FIGURE 4.

Bystander apoptosis induced by HIV-infected astrocytes microinjected in mice brain involves IP₃Rs and Ca²⁺ release. Human U87-CD4-CCR5 cells infected with HIV_ADA (50 ng/ml) were microinjected into the cortices of Cx43^fl/fl as well as hGFAP-cre Cx43^fl/fl mice, and simultaneously either IP₃R blocker, XeC (10 μM), or intracellular Ca²⁺ chelator, BAPTA-AM (100 μM), was administered to assess Ca²⁺ dependency. After 48 h, animals were sacrificed, and the brain sections were analyzed for apoptosis by TUNEL assay. (A,B) Bystander apoptosis induced by microinjected HIV-infected cells in Cx43^fl/fl mice involved IP₃R-mediated Ca²⁺ release, since inhibition of IP₃R activation by XeC prevented bystander toxicity as assessed by percentage of apoptotic cells (A) as well as by radius of bystander apoptosis from the site of microinjection (B). (C,D) Administration of BAPTA-AM along with microinjection of HIV-infected cells in Cx43^fl/fl mice also averted toxicity by regulating [Ca²⁺]_i levels leading to lesser percentage of apoptotic cells (C) and smaller radius of bystander apoptosis (D) as compared to control condition. However, in case of hGFAP-cre Cx43^fl/fl mice, use of either XeC (A,B) or BAPTA-AM (C,D) had no significant effect as these animals did not exhibit bystander apoptosis due to lack of Cx43 GJs and HCs. ^∗∗p < 0.005; n.s. – not significant.