Improvement of the basic knowledge to better determine the biomechanisms at work when using this synaptic assay.
Development of specific software and hardware to improve and automatize the quantitation of the fluorescent signals.
This method will be further extended by adding the simultaneous quantitation of a third fluorescent signal, either a specific synaptic antigen or some other functional fluorescent probe.
Standardisation of the assay and development of novel tools such as specific monoclonal antibodies and fluorescent derivatives.
The assay will be used to test the effects of various known drugs and toxicants, including free radicals generators, in physiological and pathological conditions (such as ischemic and/or excitotoxic conditions) to determine how much of the toxic or protective potentials can be predicted by this novel test.
A series of transgenic mice, where neuronal functioning has been impaired by genetic manipulation, will be generated and tested. The effects of these genetic manipulations on neuronal response to toxic insult will be analysed, and the ability of this synaptic assay to predict this susceptibility determined.
Progress in the field of brain pharmacology has been severely limited by the complexity of the underlying processes and in many cases by the lack of adequate animal models. For example, at the moment no simple test exists to check for possible actions of drugs and toxicants on complex brain functions such as learning and memory and this strongly reduces our therapeutic potential when treating neurodegenerative diseases and brain ischemic insults, where these functions are strongly disrupted. The task of this project is to validate a novel methodology that could be used as a screening assay for drugs and toxicants action on synaptic communication and plasticity mechanisms, the cellular mechanisms that underlie many of these complex behaviors. After validation, this simple test could be possibly used as a predictive model for drugs and toxicants effects on cognitive functions. This methodology uses specific tools that have been already developed or that are going to be developed, and an advanced culturing technology to produce primary neuronal cultures from different brain regions. It allows the direct visualization of early changes in the level of synaptic activity that could be induced by exogenously added compounds. Thus, the positive or negative modulation of synaptic plasticity can be studied with great sensitivity and spatial resolution. As this assay is very sensitive, simple and inexpensive, it is believed that it could easily became a standard test for drug companies interested in testing drugs and toxicants for specific effects on synaptic communication and synaptic plasticity mechanisms.
Funding SchemeCSC - Cost-sharing contracts