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Journal of Neuroscience, Vol 14, 7511-7520, Copyright © 1994 by Society for Neuroscience
Disruption of microfilaments in growth cones following depolarization and calcium influx
MD Neely and M Gesemann
Department of Pharmacology, Biozentrum of the University of Basel, Switzerland.
Depolarization of leech neurons growing on extracellular matrix extract
(ECM) leads to cessation of neurite outgrowth, rounding up of the
peripheral regions of the growth cone, loss of filopodia, and neurite
retraction. These responses depend on the influx of calcium (Neely, 1993).
The aim of the present experiments was to analyze how the cytoskeleton
becomes reorganized as growth cones change their morphology.
Immunocytochemistry revealed a loss of microfilaments in the tips of
neurites growing on ECM after depolarization. Leech neurons cultured on a
different substrate, the plant lectin concanavalin A (ConA), continue to
grow during and after depolarization (Grumbacher- Reinert and Nicholls,
1992; Neely, 1993). As expected, we did not observe any change in the
distribution of microfilaments after depolarization on ConA. Since there is
evidence that this lack of response is due to a reduced calcium influx
during depolarization of neurons on ConA (Ross et al., 1988), the effect of
the calcium ionophore A23187 on the outgrowth of these cells was analyzed.
In the absence of depolarization, this ionophore caused cessation of growth
cone motility and a loss of microfilaments, while microtubules were not
affected. Cytochalasin D, a microfilament-disrupting agent, induced changes
in growth cone morphology and neurite retraction similar to those observed
after depolarization and calcium influx. Application of phalloidin, a drug
that stabilizes microfilaments, inhibited depolarization-induced retraction
of neurites on ECM. By contrast, stabilization of microtubules with taxol
did not prevent depolarization from inducing changes in growth cone
morphology and neurite growth. These experiments show that changes in
growth cone morphology and motility of leech neurons induced by
depolarization and calcium influx are accompanied by a dramatic change in
the organization of microfilaments, but not microtubules.
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