Implantable Fluorescence Endoscopic Microscopy and Its Application in In Vivo Brain Imaging (Invited)

Significance The neurovascular unit (NVU), a critical component of the brain, regulates almost all physiological process. The precision of the morphology and function presentation regarding the NVU provides hope for advancing research on basic neuroscience,as well as diagnosing brain diseases, which are common desires of the "Brain Project" worldwide. Accordingly, high temporal and spatial resolution visualization techniques are required. Fluorescence microscopic imaging technology has significant advantages in terms of specificity, diversity, image contrast, and spatio-temporal resolution; however, due to the limited penetration depth of light in tissue, use of noninvasive fluorescence imaging to obtain high-resolution structural and functional information of NVU is difficult in deep brain regions in vivo. As a result, fluorescence endoscopic microscopy imaging technologies based on micro probes are becoming more popular among brain science researchers. ProgressOver the last two decades, a series of neurobehavioral studies in vivo have been conducted using fluorescence endoscopic microscopy. With endoscopic probes implanted into the brain, the NVU in most deep regions can be observed clearly in living mice,including the hippocampus, dorsal striatum, amygdaloid nucleus, and epithalamus. Incorporating an upright microscope or a head-mounted mini microscope, gradient refractive index (GRIN) lenses have been widely employed as an implantable probe, with the advantage of excellent stability, high resolution, and low cost. In addition, a potential strategy for implantable imaging of the brain in vivo involves using a single multimode fiber, based on modulation of the light field, to focus and scan spot at the end of multimode fiber. This reduces tissue damage, with resolution at the cellular level. Herein, the recent progression of implantable fluorescence endoscopic microscopy is reviewed based on both GRIN lens and a single multimode fiber, besides application research in vivoincluding blood velocity, neurons growth, calcium ion conduction, and so on. Finally, fluorescence endoscopic microscopy imaging technologies for clinical diagnosis of brain tumors are also introduced, demonstrating that these advanced optical imaging methods expand the toolbox for brain science research and disease diagnosis. Conclusions and ProspectsEndoscopic probes have been miniaturized, providing greater flexibility while maintaining high performance; thus, probes can be implanted at different depths in the living brain to carry out functional modulation studies in specific deep brain regions. With micromachining or adaptive optics technologies, GRIN lens provides an effective method to obtain high resolution images. Although the nonmechanical scan imaging through a single multimode fiber is a relatively new exploration for brain research in vivo, it has already exhibited the unique advantages of minimally invasive and flexibility. In future, the following considerations are worth exploring:(1) development of a high-performance multimode fiber with enhanced anti-interference ability to external disturbances;(2) processing of a microlens on the face of multimode fiber with precise 3D printing technology, to optimize imaging resolution, depth of field, and field of view;(3) introduction of fluorescence polarization and fluorescence lifetime imaging modes to analyze neuronal physiological information, such as protein dipoles and cellular microenvironment