The increased osteocyte death by oxidative stress (OS) during aging is a major cause contributing to the impairment of bone quality and bone loss. However, the underlying molecular mechanism is largely unknown. Here, we show that H2O2 induced cell death of primary osteocytes and osteocytic MLO-Y4 cells, and also caused dose-dependent decreased expression of gap junction and hemichannel-forming connexin 43 (Cx43). The decrease of Cx43 expression was also demonstrated with the treatment of other oxidants, rotenone and menadione. Antioxidant reversed the effects of oxidants on Cx43 expression and osteocyte cell death. Cx43 protein was also much lower in the osteocytes from 20-month-old as opposed to the 5-week-old or 20-week old mice. Dye transfer assay showed that H2O2 reduced the gap junction intercellular communication (GJIC). In contrast to the effect on GJIC, there was a dose-dependent increase of hemichannel function by H2O2, which was correlated with the increased cell surface expression of Cx43. Cx43(E2) antibody, an antibody that specifically blocks Cx43 hemichannel activity but not gap junctions, completely blocked dye uptake induced by H2O2 and further exacerbated H2O2-induced osteocytic cell death. In addition, knockdown of Cx43 expression by small interfering RNA (siRNA) increased the susceptibility of the cells to OS-induced death. Together, our study provides a novel cell protective mechanism mediated by osteocytic Cx43 channels against OS.