Méhes E$, Czirók A, Hegedüs B, Szabó B, Vicsek T, Satz J*, Campbell K*, Jancsik V$:
Dystroglycan is involved in laminin1 stimulated motility of Muller glial cells; a combined velocity and directionality analysis
accepted in GLIA
$ Department of Anatomy and Histology, Faculty of Veterinary Science, Szent István University, Budapest
* Howard Hughes Medical Institute, University of Iowa, Roy J. and Lucille A. Carver College of Medicine, Iowa City, USA


We investigate the role of dystroglycan, a major laminin-1 receptor and central member of the dystrophin glycoprotein complex, in the laminin-1 induced motility of cultured Muller glial cells. Binding of laminin-1 to dystroglycan was prevented by IIH6, a function-blocking monoclonal antibody against alpha-dystroglycan. As an alternative way of inhibition, we used heparin to mask the dystroglycan binding site of the laminin-1, known to overlap with heparin binding sites. Cell motility was characterized in a two dimensional motility assay based on computer-controlled videomicroscopy and statistical analysis of cellular trajectories. We obtained data on both cell velocity and diffusion index, a measure of direction-changing frequency. Both ways of inhibition of dystroglycan function resulted in a significant decrease in the ability of laminin-1 to stimulate cell migration. At the same time, dystroglycan function does not seem to be involved in laminin-1 dependent increase in process dynamism and direction-changing activity.

EPS or JPEG image showing randomly chosen (30) cell trajectories from movies:

(A: untreated surface, B: laminin-1 covered surface, C: laminin-1 covered surface + heparin masking, D: laminin-1 covered surface + a-DG antibody). Scales are plotted in microns. Two representative trajectories are plotted with thick line in each panel. Note the difference in trajectory lengths.

Movies:

Composite movie of isolated rat Muller glial cells on untreated (left) or laminin-1 covered (right) surfaces:
MPEG stream
Objective: 20X (Field of view: 650um X 500um). Duration of recording: 20 hours. Number of frames: 122 (10 min/frame).
Cells isolated at postnatal day 10 were kept in culture for 12 days.
Note the differences in migratory activity in terms of both cell velocity and turning frequency. Cell trajectories are plotted on the movie. Bar: 20 microns.

Composite movie of isolated rat Muller glial cells on laminin-1 covered surface (left) or laminin-1 covered surface + anti-dystroglycan function-blocking antibody (right) :
MPEG stream
Objective: 20X x 1.7 (Field of view: 400um X 300um). Duration of recording: 21 hours. Number of frames: 126 (10 min/frame).
Cells isolated at postnatal day 10 were kept in culture for 9 days.
Note the differences in cell velocity. Cell trajectories are plotted on the movie. Bar: 20 microns.

Composite movie of isolated rat Muller glial cells on laminin-1 covered (left) or laminin-1 covered + heparin masked (right) surfaces:
MPEG stream
Objective: 20X (Field of view: 650um X 500um). Duration of recording: 17 hours. Number of frames: 103 (10 min/frame).
Cells isolated at postnatal day 10 were kept in culture for 10 days
Note the differences in cell velocity. Cell trajectories are plotted on the movie. Bar: 20 microns.