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 (CO) 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.
The effect of extremely low frequency electromagnetic fields (EMF) on microvesicles was examined in rat astrocytes by video-enhanced microscopy in combination with a perfusable cell chamber. The EMF effect was compared with the effect of heat shock (HS) and with a combination of them both. The velocity of microvesicles was measured using image processing software (NIH Scion image 1.61). After exposure of astrocytes to EMF (50 Hz, 100microT, 1 h), the velocity of microvesicles in astrocytes increased from 0.32 +/- 0.03 microm/s (n = 120, 95% CI) in the untreated control group to 0.41 +/- 0.03 microm/s (n = 175, 95% CI). Fifteen minutes after HS (45 degrees C, 10 min) the microvesicles showed a velocity of 0.56 +/- 0.03 microm/s (n = 125, 95% CI). Combination of HS and EMF led to an increase in velocity up to 0.54 +/- 0.03 microm/s (n = 110, 95% CI). No significant difference between HS and HS+EMF was found. Compared to the untreated control group, the increased microvesicle velocity of the exposed cells might be a stress response of the cell. It is possibly a sign of intensified intracellular traffic required to adjust the metabolic needs.
EINLEITUNG:Ataxie ist ein Symptom sowohl peripher-, als auch zentral-neurologischer Störungen. Eine Differenzierung ist allein nach klinisch-neurologischer Untersuchung nicht immer sicher möglich. Auf der Suche nach einer einfachen Zusatzuntersuchung, die zur Trennung beitragen kann, wurden Patienten mit Kleinhirnerkrankungen und Polyneuropathien mittels Kraftmeßplatte und modifiziertem Romberg-Test untersucht.
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