The ?-opioid receptor (MOR) is known to undergo extensive alternative splicing as numerous splice variants of MOR have been identified. Thus, different splice variants might confer modified functions to the receptor. Despite this knowledge, how splice variants other than the canonical MOR-1 might differentially regulate human immunodeficiency virus type-1 (HIV-1) pathogenesis in the central nervous system (CNS), or elsewhere, has been largely ignored. We began profiling MOR variants in primary human CNS cell types and discovered differing expression profiles across astrocytes, microglia, and neurons as well as differences in their regulation of expression by HIV in individual cell types and infected human brain tissue. Furthermore, we found that the MOR variants we examined differed in morphine-mediated calcium signaling. We recently published these combined findings in the Journal of NeuroVirology (PMID: 22528479). Given these differential expression patterns, regulation of expression, and cellular signaling, we hypothesize that MOR variants may mediate MOR-related HIV effects by a mechanism separate from that of MOR-1. This grant will further expand on our recently published findings to further determine how this variation might affect the progression of HIV-related neurological complications in the CNS of opioid abusers through the following specific aims: 1) determine the expression profile and regulation of expression levels with the unique N-terminal truncated MOR-1K variant by HIV in region-specific human brain tissue, and in individual human CNS cell types following HIV-1 and/or morphine treatment and 2) further examine the variants MOR-1, MOR-1A, and MOR-1X from our recently published study for functional differences in cellular signaling other than calcium, a well as for morphine binding, agonist-induced internalization, and how these variants might affect HIV-1 cellular binding and entry in the context of morphine. MOR-1K expression levels will be examined in uninfected verses HIV infected human brain tissue from frontal cortex, frontal lobe white matter, and basal ganglia as well as in HIV-1 infected verses control primary CNS cell types using the real-time polymerase chain reaction. Luciferase-based cellular signaling assays will use plasmid constructs with which luciferase expression is under the control of response elements for various cellular signaling pathways. Morphine binding will be assessed using GTP gamma S binding assays. Agonist-induced internalization will be examined using in-cell/on-cell western assays. The effects of MOR variants on HIV-1 binding and entry will be examined using cell fusion and infectious assays.
Drug abuse and human immunodeficiency virus type-1 (HIV-1) are interlinked epidemics in the United States and HIV is now largely spread through the injection drug use of opiates such as morphine, which act at ?-opioid receptors (MORs). Opiate drugs exacerbate HIV-related complications through direct actions in HIV infected cells, which make understanding how the opioid system modulates HIV pathogenesis fundamental toward understanding progression of the HIV disease. The proposed work will examine how MOR splicing variation contributes to the progression of HIV-related neurological complications in the central nervous system (CNS), which could lead to new therapeutic targets and strategies through the identification of novel molecular mechanisms that are regulated by HIV-MOR interactions on particular CNS cell types expressing a particular MOR variant.
|Dever, Seth M; Costin, Blair N; Xu, Ruqiang et al. (2014) Differential expression of the alternatively spliced OPRM1 isoform ?-opioid receptor-1K in HIV-infected individuals. AIDS 28:19-30|
|El-Hage, Nazira; Rodriguez, Myosotys; Podhaizer, Elizabeth M et al. (2014) Ibudilast (AV411), and its AV1013 analog, reduce HIV-1 replication and neuronal death induced by HIV-1 and morphine. AIDS 28:1409-19|
|El-Hage, Nazira; Dever, Seth M; Podhaizer, Elizabeth M et al. (2013) A novel bivalent HIV-1 entry inhibitor reveals fundamental differences in CCR5-*-opioid receptor interactions between human astroglia and microglia. AIDS 27:2181-90|