All lentiviruses infect the brain, causing chronic neurological disease in their respective hosts. To examine the relationship(s) between lentivirus molecular diversity and the development of neurological disease, we examined in vitro and in vivo models of lentivirus neurovirulence using different recombinant viruses derived from human (HIV-1) and feline (FIV) immunodeficiency viruses. Both in vitro and in vivo studies of FIV neurovirulence showed that the FIV envelope derived from a neurovirulent strain was a principal determinant of neuropathogenesis, although systemic immunosuppression was also an integral feature of FIV neurovirulence. Studies of HIV-1 envelope sequences derived from brain or blood indicate that molecular diversity is greater in viruses from patients with HIV-associated dementia (HAD), compared to nondemented individuals. Moreover, the hypervariable V3 domain of HIVgp120, regardless of the HIV-1 clade from which it was derived, was an important region for mediating neurotoxicity in vitro but the level of viral replication did not influence neurotoxicity. For both the HIV-1 and FIV envelopes and HIV-1 Tat, induction of matrix metalloproteinase (MMP)-2 in macrophages was a consistent finding. Neurotoxicity caused by supernatants from HIV-infected or transfected macrophages, containing MMP-2, was greater than direct neurotoxicity levels caused by direct exposure of neurons to virus in assays of total neuronal death, but not in assays of neuronal apoptosis. Proteinase-activated receptor (PAR)-1 and its ligand thrombin were also induced during HIV infection, chiefly on astrocytes. PAR-1 activation resulted in gliosis and neurobehavioral changes in an animal model and resulted in N-methyl-D-aspartate (NMDA) receptor-mediated neuronal death. These findings suggest that the lentivirus envelope, which is a domain of extensive molecular diversity in brain-derived lentivirus isolates, directly influences neuropathogenesis through the activation of select proteases, underscoring the importance of concentrating on individual viral genes and proteases in the development of neuroprotective agents for HIV-related neurological disease.