Fig. 5.

Electron microscopic visualization of EAAT2-protein in rat hippocampal slices. (A) Labeling was observed in glial (g) membrane (large arrowheads point to two of the gold particles) and also over nerve terminal (t) membranes (e.g. arrows). No significant labeling was found on the membranes of dendritic spines (s). Particles (e.g. small arrowheads) are counted as unattributable as they are closer than 40 nm to two different types of membranes. About 10% of the gold particles were classified as unattributable (Table 1). (B) Gold particles (arrows) were observed in axonal (a) membranes just prior to the axon terminal (t). (C) Glial membranes (g) were heavily labeled, while nerve terminals were weakly, but indisputably labeled. Some of the labeling of terminals was intracellular (asterisk), but this is likely to mostly represent unwanted reactivity of the antibodies as some labeling of terminal cytoplasm was also observed in −/− mice (see text). The arrow points to one of the only six gold particles (out of a total of 1532+741, Table 1) found in unambiguous association with identifiable spine (s) membranes. Although there are some unattributable particles, we believe that most of the attributable labeling is due to glia and to axons because the vast majority of identifiable spines were unlabeled. The one shown in A is typical (see also Results). (D) Labeling of the terminal membranes was also obtained with antibodies specific to the A-variant (the predominant form) of EAAT2. Antibodies: A—C, anti-73 kDa (AB#171) at 5 μg/ml; D anti-B563 (AB#355) at 1 μg/ml. Scale bars=200 nm.