Sheet Illumination microscopy is a well developed tool in the bio-imaging community due to many inherent advantages. Increased photonic efficiency allows for lower power light sources to be used, which in turn reduce phototoxic damage at the sample over time. High fidelity, high SNR data can be collected from these systems due to their optical sectioning capabilities, and as such, many have been designed to operate within a limited parameter space. These instruments often follow traditional design logic, for specialization in one imaging condition, to observe only a single type of scientific data. This mentality yields high resolution results for a particular class of data, yet is limited in terms of flexibility. Here, the design and development of a user-configurable light sheet microscope is discussed, which uses a spatial phase modulator to dynamically shape and optimize the incident light sheet to perform highly in a number of temporal and spatial scales. The result is a general purpose microscopy platform for the observation of many biological phenomena.

This dissertation is focused on the design, development, and implementation of two high performance optical microscopy systems that enable study of neuronal circuit function in new and innovative ways.

Firstly, an easily adopted, open-source miniature 2-photon microscope (UCLA 2P Miniscope) capable of recording calcium dynamics from neurons located in deep structures and in dendrites over a (445µm x 380µm) field of view (FOV) during free behavior is described. The system weighs approximately 4g and utilizes two on-board silicon-based photon detectors for highly sensitive measurements. All hardware is designed for high performance and ease of assembly, while minimizing cost. To test the 2P miniature microscope, recordings in three experimental conditions were conducted to highlight system capabilities during free behavior in mice. First, calcium dynamics from place cells in hippocampal area CA1 were recorded. Next, calcium transients from dendrites in retrosplenial cortex were resolved during 30 minutes of free behavior. Lastly, dentate granule cell activity was recorded at a depth of over 620µm, through an intact hippocampal formation during an open field behavior. The dentate granule cell recordings, to our knowledge, are the first optical recordings from these neurons ever performed in the intact hippocampus during free behavior. The miniature microscope itself and all supporting equipment are open-source and all files needed for building the scope can be accessed through the UCLA Golshani Lab GitHub repository.

Secondly, a high-speed, open-source voltage imaging microscope is presented, capable of resolving (920µm x 460µm) fields of view at 500 frames per second. This microscope enables the study of fast-spiking neural populations such as interneurons with single action potential sensitivity over periods greater than 10 minutes. The system is also able to resolve activity from populations of many neurons simultaneously, with some recordings exceeding 30 active neurons in a single FOV. Additionally, this microscope system is able to quantify sub-threshold membrane potentials, making the recording of oscillatory dynamics and excitatory post synaptic potentials possible in large neural circuits in-vivo. Altogether, this high-speed, large FOV system enables new studies into the critical mechanisms key subpopulations of neurons employ to support computation and learning over time.

Both of the microscope systems described here enable new insights into the function of the central nervous system. The first (Part I) is a miniature multiphoton microscope capable of studying fine structures deep in the brain during free and naturalistic behavior. The second (Part II) is a high-speed system able to record rapidly changing electrical dynamics across large neural populations. Together they will lead to new understandings of how the brain functions both in disease and health.