MOTIVATION: Multispectral biological fluorescence microscopy has enabled the identification of multiple targets in complex samples. The accuracy in the unmixing result degrades (i) as the number of fluorophores used in any experiment increases and (ii) as the signal-to-noise ratio in the recorded images decreases. Further, the availability of prior knowledge regarding the expected spatial distributions of fluorophores in images of labeled cells provides an opportunity to improve the accuracy of fluorophore identification and abundance. RESULTS: We propose a regularized sparse and low-rank Poisson regression unmixing approach (SL-PRU) to deconvolve spectral images labeled with highly overlapping fluorophores which are recorded in low signal-to-noise regimes. First, SL-PRU implements multipenalty terms when pursuing sparseness and spatial correlation of the resulting abundances in small neighborhoods simultaneously. Second, SL-PRU makes use of Poisson regression for unmixing instead of least squares regression to better estimate photon abundance. Third, we propose a method to tune the SL-PRU parameters involved in the unmixing procedure in the absence of knowledge of the ground truth abundance information in a recorded image. By validating on simulated and real-world images, we show that our proposed method leads to improved accuracy in unmixing fluorophores with highly overlapping spectra. AVAILABILITY AND IMPLEMENTATION: The source code used for this article was written in MATLAB and is available with the test data at https://github.com/WANGRUOGU/SL-PRU.

Glutamate transporters preserve the spatial specificity of synaptic transmission by limiting glutamate diffusion away from the synaptic cleft, and prevent excitotoxicity by keeping the extracellular concentration of glutamate at low nanomolar levels. Glutamate transporters are abundantly expressed in astrocytes, and previous estimates have been obtained about their surface expression in astrocytes of the rat hippocampus and cerebellum. Analogous estimates for the mouse hippocampus are currently not available. In this work, we derive the surface density of astrocytic glutamate transporters in mice of different ages via quantitative dot blot. We find that the surface density of glial glutamate transporters is similar in 7-8 week old mice and rats. In mice, the levels of glutamate transporters increase until about 6 months of age and then begin to decline slowly. Our data, obtained from a combination of experimental and modeling approaches, point to the existence of stark differences in the density of expression of glutamate transporters across different sub-cellular compartments, indicating that the extent to which astrocytes limit extrasynaptic glutamate diffusion depends not only on their level of synaptic coverage, but also on the identity of the astrocyte compartment in contact with the synapse. Together, these findings provide information on how heterogeneity in the spatial distribution of glutamate transporters in the plasma membrane of hippocampal astrocytes my alter glutamate receptor activation out of the synaptic cleft.