Synergistic neurotoxicity of opioids and human immunodeficiency virus-1 Tat protein in striatal neurons in vitro

Abstract

Human immunodeficiency virus (HIV) infection selectively targets the striatum, a region rich in opioid receptor-expressing neural cells, resulting in gliosis and neuronal losses. Opioids can be neuroprotective or can promote neurodegeneration. To determine whether opioids modify the response of neurons to human immunodeficiency virus type 1 (HIV-1) Tat protein-induced neurotoxicity, neural cell cultures from mouse striatum were initially characterized for mu and/or kappa opioid receptor immunoreactivity. These cultures were continuously treated with morphine, the opioid antagonist naloxone, and/or HIV-1 Tat (1-72) protein, a non-neurotoxic HIV-1 Tat deletion mutant (TatDelta31-61) protein, or immunoneutralized HIV-1 Tat (1-72) protein. Neuronal and astrocyte viability was examined by ethidium monoazide exclusion, and by apoptotic changes in nuclear heterochromatin using Hoechst 33342. Morphine (10nM, 100nM or 1microM) significantly increased Tat-induced (100 or 200nM) neuronal losses by about two-fold at 24h following exposure. The synergistic effects of morphine and Tat were prevented by naloxone (3microM), indicating the involvement of opioid receptors. Furthermore, morphine was not toxic when combined with mutant Tat or immunoneutralized Tat. Neuronal losses were accompanied by chromatin condensation and pyknosis. Astrocyte viability was unaffected. These findings demonstrate that acute opioid exposure can exacerbate the neurodegenerative effect of HIV-1 Tat protein in striatal neurons, and infer a means by which opioids may hasten the progression of HIV-associated dementia.

Differential involvement of p38 and JNK MAP kinases in HIV-1 Tat and gp120-induced apoptosis and neurite degeneration in striatal neurons

“…1A) or 48 h (data not shown) following exposure as previously reported (Gurwell et al, 2001;Singh et al, 2004). The toxic actions of both virotoxins were highly selective and toxicity was not observed when nontoxic, mutant Tat (Tat Δ31-61 ) or denatured gp120 (20 min boiling) were used (Fig.…”

Apoptotic death of striatal neurons induced by human immunodeficiency virus-1 Tat and gp120: Differential involvement of caspase-3 and endonuclease G

“…Striatal neurons were seeded at identical densities (4 × 10 5 cells/cm 2 ) and grown on poly-D-lysine (0.1 mg/ml) coated Costar-24 well-plates in an incubator maintained at 35-36 °C in 5% CO 2 95% air and high humidity. The cultures described in the present study are enriched in medium spiny neurons, which comprise about 85-95% of neurons in the mouse striatum, and contain about 3-8% astroglia and about 0.4% monocytes/microglia (Hauser, unpublished), but which differ from an alternative mixed neuronal-glial striatal culture system previously characterized by us (Gurwell et al, 2001).…”

“…Recombinant Tat 1-72 was prepared as described previously with minor modifications (Gurwell et al, 2001). Recombinant gp120 from HIV-SF2 made in CHO cells was a gift from Chiron Inc. (Emeryville, CA, USA), and purified and handled as previously described (Holden et al, 1999;Haughey, 2002).…”

“…The neuronal identity of the dying cells was further confirmed by exposing cells to 0.5 µg/ml ethidium monoazide (EMA) in Dulbecco's PBS for 30 min, permanently binding the EMA to the dying cells through photoaffinity labeling, and co-detecting EMA-positive cells with neuron-specific antigenic markers as previously described (Gurwell et al, 2001). Briefly, EMA is excluded from living cells, but intercalates to the DNA of dying cells, where it can be permanently linked via photoaffinity by exposure to a 45 W fluorescent light (15 cm distance) for 30 minutes at room temperature.…”

Fulminant encephalopathy with basal ganglia hyperintensities in HIV-infected drug users

“…Opiates are not intrinsically toxic. 18 However, chronic, intermittent opiate exposure disrupts secondary messenger cascades and increases oxidative stress, 19 which may enhance the vulnerability of neurons to HIV proteins such as Tat and gp120. 20 In transgenic mice that express the HIV-Tat protein, morphine exposure had profound effects in the basal ganglia.…”