Effects of COX‑2 depletion on the learning and memory ability.
Firstly, it was observed that after genotyping COX-2KO
(COX-2-/-) mice by PCR (Fig. 1A), no COX-2 protein was
detected by Western blot (Fig. 1B).Then a comparison between WT and
COX-2KO mice revealed no differences in the body weights, body
temperatures, or basal inflammation (TNF-a, iNOS, and IL-1β) in the
hippocampus (Fig.1C-E). Furthermore, to determine whether COX-2
contributes to learning and memory ability, we used Morris water maze
(MWM) memory test and novel object recognition task (NORT). In the MWM
test, the results showed that the escape latency of mice in each group
decreased with the increase in training times, indicating that the
training of mice was effective. Compared with WT group, the escape
latency at training day 4 and 5 were significantly shorter for COX-2
knocked out mice (Fig.1C). Furthermore, to measure spatial learning and
memory retention, we next performed probe trials on the 6th day. In the
comparison with WT mice, COX-2KO ones failed to find the target platform
with less time spending (Fig.1D, F) and fewer platform crossings
(Fig.1E) in the target quadrant. In addition, no differences were
observed between WT and COX-2KO mice in terms of swimming speed (Fig.1I)
and total distance traveled (Fig.1J).Next, we used NORT to assess the
recognition memory of different groups. As illustrated in Figure 1G, the
recognition index of COX-2KO mice were reduced compared to the WT group
at the test of both 1h and 24h. Moreover, the total distance traveled by
COX-2KO and WT mice in the NORT (Fig.1M) was also not different. These
results suggested that COX-2KO mice were impaired in learning and memory
ability.
Effects
of COX‑2 depletion on the SYP and PSD95.
Then we detected synaptophysin (SYP), PSD95 by double immunofluorescence
and Western blot. We stained PSD95 with red and SYP with green in the
hippocampus CA1 (Fig. 2A). We found there was almost no change in the
fluorescence intensity of SYP between WT and COX-2KO those two groups,
however, compared with WT group, the expression of PSD95 was almost
absent in COX-2KO samples with a faint green staining (Fig. 2B).
Consistent with the immunofluorescence result, we found COX-2 depletion
could significantly downregulate the expression of PSD95 protein but not
SYP in the hippocampus (Fig. 2C). These results indicated that COX-2KO
could influence the expression of PSD95 instead of SYP in the brain.