The proposal aims to acquire an integrated multi-photon live imaging system that is also capable of simultaneous in vitro and in vivo electrophysiology. The system will consist of three main components: 1) a two-photon microscope including an ultrafast IR laser for multi-photon excitation, a laser scanning head, optics, an air table, an acquisition computer and software; 2) patch clamp equipment including an amplifier, software, a digital acquisition system, and manipulators; and 3) multiple imaging platforms for various samples including microscopic slides, dissociated single live cells, live tissue (i.e. a brain slice), and brain of a behaving animal. The system will be a multi-purpose rig for various projects and is intended to be shared primarily by a group of investigators within the Emory Neuromodulation and Technology Innovation Center and Emory/Georgia Tech Biomedical Engineering Departments, but will also be available to investigators in other departments. We combine interdisciplinary approaches to understand neural circuits underlying brain functions (e.g. tactile and auditory sensation) as well as to develop interventions for various neurological and psychiatric disorders in rodent models towards the treatment of human conditions (e.g. epilepsy, stroke, depression, Parkinson?s disease). We dissect these mechanisms at molecular and cellular levels and then evaluate neural circuits in whole animals. This system can be used for in vitro imaging in single cells or in acute brain slices while simultaneously monitoring neuronal electrical activity through patch clamp recording. It can be also used for in vivo imaging, such as calcium and membrane potential imaging in the brain to monitor activity of large numbers of neurons at a time, and in vivo optogenetic photostimulation in an awake, behaving animal to study neural circuits underlying animal behavior and diseases. In addition to physiological experiments, the microscope will also serve for histology using a whole, uncut brain treated with a tissue-clearing reagent (i.e. the CLARITY method). This method provides unperturbed morphology of neurons and circuits from the deep brain when two-photon excitation is utilized. Thus, the proposed system is intended to maximize capability to examine brain function in normal and disease conditions at different levels seamlessly. Currently available microscopes in our labs do not have such capability and microscopes in the university?s core facilities are designed for the single purpose of imaging and lack physiology capability, justifying an eminent need for the proposed system.
We will obtain a multi-purpose microscope that will support various scientific projects related to normal brain functions (e.g. perception and axon regeneration), and neurological and psychiatric disorders (e.g. epilepsy, Parkinson and stroke). The proposed system is intended to maximize capability to examine these mechanisms seamlessly at different levels, such as cellular levels, circuit levels, and in the whole awake and behaving animal. The last approach will be particularly important for conducting long-term studies in the same animals and future clinical translation.
