Membrane depolarization inhibits spiral ganglion neurite growth via activation of multiple types of voltage sensitive calcium channels and calpain.
The effect of membrane electrical activity on spiral ganglion neuron (SGN) neurite growth remains unknown despite its relevance to cochlear implant technology. We demonstrate that membrane depolarization delays the initial formation and inhibits the subsequent extension of cultured SGN neurites. This inhibition depends directly on the level of depolarization with higher levels of depolarization causing retraction of existing neurites. Cultured SGNs express subunits for L-type, N-type, and P/Q type voltage-gated calcium channels (VGCCs) and removal of extracellular Ca(2+) or treatment with a combination of L-type, N-type, and P/Q-type VGCC antagonists rescues SGN neurite growth under depolarizing conditions. By measuring the fluorescence intensity of SGNs loaded with the fluorogenic calpain substrate t-butoxy carbonyl-Leu-Met-chloromethylaminocoumarin (20 microM), we demonstrate that depolarization activates calpains. Calpeptin (15 microM), a calpain inhibitor, prevents calpain activation by depolarization and rescues neurite growth in depolarized SGNs suggesting that calpain activation contributes to the inhibition of neurite growth by depolarization.
Department of Otolaryngology-Head and Neck Surgery, University of Iowa, Iowa City, IA 52242, USA.
Pamela C
PC
Roehm
Ningyong
N
Xu
Erika A
EA
Woodson
Steven H
SH
Green
Marlan R
MR
Hansen
D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000378:metabolism; D020864:Calcium Channels, N-Type / Q000187:drug effects / Q000378:metabolism; D020907:Calcium Channels, Q-Type / Q000187:drug effects / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016501:Neurites / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D013136:Spiral Ganglion / Q000187:drug effects / Q000254:growth & development* / Q000378:metabolism*; D014162:Transfection / Q000379:methods
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D020864:Calcium Channels, N-Type; D020907:Calcium Channels, Q-Type; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D002154:Calpain; D002118:Calcium