Nerve terminals continuously maintain the synaptic vesicle cycle. Unknown mechanisms overcome the use-dependent damage of proteins. Cysteine String Protein-alpha (CSP-alpha) is a synaptic vesicle protein that prevents presynaptic neurodegeneration (Neuron 42:237-51,2004). In mice lacking CSP-alpha, we have studied hippocampal neurons in culture.

Low density autaptic cultures display a robust synaptic transmission and, after two weeks, a subtle decrease in EPSC size. Higher density cultures where synaptic contacts form between multiple neurons, display several alterations: (i) smaller size of the recycling vesicle pool (FM4-64), (ii) a progressive decrease in the density of synaptic punctae and (iii) a strikingly reduction in the size of spontaneous AMPA-mEPSCs after 30 days in culture. Our results indicate that, in conventional cultures, the absence of CSP-alpha leads to a progressive neurodegenerative phenotype similar to the "in vivo" phenotype in mice. That phenotype in culture is ideally suited to investigate the protective role of CSP-alpha against synaptic activity overloads. We are also pursuing the identification of synaptic vesicle cycle alterations that precede morphological degeneration. We have generated Thy-1 transgenic mice that, lack CSP-alpha and express synaptopHluorin, a pH-sensitive GFP that reports fluorescence changes due to exo- and endocytosis. This approach is turning out to be sensitive enough to uncover alterations in synaptic vesicle exo- and endocytosis at the neuromuscular junction that only appear after the second week of life. Support: HFSP and Spanish Ministries of Education and Science and Health (BFU2005-08130, Juan de la Cierva, FPU and BEFI).