HIV-1 infection causes, with increasing prevalence, neurological disorders characterized in part by neuronal cell death. The HIV-1 protein Tat has been shown to be directly and indirectly neurotoxic. Here, we tested the hypothesis that a non-neurotoxic epitope of Tat can, through actions on immune cells, increase neuronal cell death. Tat(1-72) and a mutant Tat(1-72) lacking the neurotoxic epitope (Tat(Delta31-61)) concentration-dependently and markedly increased TNF-alpha production in macrophage-like differentiated human U937 and THP-1 cells, in mouse peritoneal macrophages and in mouse brain microglia. Tat(1-72) was but Tat(Delta31-61) was not neurotoxic when applied directly to neurons. Supernatants from U937 cells treated with either Tat(1-72) or Tat(Delta31-61) were neurotoxic and their immunoneutralization with an anti-TNF-alpha antibody decreased Tat(1-72)- and Tat(Delta31-61)-induced neurotoxicity. Together, these results demonstrate that the neurotoxic epitope of Tat(1-72) is different from the epitope that is indirectly neurotoxic following production of TNF-alpha from immune cells, and suggest that therapeutic interventions against TNF-alpha might be beneficial against HIV-1 associated neurological disorders.