Despite advances in antiviral therapy, HIV-1 associated neurological disorders (HAND) have significantly increased in incidence, particularly in hosts who engage in substance abuse. Cocaine is a commonly abused drug and strongly implicated in HIV-1 infection and neuropathogenesis. However, the molecular basis of these pathological changes in the central nervous system (CNS) is still not yet fully understood. Recently, extracellular vesicles/exosomes (EVs) have been implicated in HIV-1 pathogenesis and elicitation of neuroinflammation. Based on our preliminary studies, we hypothesize that HIV-1 and cocaine enhance neuropathogenesis in HIV-1 infected illicit drug using individuals by altering the molecular cargo and release of EVs from HIV-1 infected immune cells. Such EVs facilitate the transfer of inflammatory mediators and viral components to the CNS by subverting the integrity of the blood brain barrier (BBB). The overall objective of this proposal is to characterize the molecular components of large EVs (lEVs) derived from HIV-1 infected and cocaine treated monocyte-derived macrophages, and then study the mechanisms involved in cocaine mediated enhanced transcytosis and inflammatory effects of these EVs on the BBB. Specific points of innovation include: (i) utilize cutting edge methodologies to characterize the lEVs derived from HIV-1 infected and/or cocaine treated monocyte-derived macrophages, (ii) analyze the biological effects of these lEVs on human brain microvascular endothelial cells (HBMECs), microglial cells and astrocytes, (iii) explore molecular mechanisms involved in cocaine mediated enhanced uptake of lEVs by HBMECs , (iv) study the effects of lEVs derived from HIV-1 infected macrophages on BBB, in the presence or absence of cocaine, (v) characterize how cocaine induced increased expression of miR-34a in lEVs enhances inflammation in HBMECs, (vi) elucidate whether so-called “ Trojan EVs”, that possess both retroviral and host components, can transmit HIV-1 infection to microglia and astrocytes. We set out three specific aims to: (1) Study the effects of cocaine on large EVs (lEVs) derived from HIV-1 infected monocyte-derived macrophages, (2) Explore the effects of these lEVs on a HBMEC monolayer, in the presence or absence of cocaine, (3) Analyze the effects of cocaine on lEV induced inflammation and HIV-1 infection in microglial cells and astrocytes. By deciphering these molecular mechanisms involving EVs, we hope to provide a framework for novel strategies to more effectively combat virus spread and development of HAND in the high risk population of cocaine users.