The advent of Human Immunodeficiency Virus (HIV) antiretrovirals have reduced the severity of HIV related neurological comorbidities but they nevertheless remain prevalent. Synaptic degeneration due to the action of several viral factors released from infected brain myeloid and glia cells and inflammatory cytokines has been attributed to the manifestation of a range of cognitive and behavioral deficits. The contributions of specific pro-inflammatory factors and their interplay with viral factors in the setting of treatment and persistence are incompletely understood. Exposure of neurons to chemokine receptor-4(CXCR4)-tropic HIV-1 envelope glycoprotein (Env) can lead to post-synaptic degradation of dendritic spines. The contribution of members of the extracellular matrix (ECM) and specifically, of perineuronal nets (PNN) toward synaptic degeneration, is not fully known, even though these structures are found to be disrupted in post-mortem HIV-infected brains. Osteopontin (Opn, gene name SPP1 ), a cytokine-like protein, is found in abundance in the HIV-infected brain. In this study, we investigated the role of Opn and its ECM integrin receptors, β 1- and β 3 integrin in modifying neuronal synaptic sculpting. We found that in hippocampal neurons incubated with HIV-1 Env protein and recombinant Opn, post-synaptic-95 (PSD-95) puncta were significantly increased and distributed to dendritic spines when compared to Env-only treated neurons. This effect was mediated through β 3 integrin, as silencing of this receptor abrogated the increase in post-synaptic spines. Silencing of β 1 integrin, however, did not block the increase of post-synaptic spines in hippocampal cultures treated with Opn. However, a decrease in the PNN to β III-tubulin ratio was found, indicating an increased capacity to support spine growth. From these results, we conclude that one of the mechanisms by which Opn counters the damaging impact of the HIV Env protein on hippocampal post-synaptic plasticity is through complex interactions between Opn and components of the ECM which activate downstream protective signaling pathways that help maintain the potential for effective post-synaptic plasticity.