Attractive axon guidance involves asymmetric membrane transport and exocytosis in the growth cone

Abstract

Asymmetric elevation of the Ca(2+) concentration in the growth cone can mediate both attractive and repulsive axon guidance. Ca(2+) signals that are accompanied by Ca(2+)-induced Ca(2+) release (CICR) trigger attraction, whereas Ca(2+) signals that are not accompanied by CICR trigger repulsion. The molecular machinery downstream of Ca(2+) signals, however, remains largely unknown. Here we report that asymmetric membrane trafficking mediates growth cone attraction. Local photolysis of caged Ca(2+), together with CICR, on one side of the growth cone of a chick dorsal root ganglion neuron facilitated the microtubule-dependent centrifugal transport of vesicles towards the leading edge and their subsequent vesicle-associated membrane-protein 2 (VAMP2)-mediated exocytosis on the side with an elevated Ca(2+) concentration. In contrast, Ca(2+) signals without CICR had no effect on the vesicle transport. Furthermore, pharmacological inhibition of VAMP2-mediated exocytosis prevented growth cone attraction, but not repulsion. These results strongly suggest that growth cone attraction and repulsion are driven by distinct mechanisms, rather than using the same molecular machinery with opposing polarities.

Calcium signaling in axon guidance

“…Tojima et al [18] identified enhanced vesicle-associated membrane protein (VAMP) 2-mediated exocytosis in response to attractive Ca 2+ signals elicited by FLIP of caged Ca 2+ . Furthermore, inhibition of VAMP2-mediated exocytosis by tetanus neurotoxin (TeNT) abolished attraction [18].…”

“…Tojima et al [18] identified enhanced vesicle-associated membrane protein (VAMP) 2-mediated exocytosis in response to attractive Ca 2+ signals elicited by FLIP of caged Ca 2+ . Furthermore, inhibition of VAMP2-mediated exocytosis by tetanus neurotoxin (TeNT) abolished attraction [18]. Interestingly, TeNT did not stop axon growth, suggesting that axon outgrowth mediation is independent of VAMP2 and can be regulated distinctly from attraction [18].…”

“…Furthermore, inhibition of VAMP2-mediated exocytosis by tetanus neurotoxin (TeNT) abolished attraction [18]. Interestingly, TeNT did not stop axon growth, suggesting that axon outgrowth mediation is independent of VAMP2 and can be regulated distinctly from attraction [18]. VAMP2-mediated exocytosis is not only necessary but also is sufficient for attraction [17].…”

“…A calmodulin-dependent protein kinase II (CaMKII)-calcineurin (CaN) switch often initiates either an attractive (turning towards the highest guidance cue concentration) or repulsive (turning away from the highest guidance cue concentration) response to these intracellular calcium elevations [8]. These responses are effected via the regulation of cytoskeletal components, such as microtubules and actin filaments [11][12][13][14][15], and by membrane dynamics, including vesicle trafficking [16][17][18]. In the past few years many new insights have emerged regarding the central and multifaceted roles of calcium in mediating axon growth and guidance.…”

Role of Glycine Receptors in Glycine-Induced LTD in Hippocampal CA1 Pyramidal Neurons

“…To investigate whether the Gly-LTD in AMPAR-mediated EPSCs was due to postsynaptic AMPAR internalization, we loaded cells with a specific dynamin-dependent endocytosis inhibitor, D15 (2 mM), which specifically interferes with the binding of dynamin to amphiphysin (Luscher et al, 1999;Morishita et al, 2005). We were then able to examine whether Gly-LTD could be induced under these conditions (Kang et al, 2005;Tojima et al, 2007). We found that there were no obvious changes in EPSCs following D15 treatment (0.99±0.04, n ¼ 7, compared with baseline, paired-samples t-test, P40.05; Figure 6b and d).…”