Molecular Mechanisms Of Synapse Development &Plasticity

Child Health And Human Development

Investigators

Linked publications, trials & patents

Abstract

Neurotrophic factors are defined as secretory proteins that regulate neuronal survival and differentiation. Recent studies indicate that a major function of neurotrophic factors in the brain is to regulate synaptic transmission and plasticity. This laboratory was among the first to reveal this novel function. We demonstrated, using rodent models, that brain derived neurotrophic factor (BDNF) plays a key role in hippocampal long-term potentiation (LTP), a cellular model for learning and memory. Using the Xenopus neuromuscular synapse as a model, we identified two modes of regulation by neurotrophins: acute modulation of synaptic transmission and plasticity, and long-term alteration of the structure and function of synapses. In the period covered by this report, we have made two significant discoveries: 1) Proteolytic conversion of proBDNF to mature BDNF by the tPA/plasmin system is essential for long-term hippocampal plasticity. Long-term memory is believed to be mediated by protein synthesis-dependent, L-LTP. Two secretory proteins, tPA and BDNF, have been implicated in this process, but their relationship is unclear. We demonstrate that tPA, by activating the extracellular protease plasmin, converts the precursor proBDNF to the mature BDNF (mBDNF) in the hippocampus, and such conversion is critical for L-LTP expression. Our elctrophysilogical studies demonstrate that mBDNF, but not the cleavage-resistant proBDNF, rescues L-LTP in tPA and plasminogen knockout mice. Biochemical experiments also show that tPA, by converting plasminogen to plasmin, cleaves proBDNF to form mBDNF. In addition, genetic and pharmacological experiments reveal that mBDNF is downstream of plasmin, which is downstream of tPA, in L-LTP expression. Moreover, application of mBDNF converts early-phase LTP (E-LTP) to L-LTP and rescues L-LTP when protein synthesis is blocked by the inhibitor anisomycin. These results suggest that mBDNF is key protein synthesis product responsible for L-LTP expression. Taken together, the present study has identified tPA/plasmin as an endogenous enzyme system that converts proBDNF to mBDNF in the hippocampus, and revealed a physiological role of such conversion in the brain. Further, these results have provided a mechanistic link between these two seemingly independent molecule systems in L-LTP expression. (Science). 2) Lipid rafts mediate chemotropic guidance of nerve growth cones. We have tested the role of lipid rafts in BDNF signaling during axon growth, which requires signal transduction of extracellular cues for directional motility. We show that lipid raft disruption abolished growth cone attraction and repulsion in gradients of BDNF and netrin-1, respectively, but exerted no effects on glutamate-induced attraction. Interestingly, local raft disruption on one side of the growth cone exposed to uniform BDNF or netrin-1 produced opposite turning responses to that induced by the gradients. Lipid raft manipulation also blocked Semaphorin 3A-induced growth cone repulsion, inhibition, and collapse. Finally, we found that guidance cue-elicited MAP kinase activation depended on raft integrity and specific guidance receptors were associated with lipid rafts. These results demonstrate an essential role for lipid rafts in growth cone guidance and suggest that localized signaling through these dynamic microdomains underlies specific actions of extracellular cues on developing axons. (Neuron). 3) Activity-dependent but Ca2+-independent endocytosis in dorsal root ganglion neurons. One of the essential mechanisms is the activity-dependent endocytosis of synaptic vesicles. Synaptic vesicle endocytosis is believed to require Ca2+ and the small GTPase dynamin. We have recently identified a novel form of rapid endocytosis (RE) in dorsal root ganglion (DRG) neurons that, unlike previously described forms of endocytosis, is completely independent of Ca2+ and the GTPase dynapmin. The RE is tightly coupled to Ca2+-independent but voltage-dependent secretion (CIVDS). Using FM dye and capacitance measurements, we show that membrane depolarization induces RE in the absence of Ca2+. Inhibition of dynamin function does not affect RE. Inhibitors of protein kinase A (PKA) suppress RE induced by high-frequency depolarization, while PKA activators enhance RE induced by low-frequency depolarization. Biochemical experiments demonstrate that depolarization directly upregulates PKA activity in Ca2+-free medium. These results reveal a Ca2+- and dynamin-independent form of endocytosis, which is controlled by neuronal activity and PKA-dependent phosphorylation, in DRG neurons. (Neuron).