Claudio Moretti, Sylvain Gigan
Fluorescence is a powerful mean to probe information processing in the mammalian brain. However, neuronal tissues are highly heterogeneous and thus opaque to light. A wide set of non-invasive or invasive techniques for scattered light rejection, optical sectioning or localized excitation, have been developed, but non-invasive optical recording of activity through highly scattering layer beyond the ballistic regime is to date impossible. Here, we show that functional signals from fluorescent time-varying sources located below an highly scattering tissue can be retrieved efficiently, by exploiting matrix factorization algorithms to demix this information from low contrast fluorescence speckle patterns.
Caio Vaz Rimoli, Claudio Moretti, Fernando Soldevila, Enora Brémont, Cathie Ventalon, Sylvain Gigan
Optical methods based on thin multimode fibers (MMFs) are promising tools for measuring neuronal activity in deep brain regions of freely moving mice thanks to their small diameter. However, current methods are limited: while fiber photometry provides only ensemble activity, imaging techniques using of long multimode fibers are very sensitive to bending and have not been applied to unrestrained rodents yet. Here, we demonstrate the fundamentals of a new approach using a short MMF coupled to a miniscope. In proof-of-principle in vitro experiments, we disentangled spatio-temporal fluorescence signals from multiple fluorescent sources transmitted by a thin (200 μm) and short (8 mm) MMF, using a general unconstrained non-negative matrix factorization algorithm directly on the raw video data. Furthermore, we show that low-cost open-source miniscopes have sufficient sensitivity to image the same fluorescence patterns seen in our proof-of-principle experiment, suggesting a new avenue for novel minimally invasive deep brain studies using multimode fibers in freely behaving mice.
Lei Zhu, Fernando Soldevila, Claudio Moretti, Alexandra d'Arco, Antoine Boniface, Xiaopeng Shao, Hilton B. de Aguiar, Sylvain Gigan
On-invasive optical imaging techniques are essential diagnostic tools in many fields. Although various recent methods have been proposed to utilize and control light in multiple scattering media, non-invasive optical imaging through and inside scattering layers across a large field of view remains elusive due to the physical limits set by the optical memory effect, especially without wavefront shaping techniques. Here, we demonstrate an approach that enables non-invasive fluorescence imaging behind scattering layers with field-of-views extending well beyond the optical memory effect. The method consists in demixing the speckle patterns emitted by a fluorescent object under variable unknown random illumination, using matrix factorization and a novel fingerprint-based reconstruction. Experimental validation shows the efficiency and robustness of the method with various fluorescent samples, covering a field of view up to three times the optical memory effect range. Our non-invasive imaging technique is simple, neither requires a spatial light modulator nor a guide star, and can be generalized to a wide range of incoherent contrast mechanisms and illumination schemes.
Fernando Soldevila, Claudio Moretti, Tobias Nöbauer, Hossein Sarafraz, Alipasha Vaziri, Sylvain Gigan
Recently, fluorescence-based optical techniques have emerged as a powerful tool to probe information in the mammalian brain. However, tissue heterogeneities prevent clear imaging of deep neuron bodies due to light scattering. While several up-to-date approaches based on ballistic light allow to retrieve information at shallow depths inside the brain, non-invasive localization and functional imaging at depth still remains a challenge. It was recently shown that functional signals from time-varying fluorescent emitters located behind scattering samples could be retrieved by using a matrix factorization algorithm. Here we show that the seemingly information-less, low-contrast fluorescent speckle patterns recovered by the algorithm can be used to locate each individual emitter, even in the presence of background fluorescence. We test our approach by imaging the temporal activity of large groups of fluorescent sources behind different scattering phantoms mimicking biological tissues, and through a brain slice with a thickness of ~200 micron.