Filament displacement image analytics tool for use in investigating dynamics of dense microtubule networks.

The fate and motion of cells is influenced by a variety of physical characteristics of their microenvironments. Traditionally, mechanobiology focuses on external mechanical phenomena such as cell movement and environmental sensing. However, cells are inherently dynamic, where internal waves and internal oscillations are a hallmark of living cells observed under a microscope. We propose that these internal mechanical rhythms provide valuable information about cell health. Therefore, it is valuable to capture the rhythms inside cells and quantify how drugs or physical interventions affect a cell's internal dynamics. One of the key dynamical entities inside cells is the microtubule network. Typically, microtubule dynamics are measured by end-protein tracking. In contrast, this paper introduces an easy-to-implement approach to measure the lateral motion of the microtubule filaments embedded within dense networks with (at least) confocal resolution image sequences. Our tool couples the computer vision algorithm Optical Flow with an anisotropic, rotating Laplacian of Gaussian filtering to characterize the lateral motion of dense microtubule networks. We then showcase additional image analytics used to understand the effect of microtubule orientation and regional location on lateral motion. We argue that our tool and these additional metrics provide a fuller picture of the active forcing environment within cells.

Decoding Natural Astrocyte Rhythms: Dynamic Actin Waves Result from Environmental Sensing by Primary Rodent Astrocytes.

Astrocytes are key regulators of brain homeostasis, equilibrating ion, water, and neurotransmitter concentrations and maintaining essential conditions for proper cognitive function. Recently, it has been shown that the excitability of the actin cytoskeleton manifests in second-scale dynamic fluctuations and acts as a sensor of chemophysical environmental cues. However, it is not known whether the cytoskeleton is excitable in astrocytes and how the homeostatic function of astrocytes is linked to the dynamics of the cytoskeleton. Here it is shown that homeostatic regulation involves the excitable dynamics of actin in certain subcellular regions of astrocytes, especially near the cell boundary. The results further indicate that actin dynamics concentrate into "hotspot" regions that selectively respond to certain chemophysical stimuli, specifically the homeostatic challenges of ion or water concentration increases. Substrate topography makes the actin dynamics of astrocytes weaker. Super-resolution images demonstrate that surface topography is also associated with the predominant perpendicular alignment of actin filaments near the cell boundary, whereas flat substrates result in an actin cortex mainly parallel to the cell boundary. Additionally, coculture with neurons increases both the probability of actin dynamics and the strength of hotspots. The excitable systems character of actin thus makes astrocytes direct participants in neural cell network dynamics.