Acetylcholinesterase inhibitor treatment delays recovery from axotomy in cultured dorsal root ganglion neurons.
We have previously reported that dorsal root ganglion neurons cultured in the presence of the highly specific, reversible acetylcholinesterase inhibitor 1,5-bis-(4-allyldimethylammoniumphenyl) pentan-3-one dibromide (BW284c51), showed significantly reduced neurite outgrowth and contained massive perikaryal inclusions of neurofilaments. In the present report we have more closely examined these changes in a time course study over a 21-day culture period using a combined morphological, immunocytochemical and enzymatic approach and additionally, describe, the effects of acetylcholinesterase inhibitor treatment on the state of neurofilament phosphorylation. Finally, we have examined the effects of co-administration of N6,2'-0-dibutyryladenosine 3':5'-cyclic monophosphate (dbcAMP) with BW284c51. At 1 day in culture, both control and treated cells displayed eccentrically located nuclei, numerous polysomes and perikaryal accumulations of neurofilaments which were immunoreactive with both phosphorylation- and nonphosphorylation-dependent neurofilament antibodies. These cytological changes, which are common features of the chromatolytic reaction following axotomy in vivo, rapidly resolved in the control neurons, where by 7 days in culture, the neurofilament accumulations had completely disappeared and neurite outgrowth was robust. In contrast, inhibitor-treated neurons retained the post-axotomy features up to 21 days and had significantly reduced neurite outgrowth. In addition, we have investigated a possible role of cyclic adenosine monophosphate (cAMP) in the recovery process since it has been shown to enhance neuritic outgrowth in cultured neurons. Our results demonstrate that the addition of dbcAMP, a membrane permeable analog of cAMP, significantly enhanced neuritic outgrowth and accelerated the recovery of BW284c51-treated dorsal root ganglion cells, as gauged by the disappearance of the axotomy-related cytological changes. Treatment with dbcAMP also increased acetylcholinesterase activity which has been positively correlated with neurite outgrowth both in vivo and in vitro. Together, these observations suggest that acetylcholinesterase has a non-cholinolytic, neurotrophic role in neuronal regeneration and development.
Department of Anatomy, Virginia Commonwealth University, Richmond 23298-0709, USA.
J L
JL
Dupree
J W
JW
Bigbee
D000110:Acetylcholinesterase / Q000378:metabolism*; D000818:Animals; D001369:Axons / Q000502:physiology*; D002093:Benzenaminium, 4,4'-(3-oxo-1,5-pentanediyl)bis(N,N-dimethyl-N-2-propenyl-), Dibromide / Q000494:pharmacology*; D003994:Bucladesine; D002467:Cell Nucleus / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D002478:Cells, Cultured; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D004622:Embryo, Mammalian; D005455:Fluorescent Antibody Technique; D005727:Ganglia, Spinal / Q000166:cytology / Q000502:physiology*; D007150:Immunohistochemistry; D007700:Kinetics; D008854:Microscopy, Electron; D016501:Neurites / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D016900:Neurofilament Proteins / Q000032:analysis / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D051381:Rats
D002800:Cholinesterase Inhibitors; D016900:Neurofilament Proteins; D002093:Benzenaminium, 4,4'-(3-oxo-1,5-pentanediyl)bis(N,N-dimethyl-N-2-propenyl-), Dibromide; D003994:Bucladesine; D000110:Acetylcholinesterase