<p>Emerging evidence suggests brain-resident myeloid cells, including perivascular macrophages and microglia, provide a reservoir for HIV infection in the central nervous system (CNS), and their inflammatory activation is a proposed pathogenic mechanism in HIV-associated neurocognitive disorders (HAND). We investigated whether cannabinoid receptor 2 (CB₂), an immunomodulatory receptor expressed in myeloid cells, regulates viral replication and inflammation in HIV-infected macrophages and microglia. Using the synthetic CB₂-specific agonist JWH-133, we found that CB₂ activation reduced HIV replication in primary human monocyte-derived macrophages (MDMs) and human induced pluripotent stem cell-derived microglia (iMg) at differing doses, corresponding to the basal expression of <i>CNR2</i>, which encodes CB₂, and related endocannabinoid transcripts in each cell type. JWH-133 broadly reduced release of cytokines from HIV-infected MDMs but not iMg. RNA-seq revealed that CB₂ agonism primarily altered interferon and integrated stress response pathways in MDMs while altering homeostatic pathways, including synapse maintenance and phagocytosis, in iMg. Further analyses in iMg revealed that NLRP3 inflammasome activation, but not priming, was reduced by CB₂ activation, which did not inhibit HIV-induced nuclear factor kB activation. This study identifies key differences in CB₂ response between myeloid lineage cell types and implicates CB₂-specific agonists as promising candidates for the regulation of HIV-associated neuroinflammation.</p> Graphical Abstract <p>Created in BioRender. Espinoza, C. (2025) <a href="https://BioRender.com/mxfla3i">https://BioRender.com/mxfla3i</a></p> <p></p>