Fast Three-Dimensional Single-Particle Tracking in Natural Brain Tissue

Abstract

Observation of molecular dynamics is often biased by the optical very heterogeneous environment of cells and complex tissue. Here, we have designed an algorithm that facilitates molecular dynamic analyses within brain slices. We adjust fast astigmatism-based three-dimensional single-particle tracking techniques to depth-dependent optical aberrations induced by the refractive index mismatch so that they are applicable to complex samples. In contrast to existing techniques, our online calibration method determines the aberration directly from the acquired two-dimensional image stream by exploiting the inherent particle movement and the redundancy introduced by the astigmatism. The method improves the positioning by reducing the systematic errors introduced by the aberrations, and allows correct derivation of the cellular morphology and molecular diffusion parameters in three dimensions independently of the imaging depth. No additional experimental effort for the user is required. Our method will be useful for many imaging configurations, which allow imaging in deep cellular structures.

Multiple particle tracking detects changes in brain extracellular matrix structure and predicts neurodevelopmental age

“…Collectively, this shows MPT can be used to capture changes in brain ECM structure. While diffusion-based techniques for characterizing the brain microenvironment already exist, researchers have been limited to either taking macroscopic approaches to quantifying diffusion related parameters (Hrabetova and Nicholson, 2007;Nicholson et al, 1979;Nicholson and Tao, 1993;Patlak and Fenstermacher, 1975;Sykova and Nicholson, 2008) or only being able to track individual particles, separately (Godin et al, 2017;Sokoll et al, 2015). Macroscopic approaches fall short in their ability to provide microscopic spatial resolution, the resolution needed to make inferences on the cellular and local extracellular level.…”

Surface trafficking of neurotransmitter receptors: From cultured neurons to intact brain preparations

“…semiconductor nano-crystals typically composed of a core of CdSe beneath a shell of ZnS -are particularly well suited for this purpose with their relatively small size (10 nm), wide excitation but narrow emission spectra, and outstanding brightness and photostability and have been used to pioneer investigations in brain slices (Michalet et al, 2005). Using spinning disk confocal microscopy, Biermann and collaborators first demonstrated the feasibility of measuring the surface dynamics of lipids (glycosylphosphatidylinositol, GPI) and cell adhesion molecules (neuroligin 1, Nlg1; β-neurexin) in three dimensions (3D) in organotypic brain slices using antibodies coupled to red light-emitting QDs (QD605 nm / QD655 nm) (Biermann et al, 2014;Sokoll et al, 2015).…”