Title Page

Contents

ABSTRACT ENGLISH 7

INTRODUCTION 9

MATERIAL AND METHODS 12

1. Experimental animals 12

2. Primary neuron culture 12

3. Oxygen-glucose deprivation model 13

4. Cell viability assay (WST assay) 13

5. Global cerebral ischemia induction 14

6. Brain tissue preparation 15

7. Measurement of neuronal death 15

8. Detection of blood-brain barrier (BBB) disruption 16

9. Immunohistochemistry 17

10. Western blot analysis 18

11. Behavior testing 18

11.1. Modified neurological severity score (mNSS). 18

11.2. Morris water maze (MWM). 19

12. Quantification 19

13. Statistical Analysis 20

RESULTS 21

1. Inhibition of PDE4B reduces neuronal cell death after global cerebral ischemia or oxygen-glucose deprivation 21

2. Inhibition of PDE4B reduced microtubule damage and oxidative injury after global cerebral ischemia. 22

3. Inhibition of PDE4B reduced glial cell activation after global cerebral ischemia 23

4. Amlexanox reduced PDE4B expression, and improved lysosomal and autophagy function 24

5. Amlexanox improved mitochondrial function and reduced apoptotic cell death 25

6. Amlexanox recovered motor and cognitive function after global cerebral ischemia 26

DISCUSSION 29

REFERENCES 50

ABSTRACT KOREAN 60

Figure 1. (A) Experimental timeline for three-day animal experiments. (B) Representative images of hippocampal neurons stained with Fluoro-Jade B (FJB)... 34

Figure 2. (A) Representative images show hippocampal microtubule damage (MAP2) in subiculum, CA1, and CA2 regions three days after GCI. AMX-treated... 36

Figure 3. (A) Representative image: Astrocyte activation (GFAP, green) and complement 3 (C3, red) in hippocampal CA1 region at 3 days after GCI. Scale bar... 38

Figure 4. (A) Experimental timeline for 24-hour animal experiments. (B) Representative image: PDE4B expression (red) in hippocampal CA1 region with... 40

Figure 5. (A) Experimental timeline: Cells cultured for 2 weeks, subjected to oxy-gen-glucose deprivation/re-oxygenation (OGD/R) one day before fixation, and... 42

Figure 6. (A) Experimental timeline: AMX administered for 13 days prior to sacrifice on the 14th day after GCI or sham operation. (B) Modified Neurological... 44

Figure 7. (A) Schematic illustrations outline the PDE4B pathway contributing to neuronal damage after global cerebral ischemia (GCI). This pathway involves... 46

Figure S1A-B. Phosphodiesterase 4B Inhibition Mitigates Blood-Brain Barrier Disruption in Global Cerebral Ischemia. 48

Figure S2A. mNSS score rate board; S2B. 30cm diameter circle maze. 49