title: Quantifying Filopodia in Cultured Astrocytes by an Algorithm.
abstract: Astrocytes in vivo extend thin processes termed peripheral astrocyte processes (PAPs), in particular around synapses where they can mediate glia-neuronal communication. The relation of PAPs to synapses is not based on coincidence, but it is not clear which stimuli and mechanisms lead to their formation and are active during process extension/ retraction in response to neuronal activity. Also, the molecular basis of the extremely fine PAP morphology (often 50 to 100 nm) is not understood. These open questions can be best investigated under in vitro conditions studying glial filopodia. We have previously analyzed filopodial mechanisms (Lavialle et al. PNAS 108:12915) applying an automated method for filopodia morphometry, which is now described in greater detail. The Filopodia Specific Shape Factor (FSSF) developed integrates number and length of filopodia. It quantifies filopodia independent of overall astrocytic shape or size, which can be intricate in itself. The algorithm supplied here permits automated image processing and measurements using ImageJ. Cells have to be sampled in higher numbers to obtain significant results. We validate the FSSF, and characterize the systematic influence of thresholding and camera pixel grid on measurements. We provide exemplary results of substance-induced filopodia dynamics (glutamate, mGluR agonists, EGF), and show that filopodia formation is highly sensitive to medium pH (CO2) and duration of cell culture. Although the FSSF was developed to study astrocyte filopodia with focus on the perisynaptic glial sheath, we expect that this parameter can also be applied to neuronal growth cones, non-neural cell types, or cell lines.
authors.affiliation: Institute of Anatomy, University of Dresden, Dresden, Germany.|Hospital of Günzburg, Günzburg, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Aumann
authors.affiliation: Department of Neurology, University Hospital Frankfurt, Goethe-University Frankfurt, Frankfurt am Main, Germany.
authors.forename: Felix
authors.initials: F
authors.lastname: Friedländer
authors.affiliation: Department of Medical Physics and Biomedical Engineering, University of Dresden, Dresden, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Thümmler
authors.affiliation: Institute of Anatomy II, Goethe-University Frankfurt, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Keil
authors.affiliation: Department of Neurology, University Hospital Frankfurt, Goethe-University Frankfurt, Frankfurt am Main, Germany.
authors.forename: Robert
authors.initials: R
authors.lastname: Brunkhorst
authors.affiliation: Institute of Anatomy II, Goethe-University Frankfurt, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany.|Dr. Senckenbergisches Chronomedizinisches Institut, University of Frankfurt, Frankfurt am Main, Germany.
authors.forename: Horst-Werner
authors.initials: HW
authors.lastname: Korf
authors.affiliation: Institute of Anatomy II, Goethe-University Frankfurt, Theodor-Stern-Kai 7, 60590, Frankfurt am Main, Germany. Derouiche@em.uni-frankfurt.de.|Dr. Senckenbergisches Chronomedizinisches Institut, University of Frankfurt, Frankfurt am Main, Germany. Derouiche@em.uni-frankfurt.de.
authors.forename: Amin
authors.identifier: http://orcid.org/0000-0002-7309-2044
authors.initials: A
authors.lastname: Derouiche
mesh_terms: D000465:Algorithms; D000818:Animals; D000831:Animals, Newborn; D002452:Cell Count / Q000379:methods; D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology* / Q000502:physiology*; D011554:Pseudopodia / Q000502:physiology*; D051381:Rats
keywords: Actin cytoskeleton; Cell culture; Cell motility; Glia-synaptic interaction; Glutamate; Morphometry