place fields shift toward landmarks during septal inactivation

Spatial location in the environment can be defined in relation to specific landmarks or in relation to the global context, and is estimated from both the sensing of landmarks and the inner sense of cumulated locomotion referred to as path-integration. The respective contribution of landmark and path-integration to place-cell activity in the hippocampus is still unclear and complicated by the fact that the two mechanisms usually overlap. To bias spatial mechanisms toward landmark or path-integration, we use a treadmill equipped with a long belt on which male mice run sequentially through a zone enriched and a zone impoverished in visual-tactile cues. We show that inactivation of the medial septum (MS), which is known to disrupt the periodic activity of grid cells, impairs mice ability to anticipate the delivery of a reward in the cue-impoverished zone and transiently alter the spatial configuration of place fields in the cue-impoverished zone selectively: following MS inactivation, place fields in the cue-impoverished zone progressively shift backward and stabilize near the cues, resulting in the contraction of the spatial representation around cues; following MS recovery, the initial spatial representation is progressively restored. Furthermore, we found that place fields in the cue-rich and cue-impoverished zones are preferentially generated by cells from the deep and superficial sublayers of CA1, respectively. These findings demonstrate with mechanistic insights the contribution of MS to the spread of spatial representations in cue-impoverished zones, and indicate a segregation of landmark-based and path-integration-assisted spatial mechanisms into deep and superficial CA1, respectively. SIGNIFICANCE STATEMENT Cells encoding a cue-impoverished zone and the vicinity of landmarks responded differentially to septal inactivation and resided in distinct sublayers of CA1. These findings provide new insights on place field mechanisms: septal activity is critical for maintaining the spread of place fields in cue-impoverished areas, but not for the generation of place fields; Following MS inactivation, trial-by-trial network modifications by activity-dependent mechanisms are responsible for the gradual collapse of spatial representations. Furthermore, the findings suggest parallel coding streams for landmark and self-motion information. Superficial CA1 cells are better suited for encoding global position via the assist of path-integration, whereas deep CA1 cells can support spatial memory processes on an object-specific basis.

Implantation of Chronic Silicon Probes and Recording of Hippocampal Place Cells in an Enriched Treadmill Apparatus

An important requisite for understanding brain function is the identification of behavior and cell activity correlates. Silicon probes are advanced electrodes for large-scale electrophysiological recording of neuronal activity, but the procedures for their chronic implantation are still underdeveloped. The activity of hippocampal place cells is known to correlate with an animals position in the environment, but the underlying mechanisms are still unclear. To investigate place cells, here we describe a set of techniques which range from the fabrication of devices for chronic silicon probe implants to the monitoring of place field activity in a cue-enriched treadmill apparatus. A micro-drive and a hat are built by fitting and fastening together 3D-printed plastic parts. A silicon probe is mounted on the micro-drive, cleaned, and coated with dye. A first surgery is performed to fix the hat on the skull of a mouse. Small landmarks are fabricated and attached to the belt of a treadmill. The mouse is trained to run head-fixed on the treadmill. A second surgery is performed to implant the silicon probe in the hippocampus, following which broadband electrophysiological signals are recorded. Finally, the silicon probe is recovered and cleaned for reuse. The analysis of place cell activity in the treadmill reveals a diversity of place field mechanisms, outlining the benefit of the approach.