Co-Signaling by Neurotrophic Factors and the Extracellular Matrix for Axonal Growth and Neuronal Survival

Neurotrophic factors (NTFs) and the extracellular matrix (ECM) are important regulators of axonal growth and neuronal survival in mammalian nervous system. Understanding of the mechanisms of this regulation is crucial for the development of posttraumatic therapies and drug intervention in the injured nervous system. NTFs act as soluble, target-derived extracellular regulatory molecules for a wide range of physiological functions including axonal guidance and the regulation of programmed cell death in the nervous system. The ECM determines cell adhesion and regulates multiple physiological functions via short range cell-matrix interactions. The present work focuses on the mechanisms of the action of NTFs and the ECM on axonal growth and survival of cultured sensory neurons from dorsal root ganglia (DRG). We first examined signaling mechanisms of the action of the glial cell line-derived neurotrophic factor (GDNF) family ligands (GFLs) on axonal growth. GDNF, neurturin (NRTN) and artemin (ART) but not persephin (PSPN) promoted axonal initiation in cultured DRG neurons from young adult mice. This effect required Src family kinase (SFK) activity. In neurons from GFRalpha2-deficient mice, NRTN did not significantly promote axonal initiation. GDNF and NRTN induced extensive lamellipodia formation on neuronal somata and growth cones. This study suggested that GDNF, NRTN and ARTN may serve as stimulators of nerve regeneration under posttraumatic conditions. Consequently we studied the convergence of signaling pathways induced by NTFs and the ECM molecule laminin in the intracellular signaling network that regulates axonal growth. We demonstrated that co-stimulation of DRG neurons with NTFs (GDNF, NRTN or nerve growth factor (NGF)) and laminin leads to axonal growth that requires activation of SFKs. A different, SFK-independent signaling pathway evoked axonal growth on laminin in the absence of the NTFs. In contrast, axonal branching was regulated by SFKs both in the presence and in the absence of NGF. We proposed and experimentally verified a Boolean model of the signaling network triggered by NTFs and laminin. Our results put forward an approach for predictable, Boolean logics-driven pharmacological manipulation of a complex signaling network. Finally we found that N-syndecan, the receptor for the ECM component HB-GAM was required for the survival of neonatal sensory neurons in vitro. We demonstrated massive cell death of cultured DRG neurons from mice deficient in the N-syndecan gene as compared to wild type controls. Importantly, this cell death could not be prevented by NGF the neurotrophin which activates multiple anti-apoptotic cascades in DRG neurons. The survival deficit was observed during first postnatal week. By contrast, DRG neurons from young adult N-syndecan knock-out mice exhibited normal survival. This study identifies a completely new syndecan-dependent type of signaling that regulates cell death in neurons.