2.1.3. Findings by Fluoro-Jade B (FJB) Histofluorescence

In the vehicle-sham and ATX-sham groups, no FJB-stained cells (FJB-cells), which are degenerating or dead cells, were detected in CA1-3 (Figure 3(Aa,Ae,Ca,Ce)). In the vehicle-IR group, FJB-pyramidal cells in CA1-3 were observed from two days after IR (Figure 3(Ab,Ac,B,Cb,Cc,D)), and FJB-pyramidal cells of CA1-3 were dramatically increased at five days after IR; the number of FJB-pyramidal cells in CA1 and CA2/3 was 63 and 67 cells/300 μm2, respectively (Figure 3(Ad,B,Cd,D)). In the ATX-IR group, FJB-pyramidal cells in CA1-3 were not found until two days after IR (Figure 3(Af,Ag,B,Cf,Cg,D)). At five days after IR, many FJB-pyramidal cells were detected in CA1 and CA2/3 (26 and 13 cells/300 μm2, respectively), but the number was significantly low (41% in CA1 and 19% in CA2/3 of vehicle-IR group) when compared with the vehicle-IR group (Figure 3(Ah,B,Ch,D)). This finding means that the pyramidal cells in CA1-3 were lost (dead) after severe IR, and ATX could attenuate the death of pyramidal cells induced by severe IR.

**Figure 3.** (**A**,**C**) Representative photographs of FJB staining in CA1 (**A**) and CA2/3 (**C**) of the vehicletreated (left column) and ATX-treated (right column) groups in sham (**Aa**,**Ca**,**Ae**,**Ce**), and on day 1 (**Ab**,**Cb**,**Af**,**Cf**), day 2 (**Ac**,**Cc**,**Ag**,**Cg**), and day 5 (**Ad**,**Cd**,**Ah**,**Ch**) after IR. In the vehicle-IR group, FJB-pyramidal cells are detected in the SP of CA1-3 from two days after IR, and, at five days after IR, FJB-pyramidal cells (white asterisks in (**Ad**,**Cd**)) are significantly increased. However, in the ATX-IR group, decreased FJB-pyramidal cells (arrows in (**Ah**,**Ch**)) are observed in CA1-3 only at five days after IR. Scale bar = 60 μm; (**B**,**D**) Mean numbers of FJB- pyramidal cells in CA1 (**B**) and CA2/3 (**D**). The bars indicate the means ± SEM (*n* = 7, respectively; \* *p* < 0.05 vs. corresponding sham group, # *p* < 0.05 vs. vehicle-IR group).
