Carbenoxolone and 18β-glycyrrhetinic acid inhibit inositol 1,4,5-trisphosphate-mediated endothelial cell calcium signalling and depolarise mitochondria

Background and Purpose: Coordinated endothelial control of cardiovascular function is proposed to occur by endothelial cell communication via gap junctions and connexins. To study intercellular communication, the pharmacological agents carbenoxolone (CBX) and 18β-glycyrrhetinic acid (18βGA) are used widely as connexin inhibitors and gap junction blockers. Experimental Approach: We investigated the effects of CBX and 18βGA on intercellular Ca ²⁺ waves, evoked by inositol 1,4,5-trisphosphate (IP ₃) in the endothelium of intact mesenteric resistance arteries. Key Results: Acetycholine-evoked IP ₃-mediated Ca ²⁺ release and propagated waves were inhibited by CBX (100 μM) and 18βGA (40 μM). Unexpectedly, the Ca ²⁺ signals were inhibited uniformly in all cells, suggesting that CBX and 18βGA reduced Ca ²⁺ release. Localised photolysis of caged IP ₃ (cIP ₃) was used to provide precise spatiotemporal control of site of cell activation. Local cIP ₃ photolysis generated reproducible Ca ²⁺ increases and Ca ²⁺ waves that propagated across cells distant to the photolysis site. CBX and 18βGA each blocked Ca ²⁺ waves in a time-dependent manner by inhibiting the initiating IP ₃-evoked Ca ²⁺ release event rather than block of gap junctions. This effect was reversed on drug washout and was unaffected by small or intermediate K ⁺-channel blockers. Furthermore, CBX and 18βGA each rapidly and reversibly collapsed the mitochondrial membrane potential. Conclusion and Implications: CBX and 18βGA inhibit IP ₃-mediated Ca ²⁺ release and depolarise the mitochondrial membrane potential. These results suggest that CBX and 18βGA may block cell–cell communication by acting at sites that are unrelated to gap junctions.