We present artificial confocal microscopy (ACM) to achieve confocal-level depth sectioning, sensitivity, and chemical specificity non-destructively on unlabeled specimens. ACM is equipped with a laser scanning confocal microscopy with a quantitative phase imaging module, which provides optical path-length maps of the specimen colocalized with the fluorescence channel. Using pairs of phase and fluorescence images, a convolution neural network was trained to translate the former into the latter. The ACM images hold much stronger depth sectioning than the input (phase) images, enabling us to recover confocal-like tomographic volumes of microspheres, hippocampal neurons in culture, and three-dimensional liver cancer spheroids.
Traditionally the measurement of electrical activity in neurons has been carried out using microelectrode arrays that require the conducting elements to be in contact with the neuronal network. This method, also referred to as “electrophysiology”, while being excellent in terms of temporal resolution is limited in spatial resolution and is invasive. An optical microscopy method for measuring electrical activity is thus highly desired. Common-path quantitative phase imaging (QPI) systems are good candidates for such investigations as they provide high sensitivity (on the order of nanometers) to the plasma membrane fluctuations that can be linked to electrical activity in a neuronal circuit. In this work we measured electrical activity in a culture of rat cortical neurons using MISS microscopy, a high-speed common-path QPI technique having an axial resolution of around 1 nm in optical path-length, which we introduced at PW BIOS 2016. Specifically, we measured the vesicular cycling (endocytosis and exocytosis) occurring at axon terminals of the neurons due to electrical activity caused by adding a high K+ solution to the cell culture. The axon terminals were localized using a micro-fluidic device that separated them from the rest of the culture. Stacks of images of these terminals were acquired at 826 fps both before and after K+ excitation and the temporal standard deviation maps for the two cases were compared to measure the membrane fluctuations. Concurrently, the existence of vesicular cycling was confirmed through fluorescent tagging and imaging of the vesicles at and around the axon terminals.
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