Probenecid protects against cerebral ischemia/reperfusion injury by inhibiting lysosomal and inflammatory damage in rats

- Probenecid protects against cerebral I/R injury when given before reperfusion.
- Probenecid also has a protective effect when given at 2 h after reperfusion, but not at 6 h.
- The protective effect regained for continuous probenecid treatment, even first dose given at 6 h after reperfusion.
- Probenecid pre-treatment inhibits the calpain-cathepsin pathway after I/R injury.
- The inflammatory reaction after I/R injury is also suppressed by continuous probenecid given for 7 days.

Probenecid has been used for decades to treat gout, and recent studies have revealed it is also a specific inhibitor of the pannexin-1 channel. It has been reported that the pannexin-1 channel is involved in ischemic injury. Here, we investigated the neuroprotective effect and the possible mechanisms of action of probenecid in global cerebral ischemia/reperfusion (I/R) injury in rats. Twenty minutes of transient global cerebral I/R injury was induced using the four-vessel occlusion (4-VO) method in male Sprague-Dawley rats. Different doses of probenecid were administered intravenously, intraperitoneally, or by gavage before or after reperfusion. Probenecid via all three routes protected against CA1 neuronal death when given before reperfusion. This protective effect continued when probenecid was given at 2 h after reperfusion, but not at 6 h. Interestingly, the protective effect regained if probenecid was given continuously for 7 days after reperfusion. The release of cathepsin B and overexpression of calpain-1 after reperfusion were inhibited, while the upregulation of Hsp70 was strengthened by probenecid pre-treatment. Furthermore, the activation and proliferation of microglia and astrocytes after I/R injury were suppressed by continuous given for 7 days, but only partly by a single dose at 6 h of reperfusion. Thus, our data indicate that probenecid protects against transient global cerebral I/R injury probably by inhibiting calpain-cathepsin pathway and the inflammatory reaction.