Frequency domain speckle tracking for OCT based 3D vibrometry of the inner ear - The increasing prevalence of hearing loss is driving the demand for improved techniques to study the intricate structure and complex processes of the living cochlea. Optical coherence tomography (OCT) has become a widely used technology to study the vibrometry of the living cochlea due to its temporal and spatial resolution. One limitation with OCT is that it only measures the vibratory response of the cochlea along its optical axis, providing measurements in just one dimension. Given that the cochlea has a 3D structure with subcellular structures that move in all 3 dimensions when stimulated, traditional OCT does not capture important information about the full range of movement of the cochlea. To address this limitation with OCT, our lab has built a 3-beam OCT and vibrometry system that has three sample arms combined at a single interferometer. With three projections of data collected from three different angles, we can reconstruct the 3D vector of motion when the living cochlea is excited. We have used this system to collect data from the apical region of the mouse cochlea and study the 3D motion for different stimuli. However, it is not ideal as it is a complex optical system to develop, and it requires precise co-registration making it challenging for other labs to reproduce. In this proposal, we are developing a new approach to study the 3D motion of the cochlea that involves extracting the vibrational angles encoded in OCT image stacks collected using a traditional OCT system. By scanning our sample at higher frame rates, we can obtain sequential cross-sectional images in the XZ and YZ planes and extract the corresponding vibrational angles. With this information, we can compute the 3D motion vector through the vibration-based modulation of the image’s spatial frequencies. We have validated this approach for low frequencies using a piezoelectric phantom with a layer of scattering material. However, we want to extend this approach to work for higher frequencies so we can collect measurements from the apical region of the mouse cochlea (2 kHz-15 kHz). Obtaining 3D vibrometry from the mouse cochlea using a more accessible approach with a traditional OCT system will enable our lab and others to build on these studies. In Aim 1, we will be implementing changes to the acquisition of our OCT system to be able to perform interleaved sampling. This will allow us to use our approach for our target frequency range of 2 kHz to 15 kHz. We will validate these changes using a position sensor to check the scan pattern and piezo experiments to ensure we can use our speckle approach at the desired frequencies. These validation experiments will give us the confidence to move forward with Aim 2, where we will use our speckle approach to measure the 3D motion vector for different structures within the mouse cochlea. We will compare the measured 3D vector of motion from vibrometry points taken from the tectorial membrane, reticular lamina, outer hair cells, inner hair cells, and basilar membrane to our current results with the 3-beam OCT system.