Regulation of receptor‐mediated shape change in astroglial cells

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Abstract
Activation of adenylate cyclase in astroglial cells in culture results in a rapid change in cell shape that appears to occur by the active movement of cytoplasm from peripheral cell regions to the perinuclear space with processes being formed along regions that remain extended. Three series of experiments were designed to determine how shape change occurred. First, the Ca2+-dependency of shape change was determined by reducing intracellular Ca2+ concentrations to ≤50 nM or increasing intracellular Ca2+ concentrations to ≥1 μM. Neither of these changes significantly affected the rate of receptor-mediated shape change. Second the role that longer-lived, acetylated microtubules play in receptor-mediated shape change was assessed by visualizing microtubules using a polyclonal antibody to brain 6S tubulin or a monoclonal antibody to oligomers of tubulin to monitor total tubulin distribution and a monoclonal antibody to acetylated tubulin to describe the distribution of these microtubules. Three-dimensional distribution of microtubules was observed by optical sectioning of cultures using a laser scanning confocal imaging system. The distribution of acetylated tubules in control cells was similar to that observed with the antibodies to tubulin. Following treatment with 100 nM isoproterenol to stimulate shape change, there was a dramatic redistribution of microtubules; however, the distribution of acetylated tubules was again similar to the total microtubules. Analysis of the optical sections recorded using the confocal attachment revealed that while control cells were relatively flat (cell height = 4 μm), the perinuclear region of isoproterenol-treated cells extended much higher above the substrate (cell height = 13 μm). Third, the role of microtubule assembly and disassembly were assessed using colchicine and taxol. Results from these experiments suggest that microtubule reassembly is necessary for receptor-mediated shape change. Control experiments indicated that colchicine or taxol treatment did not inhibit either cAMP synthesis or another cAMP-dependent process, receptor-mediated taurine release. Together these results indicate that receptor-mediated shape change in astroglial cells occurs by a Ca2+-independent mechanism that results in active movement of cytoplasm to the perinuclear region. This process is dependent on microtubule reassembly suggesting that shape change may occur by active movement of material along microtubules or by microtubule redistribution.