High speed, high resolution and high sensitivity are desirable for optical coherence tomography (OCT). We demonstrate space-division multiplexing (SDM) technology that translates long coherence length of a commercially available wavelength tunable laser into high OCT imaging speed. In our 1st generation SDM-OCT system, we achieved an effective 800,000 A-scans/s imaging speed using a 100,000 Hz tunable vertical cavity surface-emitting laser (VCSEL). A sensitivity of 94.6 dB and a roll-off of < 2 dB over ~30 mm imaging depth were measured from a single channel. An axial resolution of ~11 μm in the air (or ~8.3 μm in tissue) was achieved throughout the entire depth range. Our 1st generation SDM-OCT system was developed based on fiber optics. However, it required extensive efforts to assemble fiber components (optical splitter, optical delay, fiber array, in the red rectangle of figure A) and control optical delays between different channels by hand, which made it challenging for mass-reproduction. As a solution, we seek to integrate all these fiber components into a photonic chip and incorporate it into our 2nd generation SDM-OCT system. Optical delays and spacing between each output beam of the chip can be customized and fabricated precisely with sub-micron tolerances. The SDM technology provides a new orthogonal dimension for further speed improvement for OCT with favorable cost scaling. With integrated photonic chips, cost scaling will be further reduced, which facilitates broad dissemination of the SDM-OCT technology. SDM-OCT also preserves image resolution and allows synchronized cross-sectional and three-dimensional (3D) imaging of biological samples, enabling new biomedical applications.

OCT is a powerful tool for assessing tissue architectural morphology. It enables 3D imaging with resolutions approaching standard histopathology (a few microns), and it can be performed in vivo and in real-time without the need to remove and process specimens. OCM combines coherence-gated detection with confocal microscopy in order to achieve high transverse resolutions, thus enabling 3D visualization of cellular features. However, current OCT and OCM imaging technologies have not been able to leverage the recent advances in molecular-targeted contrast agents that are revolutionizing biomedicine. In this project, we will develop and validate techniques that enable molecular contrast for 3D-OCT and OCM. The successful completion of this project will allow both the structure and pathological states of tissue to be imaged in 3D, in vivo, and in real time with micron-level spatial resolutions at multiple scales. This work will lay the foundation for a wide range of fundamental research, small animal imaging, and future clinical applications in humans. This work will also serve as a starting point for the OCT and OCM studies of other pathologies associated with abnormal protein expression levels, such as neurodegenerative and cardiovascular diseases. This work is supported by NIH/NIBIB through the Pathway to Independence Award (K99/R00).

Electrical stimulation is currently the gold standard for cardiac pacing. However, it is invasive and nonspecific for cardiac tissues. We recently developed a noninvasive cardiac pacing technique using optogenetic tools, which are widely used in neuroscience. Optogenetic pacing of the heart provides high spatial and temporal precisions, is specific for cardiac tissues, avoids artifacts associated with electrical stimulation, and therefore promises to be a powerful tool in basic cardiac research. We demonstrated optogenetic control of heart rhythm in a well-established model organism, Drosophila melanogaster. We developed transgenic flies expressing a light-gated cation channel, channelrhodopsin-2 (ChR2), specifically in their hearts and demonstrated successful optogenetic pacing of ChR2-expressing Drosophila at different developmental stages, including the larva, pupa, and adult stages. A high-speed and ultrahigh-resolution optical coherence microscopy imaging system that is capable of providing images at a rate of 130 frames/s with axial and transverse resolutions of 1.5 and 3.9 mm, respectively, was used to noninvasively monitor Drosophila cardiac function and its response to pacing stimulation. The development of a noninvasive integrated optical pacing and imaging system provides a novel platform for performing research studies in developmental cardiology.

Circadian rhythms are endogenous, entrainable oscillations of physical, mental and behavioral processes in response to local environmental cues such as daylight, which are present in the living beings, including humans. Circadian rhythms have been related to cardiovascular function and pathology. However, the role that circadian clock genes play in heart development and function in a whole animal in vivo are poorly understood. The Drosophila cryptochrome (dCry) is a circadian clock gene that encodes a major component of the circadian clock negative feedback loop. In this study, we utilized ultrahigh-resolution optical coherence microscopy (OCM) system to perform non-invasive and longitudinal analysis of functional and morphological changes in the Drosophila heart throughout its post-embryonic lifecycle for the first time. The Drosophila heart exhibited major morphological and functional alterations during its development. In order to study the functional role of dCry on Drosophila heart development, we silenced dCry by RNAi in the Drosophila heart and mesoderm, and quantitatively measured heart morphology and function in those flies throughout its development. Silencing of dCry resulted in slower HR, reduced CAP, smaller heart chamber size, pupal lethality and disrupted posterior segmentation that was related to increased expression of a posterior compartment protein, wingless. Collectively, our studies provided novel evidence that the circadian clock gene, dCry, plays an essential role in heart morphogenesis and function.

Normal brain function depends on the delivery of oxygen and glucose, and on clearance of the byproducts of metabolism. Thus, an understanding of the normal and pathological conditions of oxygen supply and consumption, and measurement of blood flow is important for basic neuroscience and clinical applications. To this end, a variety of tools have been developed to image cerebral hemodynamics. For example, transcranial Doppler is a common clinical tool but is limited to the measurement of blood flow within large vessels. Functional (blood oxygen level dependent – BOLD, or arterial spin labeled – ASL) MRI provides 3D tomography of the brain with moderate spatial resolution (a few millimeters). PET measures cerebral blood flow and oxygen metabolism with a decreased spatial resolution compared to MRI. Currently, the laboratory use of MRI and PET based techniques are limited due to high cost, low spatial and temporal resolution, and low mobility. Optical imaging techniques, such as optical intrinsic imaging and LSI, can be used to extract cerebral blood oxygenation and blood flow information at high spatial and temporal resolutions. However, optical intrinsic imaging and LSI are limited to the mapping of brain functions only in 2D. We will develop novel OCT imaging techniques to image 3D brain functions in animal models. Not only can OCT provide structural information of the animal cortex at micron-scale resolution, but can also be used to extract 3D cerebral hemodynamic information by using Doppler (for blood flow) and spectroscopic (for blood oxygenation) OCT techniques. The 1-2 mm penetration depth of OCT allows imaging through thinned skull rather than opened skull, which makes longitudinal studies possible. The combination of 3D mapping of blood flow and oxygenation will enable for the first time imaging of cerebral oxygen metabolism in 3D at micron-scale resolution. The successful completion of the development of this technique will enable us to investigate 3D brain functions in physiological (e.g. during forepaw, hind paw and whisker stimulations), and pathophysiological (eg. cortical spreading depression, ischemic and traumatic brain injuries) conditions in animal models.

For this project, we focused on evaluating the neuronal injury induced by spontaneous seizure in rat’s hippocampus using OCM. Compared to confocal microscopy, OCM is an imaging modality with label-free and greater imaging depth. We designed and built an OCM system to achieve 3D images of ~300µm-thick slice of rat’s hippocampus with cellular resolution in en face view. Currently, we are working on evaluating the change of neuron morphology and number in rat’s hippocampus from different days in vitro. Also, we plan to record the electrical signal from the spontaneous seizure and image the hippocampus sample using OCM at the same time to explore more about the seizure and its affection to the neurons. If successful, OCM would be a promising tool for researchers to evaluate neuronal injury induced by the spontaneous seizure with rat’s hippocampus mode and help to develop better treatments for epilepsy.