Chloride channel-dependent mechanisms of opiate and HIV-induced synaptodendritic injury

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Opioid use disorder (OUD) in people infected with HIV-1 (PWH) exacerbates the pathobiology of neuroHIV and HIV-associated neurocognitive disorder (HAND) via complex mechanisms involving direct peripheral (e.g., immune suppression) and CNS (e.g., neuroinflammation, blood-brain barrier damage, and neuronal injury) effects. Although glutamatergic excitotoxicity remains the major driving force in HIV-dependent neuronal injury, emerging evidence from our lab indicates that the loss of inhibitory GABAergic control and [Cl−]i homeostasis not only contributes to a loss of inhibition, but also increases the excitatory/neurotoxic effects of glutamate at NMDA and AMPA receptors. This proposal directly tests the assumption that [Cl−]i-dependent net losses of neuronal inhibition will increase excitation and promote excitotoxic. In mature neurons, intracellular Cl− concentration ([Cl−]i) is dynamically regulated by membrane ion channels (GABAA and glycine receptors) and Cl− channels and transporters. By controlling [Cl−]i, these channels and transporters regulate the Cl− equilibrium potential (ECl) and neuronal inhibition and we have already shown the KCC2 loss-of-function in OUD/neuroHIV models leads to [Cl−]i increases, elevated ECl, and excitotoxicity. We discovered a previously unidentified a ClC-1-like voltage-dependent Cl− channel that differentially regulates the excitability of dopamine receptor D1- and D2- expressing (D1 and D2, respectively) striatal medium spiny neurons (MSNs) and is disrupted by opioids and HIV Tat. We hypothesize that opioids and HIV-1 disrupt [Cl−]i, homeostasis via a voltage-gated ClC Cl− channel resulting in synaptodendritic injury. This hypothesis will be tested in the following specific aims and if confirmed will be a conceptual shift in our understanding of how opioids and HIV contribute to excitotoxicity. Aim 1 will identify the mechanisms by which opioids and HIV dysregulate voltage- dependent ClC-1 channels and Cl− homeostasis in vitro. Our preliminary findings suggest that ClC-1 significantly affects voltage-dependent Cl− currents in D1 and D2 MSNs. The extent to which ClC-1 mediates the HIV and opioid-induced pathology will be explored using gain-of-function, loss-of-function strategies, and confirmed in vitro and in vivo/ex vivo using electrophysiologic, pharmacologic, and genetic approaches at the cell and molecular levels. Aim 2. Establish to what extent ClC-1 modulation contributes to the synaptodendritic injury caused by opioids and HIV-induced dysregulation of Cl− homeostasis. We predict that the dysregulation of [Cl−]i homeostasis amplifies synaptodendritic injury in MSNs in OUD/neuroHIV models. Since ClC-1 can be a novel source of activity-dependent Cl− entry, we also predict that opioids and HIV Tat cause neuronal injury via a complex mechanism involving ClC-1 modulation and, thus, synaptic disinhibition. This project will test whether opioids and HIV dysregulate [Cl−]i homeostasis and cause neuronal injury via ClC-1. If ClC-1 is confirmed to be that target, as preliminary evidence indicates, this will be a conceptual shift in our understanding of how opioids and HIV contribute to excitotoxicity.