Recent studies have shown that synaptic dysfunction precedes neuronal cell death in the early stages of dementia accompanied by neurodegenerative diseases. Synaptic dysfunction is presumed as a decrease in the number of dendritic spines in neurons of the cerebral cortex and hippocampus, which are essential for learning and memory. Therefore, the risk for impairment of learning and memory can be assessed by decreased number of dendritic spines. Dendritic spines are small actin-rich projections protruding from the dendrites of neurons that form excitatory synapses in the cortex and hippocampus. Drebrin is an actin-binding protein that localizes to dendritic spines in mature neurons and plays a specific role in spine formation. Drebrin is known to decrease in Alzheimer's disease with a high correlation to symptom stage. In low-density cultures of hippocampal neurons, the number of dendritic spines can be counted as the number of drebrin clusters with immunostaining. The protocol for high-throughput imaging analysis of drebrin clusters has been developed and shown to be useful for screening chemicals that bind to the NMDA receptor. In fact, we have examined the toxicity of phencyclidine (PCP) and PCP-analogues and published results in a previous paper. We have developed not only the immunocytochemical protocol for in vitro assay using neuronal culture but also enzyme-linked immunosorbent assay (ELISA) kits to evaluate drebrin protein levels. Thus, decreased number of dendritic spines induced by chemicals can be assessed quantitatively as a loss of drebrin immunocytochemically and biochemically. Drebrin deficiency is directly related to synaptic dysfunction and leads to the impairment of learning and memory, even in the absence of neuronal cell death.