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Transecting the axons of neurons in the adult superior cervical ganglion (SCG; axotomy) results in the survival of most postganglionic neurons, the influx of circulating monocytes, proliferation of satellite cells, and changes in neuronal gene expression. In contrast, transecting the afferent input to the SCG (decentralization) results in nerve terminal degeneration and elicits a different pattern of gene expression. We examined the effects of decentralization on macrophages in the SCG and compared the results to those previously obtained after axotomy. Monoclonal antibodies were used to identify infiltrating (ED1+) and resident (ED2+) macrophages, as well as macrophages expressing MHC class II molecules (OX6+). Normal ganglia contained ED2+ cells and OX6+ cells, but few infiltrating macrophages. After decentralization, the number of infiltrating ED1+ cells increased in the SCG to a density about twofold greater than that previously seen after axotomy. Both the densities of ED2+ and OX6+ cells were essentially unchanged after decentralization, though a large increase in OX6+ cells occurred after axotomy. Proliferation among the ganglion's total non-neuronal cell population was examined and found to increase about twofold after decentralization and about fourfold after axotomy. Double-labeling experiments indicated that some of these proliferating cells were macrophages. After both surgical procedures, the percentage of proliferating ED2+ macrophages increased, while neither procedure altered the proliferation of ED1+ macrophages. Axotomy, though not decentralization, increased the proliferation of OX6+ cells. Future studies must address what role(s) infiltrating and/or resident macrophages play in regions of decentralized and axotomized neurons and, if both are involved, whether they play distinct roles., (Copyright 2002 Wiley Periodicals, Inc.)

Following peripheral nerve transection, a series of biochemical changes occurs in axons and Schwann cells both at the site of the lesion and distal to it. Macrophages differentiated from monocytes that invade the area in response to transection (elicited macrophages) and, perhaps, also macrophages normally present in the tissue (resident macrophages) play important roles in these changes. In addition, nerve transection produces changes in the cell bodies of axotomized neurons and their surrounding glial cells, located at some distance from the lesion. To determine whether macrophages might play a role in the changes occurring in the superior cervical ganglion (SCG) after axotomy, we examined the presence of macrophages before and after axonal damage. The monoclonal antibodies ED1, ED2, and OX6 were used, each of which recognizes a somewhat different population of macrophages. Ganglia from normal rats contained a population of resident cells that were ED2+ but very few that were ED1+. Within 2 days after the post-ganglionic nerves were transected, the number of ED1+ cells increased substantially, with little change in immunostaining for ED2. These data, in combination with published studies on other tissues, suggest that ED1 in the SCG is selective for elicited macrophages and ED2 for resident macrophages. OX6 immunostaining was prominent in normal ganglia but also increased significantly after axotomy, suggesting that it reflects both macrophage populations. Systemic administration of 6-hydroxydopamine, a neurotoxin that causes the destruction of sympathetic nerve endings, also produced an increase in ED1 immunostaining. Thus, the change in ED1 immunostaining in the SCG does not require surgery, with the attendant severing of local blood vessels and connective tissue, but rather only the disconnection of sympathetic neurons from their end organs. The time course of the invasion of monocytes after axotomy indicates that this process is not required to trigger the biochemical changes occurring in the ganglion within the first 24 h. On the other hand, the existence of a resident population of macrophages raises the possibility that changes in those cells might be involved.

Agrin induces discrete high-density patches of acetylcholine receptors (AChRs) and other synaptic components on cultured myotubes in a manner that resembles synaptic differentiation. Furthermore, agrin-like molecules are present at developing neuromuscular junctions in vivo. This provides us with a unique opportunity to manipulate AChR patching in order to examine the role of cytoskeletal components. Cultured chick myotubes were fixed and labeled to visualize the distributions of actin, alpha-actinin, filamin, tropomyosin, and vinculin. Overnight exposure to agrin caused a small amount of alpha-actinin, filamin, and vinculin to reorganize into discrete clusters. Double-labeling studies revealed that 78% of the AChR clusters were associated with detectable concentrations of filamin, 70% with alpha-actinin, and 58% with vinculin. Filamin even showed congruence to AChRs within clustered regions. By contrast, actin (visualized with fluorescein-phalloidin) and tropomyosin did not show specific associations with agrin-induced AChR clusters. The accumulation of cytoskeletal components at AChRs clusters raised the possibility that cytoskeletal rearrangements direct AChR clustering. However, a time course of agrin-induced clustering that focused on filamin revealed that most of the early AChR clusters (3-6 h) were not associated with detectable amounts of cytoskeletal material. The accumulation of cytoskeletal material at later times (12-18 h) may imply a role in maintenance and stabilization, but it appears unlikely that these cytoskeletal elements initiate AChR clustering on myotubes.