Two pathways control chromaffin cell cortical F-actin dynamics during exocytosis

Neurosecretory cells including chromaffin cells possess a mesh of filamento us actin underneath the plasma membrane. We have proposed that the F-actin network acts as a barrier to the secretory vesicles blocking their access t o exocytotic sites at the plasma membrane. Disassembly of cortical F-actin in chromaffin cells in response to stimulation is thought to allow the free movement of secretory vesicles to exocytotic sites, Moreover, experiments by us using morphometric analysis of resting and stimulated chromaffin cell s together with membrane capacitance measurements have shown that cortical F-actin controls the traffic of vesicles from the vesicle reserve compartme nt to the release-ready Vesicle compartment. The dynamics of the cortical F -actin is controlled by two pathways: A) stimulation-induced Ca2+ entry and scinderin activation; and B) protein kinase C (PKC) activation and MARCKS (myristoylated alanine-rich C kinase substrate) phosphorylation. When chrom affin cells are stimulated through nicotinic receptors, cortical F-actin di sassembly is mainly through the intervention of pathway A, since in the pre sence of PKC inhibitors, F-actin disassembly in response to cholinergic sti mulation is only blocked by 20%. Pathway A involves the activation of scind erin by Ca2+ with a consequent F-actin severing. Pathway B is fully activat ed by phorbol esters and in this case PKC blockers inhibit by 100% the disr uption of cortical F-actin. This pathway operates through MARCKS. A peptide with amino acid sequence corresponding to the phosphorylation site domain of MARCKS, which also corresponds to its actin binding site, blocks PMA pot entiation of Ca2+-induced catecholamine release. The results suggest that u nder physiological conditions (i.e., nicotinic receptor stimulation) pathwa y A is the principal mechanism for the control of cortical F-actin dynamic changes. (C) 2000 Society francaise de biochimie et biologie moleculaire / Editions scientifiques ct medicales Elsevier SAS.