microscopic analyses at several levels of resolution, as outlined in an earlier section. At the regional level, receptor binding studies have been used to study potential age-related changes in hippocampal GluRs in several species. While several studies have reported decreases in NMDA binding in the hippocampus of mice, rats, and monkeys (Wenk and Walker, 1991; Clark et al., 1992; Magnusson and Cotman, 1993; Le Jeune et al., 1996), other studies suggested that there is no change in NMDA receptors with aging (Nicolle et al., 1996) or perhaps even an increase in humans without Alzheimer's disease (Johnson et al., 1996).
In studying age-related changes in receptors or any of the key synaptic molecules, it is particularly important to be able to take the analysis from the regional level to that of cell classes, circuits, individual neuronal compartments, and synapses, since the changes are very likely to be cell, circuit, and synapse specific and therefore difficult to resolve at the regional level. In the case of multisubunit receptors like the NMDA and AMPA GluRs, it is optimal for the immunohistochemical data to be available at high anatomic resolution and with the highest molecular specificity, since receptor composition at the synapse is a crucial determinant of the functional characteristics of synaptic transmission.
In the same vein, it is best if shifts in GluR expression are detected at the level of GluR families (e.g., AMPA, kainate, NMDA receptors) and subunits within a family (e.g., for the AMPA receptor, GluRs1-4; for NMDA receptors, NRI and NR2A-D), given that the subunit composition profoundly impacts function (see Hollman and Heinemann, 1994, for a review). Thus, for multisubunit receptor studies in aging, the demands are particularly high in that we want the localization to: (1) have the highest level of molecular specificity, (2) be linked to identified cells and circuits, (3) be available at a quantitative level, and (4) precisely describe the receptor composition at the synaptic level. Furthermore, if the analysis is to be truly comprehensive, then a given circuit needs to be characterized beyond the receptor subunits themselves to the associated proteins that modulate receptor function (e.g., PSD-95).
We have been able to get a start on such a comprehensive analysis, although it is still in its infancy. Motivated by the importance of both the entorhinal cortex projection to the dentate gyrus and the NMDA receptor in age-related changes in memory, we investigated the GluR distribution and immunofluorescence intensity within the dentate gyrus of juvenile, adult, and aged macaque monkeys with the combined use of subunit-specific antibodies and quantitative confocal microscopy (Gazzaley et al., 1996a).
This circuit has great advantages for such an analysis because the projection from the entorhinal cortex to the dentate gyrus is strictly confined to the outer molecular layer, i.e., the distal dendrites of granule cells, whereas other excitatory inputs terminate in a nonoverlapping fashion in the inner molecular layer, the proximal dendrites (Rosene and Van Hoesen, 1987; Witter and