Recording all the neural activity in a 3D volume with millisecond timescale precision is a key goal of the BRAIN initiative. Recently, in a collaborative project with the Vaziri lab (IMP, Vienna), we adapted the strategy of lightfield microscopy for 3D volumetric imaging of fluorescent neural calcium responses (Prevedel 2014). This technology enables computational reconstruction of a 3D volume from an image by simultaneously capturing the angle of incident light rays in addition to their intensity. Imaging can occur as quickly as the fluorescent neural activity reporter allows (Chen 2013)? we imaged the entire larval zebrafish brain at 20 Hz. However, the spatial resolution for lightfield microscopy is poo, resulting in low signal to noise ratio (SNR) as well as difficulty in automatically segmenting neural anatomy, which is key to linking neural activity to underlying circuitry. This spatial resolution limit is a fundamental issue with lightfield microscopy, since to gain 3D imaging capability, one must sacrifice spatial resolution: there are only so many pixels on the camera. Accordingly, we here propose to perform the first whole brain recording of a larval zebrafish with single neuron resolution by increasing the total pixel count of our existing system by an order of magnitude whilst improving the SNR by leveraging a six fold increase in frame rate. Our existing lightfield imaging system (Prevedel 2014), and others (Levoy 2006, Cohen 2014), use an array of microlenses to effect the tradeoff of spatial for axial resolution. An alternativ approach captures the lightfield using an array of cameras, without any microlenses. Our novel design combines both approaches at an unprecedented scale. Our scalable data acquisition system (Willow, see preliminary data) combined with cameras designed inhouse limit the cost of our system to 1/10 of a traditional twophoton microscope. In this way we aim to develop a userfriendly system capable of imaging all of the neurons in a 3D volume, at speeds comparable to the natural timescales of neural activity, whilst keeping an eye towards polishing our system for marketability and widespread use.
Lightfield microscopy systems allow direct observation of neural activity in animal subjects over large 3D volumes of tissue. This facilitates the study of crucial neuroscientific topics such as development, learning and memory, and cognition, as well as brain diseases such as Alzheimer's, epilepsy, Parkinson's, and depression. Currently, these systems are custom engineered luxury items for research laboratories and have limited field of view, but LeafLabs plans to commoditize them by developing new, cheaper, and higher density cameras and data acquisition systems, and polishing this technology into a system made commercially available off the shelf.
| Piatkevich, Kiryl D; Jung, Erica E; Straub, Christoph et al. (2018) A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters. Nat Chem Biol 14:352-360 |
