We have recently identified, in the brain tissue of patients afflicted with Alzheimer's disease (AD), the non-A beta component of AD amyloid (NAC) as a new constituent of amyloid. NAC is derived from a larger precursor, NACP, a presynaptic protein. To better understand the role of NACP/NAC in the pathogenesis of AD, we used semiquantitative immunoblotting and combined double-immunocytochemistry/laser scanning confocal microscopy to study the concentration and distribution of NACP/NAC in human brain, and compared them to the concentration and distribution of the presynaptic marker synaptophysin and the amyloid marker A beta. The semiquantitative immunoblotting demonstrated that the NACP concentration is slightly increased in the AD frontal cortex without statistical significance, whereas synaptophysin was reduced in its levels in AD. Consequently the proportion of NACP/synaptophysin was more than double in the AD frontal cortex as compared with controls. In the AD neocortex, NACP was colocalized with approximately 80% of the synaptophysin-immunoreactive structures (presumably the presynaptic terminals) and with the dystrophic neuritic component of the plaques. Computer-aided analysis showed that numbers of NACP-immunoreactive structures along synaptophysin-immunoreactive structures were significantly diminished (30 to 40%) in AD. Although the overall numbers of NACP-positive structures were decreased, there was a significant increase in the intensity of NACP-immunoreactivity per structure in AD. This increased intensity of NACP immunoreactivity per structure in AD was not observed with anti-synaptophysin, consistent with immunoblotting-based quantification. Antibodies against NAC immunoreacted with amyloid in 35% of the diffuse plaques and 55% of the mature plaques. Normal aged control brains containing small groups of diffuse plaques were negative with anti-NAC. Double-immunolabeling studies with A beta antibodies showed that NAC immunoreactivity is more abundant in the center portion of amyloid rather than in the periphery. These studies suggest that there is a connection between metabolism of presynaptic proteins and amyloid formation, and that NAC might follow diffuse A beta accumulation resulting in the formation of compact amyloid and mature plaques.