Clostridium difficile toxin B, an inhibitor of the small GTPases Rho, Rac and Cdc42, influences spiral ganglion neurite outgrowth.
OBJECTIVE
Neurotrophins and extracellular matrix (ECM) molecules are involved in neurite guidance during the development of spiral ganglion (SG) neurons. Several intracellular signaling molecules can be activated by ECMs and neurotrophins via their cognate receptors. In other systems these include the Rho small GTPases, which influence reorganization of the actin cytoskeleton that is required for axon growth. The aim of this study was to determine whether neurotrophin-3 (NT-3)-mediated SG neurite outgrowth on laminin-1 (LN) is dependent on the activation of the small GTPases Rho/Rac/Cdc42.


METHODS
SG explants from postnatal day 4 rats were cultured on LN with and without NT-3 and increasing concentrations of Clostridium difficile Toxin B, an inhibitor of Rho GTPases. After fixation and immunocytochemical labeling, neurite growth was evaluated.


RESULTS
Treatment with C. difficile Toxin B without NT-3 led to a dose-dependent decrease in the length and number of processes on LN. In contrast, C. difficile Toxin B had no significant effect on NT-3-mediated stimulation of neurite growth on LN, in terms of either number or length.


CONCLUSIONS
The results suggest that the Rho GTPases (Rho, Rac and Cdc42) are not involved in the pathways linking NT-3 signals to neurite outgrowth, but appear to be involved in LN signaling in these neurons. However, NT-3 can override or bypass LN signaling to promote neurite extension.
Brors
Dominik
D
Department of Surgery, Division of Otolaryngology, UCSD School of Medicine and VA Medical Center, La Jolla, California 92093, USA.
Aletsee
Christoph
C
Dazert
Stefan
S
Huverstuhl
Jochen
J
Ryan
Allen F
AF
Bodmer
Daniel
D
D000818:Animals; D000831:Animals, Newborn; D001426:Bacterial Proteins; D001427:Bacterial Toxins / Q000633:toxicity*; D002454:Cell Differentiation / Q000187:drug effects*; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D007797:Laminin / Q000502:physiology; D008856:Microscopy, Fluorescence; D020559:Monomeric GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology; D016501:Neurites / Q000187:drug effects*; D009473:Neuronal Plasticity / Q000187:drug effects*; D009474:Neurons / Q000166:cytology / Q000187:drug effects; D020933:Neurotrophin 3 / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013136:Spiral Ganglion / Q000166:cytology / Q000187:drug effects*; D020764:cdc42 GTP-Binding Protein / Q000037:antagonists & inhibitors* / Q000502:physiology; D020744:rac GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology; D020741:rho GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D007797:Laminin; D020933:Neurotrophin 3; C121346:laminin 1; C031815:toxB protein, Clostridium difficile; D020559:Monomeric GTP-Binding Proteins; D020764:cdc42 GTP-Binding Protein; D020744:rac GTP-Binding Proteins; D020741:rho GTP-Binding Proteins