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Direct Rho-associated kinase inhibiton induces cofilin dephosphorylation and neurite outgrowth in PC-12 cells

Abstract

Axons fail to regenerate in the adult central nervous system (CNS) following injury. Developing strategies to promote axonal regeneration is therapeutically attractive for various CNS pathologies such as traumatic brain injury, stroke and Alzheimer’s disease. Because the RhoA pathway is involved in neurite outgrowth, Rho-associated kinases (ROCKs), downstream effectors of GTP-bound Rho, are potentially important targets for axonal repair strategies in CNS injuries. We investigated the effects and downstream mechanisms of ROCK inhibition in promoting neurite outgrowth in a PC-12 cell model. Robust neurite outgrowth (NOG) was induced by ROCK inhibitors Y-27632 and H-1152 in a time-and dose-dependent manner. Dramatic cytoskeletal reorganization was noticed upon ROCK inhibition. NOG initiated within 5 to 30 minutes followed by neurite extension between 6 and 10 hours. Neurite processes were then sustained for over 24 hours. Rapid cofilin dephosphorylation was observed within 5 minutes of Y-27632 and H-1152 treatment. Re-phosphorylation was observed by 6 hours after Y-27632 treatment, while H-1152 treatment produced sustained cofilin dephosphorylation for over 24 hours. The results suggest that ROCK-mediated dephosphorylation of cofilin plays a role in the initiation of NOG in PC-12 cells.

Abbreviations

Y-27632:

(R)-(+)-trans-N-(4-Pyridyl)-4-(1-aminoethyl)-cyclohexanecarboxamide

ROCK:

Rho-associated kinase

NGF:

nerve growth factor

NOG:

neurite outgrowth

DAG:

dorsal root ganglion

MAG:

myelin associated glycoprotein

CSPG:

chondroitin sulfate proteoglycan

OMgp:

oligodendrocyte myelin

FGF:

fibroblast growth factor

BDNF:

brain-derived neurotrophic factor

NT-3:

neurotrophin-3

PRK:

protein kinase C-related kinase

LIMK:

LIM-kinase

FITC:

Fluorescein isothiocyanate

CNS:

central nervous system

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Correspondence to Kevin K. W. Wang.

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Zhang, Z., Ottens, A.K., Larner, S.F. et al. Direct Rho-associated kinase inhibiton induces cofilin dephosphorylation and neurite outgrowth in PC-12 cells. Cell. Mol. Biol. Lett. 11, 12–29 (2006). https://doi.org/10.2478/s11658-006-0002-x

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Key words

  • Neurite outgrowth
  • ROCK
  • Y-27632
  • PC-12
  • Cofilin
  • Actin dynamics