Xiu-ti Hu - US grants

DESCRIPTION (provided by applicant): Dysregulation and progressive neurodegeneration in the prefrontal cortex of HIV/AIDS patients affect cognitive function and are implicated in HIV-associated neurocognitive disorders (HAND), including HIV- associated dementia (HAD). Although the precise mechanisms that underlie HAD/HAND remain unknown, HIV-1 viral proteins are most likely involved. A prominent example is HIV-1 Tat (Tat), a HIV-1 transactivating protein, causes dysregulation and ultimately death of cortical/striatal neurons and astrocytes in vitro. In the brain, Tat is released by HIV-infected microglia, macrophages and monocytes. The neurotoxic effects of Tat in vitro result from an excessive increase of cytosolic free Ca2+ levels caused by abnormally enhanced Ca2+ influx via L-type Ca2+ channels, activation of ionotropic glutamate receptors and intracellular Ca2+ release. Cocaine abuse increases the morbidity and mortality of HIV/AIDS-associated pathology with exacerbation of cognitive, psychosocial and motor deficits. Even though the medial prefrontal cortex (mPFC) is well- recognized as a critical brain region in the control of these behaviors, as well as of addiction, the mechanisms underlying its dysregulation in the cocaine abuse-HIV/AIDS comorbidity have not been examined. Also, current HIV/AIDS studies predominantly focus on cell cultures, thus, the circuit-related processes are unknown. These two situations have delayed our understanding of the neuropathogenesis of this comorbidity. Our recent work with rats demonstrate that chronic cocaine exposure significantly increases Ca2+ influx by selectively upregulating L-type Ca2+ channel function in pyramidal neurons localized in the mPFC. These novel findings strongly suggest that chronic exposure to cocaine in vivo may potentiate the dysregulation and consequent neurodegeneration of the mPFC induced by Tat. Based on these evidences, the objective of the current project is to ascertain a common biological substrate in the mPFC for cocaine abuse on HIV/AIDS. The designed experiments will test the hypothesis that chronic cocaine exposure in vivo increases susceptibility and vulnerability of the mPFC to HIV-1 Tat by increasing Ca2+ influx via L-type Ca2+ channels in pyramidal cells, and these changes contribute to the pathogenesis of cocaine abuse and HAD. This pilot study is innovative and significant because it will provide informative preliminary data pointing to the mechanisms underlying the cocaine abuse-HAD comorbidity. Results from these studies will used to develop a R01 application focusing on this comorbidity and putative therapeutic targets to help treat dually afflicted individuals. PUBLIC HEALTH RELEVANCE: This grant application is titled: "Chronic cocaine exposure &HIV-1 Tat: Dysregulation of the medial prefrontal cortex." Our goal is to explore the mechanisms of the brain and behavioral deficits induced by cocaine abuse and HIV infection. The combination of these two problems results in a much more severe condition, and unfortunately the combination is spreading in our society. This project will determine how a protein (Tat) produced by HIV-infected cells alters activity of neurons in the medial prefrontal cortex. This brain region is involved both in reward and in judgment, and it is compromised in both cocaine addiction and AIDS. We propose that changes in neuronal activity in the medial prefrontal cortex induced by the HIV protein Tat will be potentiated by repeated cocaine exposure. Findings from this study will help us to verify the common target of cocaine abuse and HIV infection, which will be very important for developing more effective treatments for cocaine abuse and HIV-related brain/behavioral deficits.

DESCRIPTION (provided by applicant): Cocaine-abusing, HIV+ individuals experience more severe and rapidly evolving neuropsychiatric disorders and HIV-associated neurocognitive disorders (HAND) than non-abusing HIV+ individuals. These disorders reflect in part, a dysregulation of the medial prefrontal cortex (mPFC), but the underlying mechanisms are unclear. To significantly advance understanding of this phenomenon, we submit this R21 application to implement a new rat model of the comorbid human condition wherein critical features of both HIV+ and cocaine abuse are emulated to investigate the functional state of mPFC neurons. The research objectives are: (1) To reveal dysfunction of the mPFC in non-infectious, young (10-12 week old) adult HIV-1 transgenic (Tg) rats that chronically express multiple HIV-1 protein toxins in the brain, but do not exhibit the overt pathologies seen in old Tg rats (i.e., 10-12 months of age). (2 To determine if an exaggerated pathophysiology exists in the mPFC of HIV-1 Tg rats trained to self-administer cocaine. Our central hypothesis is that mPFC pyramidal neurons are more prone to over-activation in cocaine self-administering Tg rats than in saline-treated Tg rats or cocaine self-administering non-Tg rats. We will test this hypothesis with two Specific Aims. Aim 1 will validate [the level of microgliosis] in the mPFC, and determine their association with abnormally- enhanced neuronal excitability mediated via over-activated L-type Ca2+ channels in the young adult Tg rats. Non-Tg rats will serve as controls. Functional activity of mPFC pyramidal neurons will be assessed in ex vivo brain slices using patch-clamp recording techniques. We expect that the Tg rats will show [mPFC microgliosis] and hyper-excitability of pyramidal neurons mediated by over-activated L-channels, reflected by abnormally-increased responses to excitatory stimuli. Aim 2 will determine that cocaine self- administration enhances the pyramidal cell pathophysiology seen in the Tg rats. We expect that cocaine will exacerbate the hyper-excitability of mPFC pyramidal neurons observed in the Tg rats. This research is significant and innovative because it will provide the first means to ascertain the functional consequences of the chronic brain exposure of HIV-1 proteins under conditions of cocaine self-administration; in so doing, it will provide a completely novel method to investigate the molecular and cellular mechanisms responsible for this comorbid condition. The outcomes expected by the end of two years will determine that the HIV-1 proteins-induced pathology and cocaine abuse converge to dysregulate mPFC neuronal function via over- activated L-channels. These outcomes will guide our future investigations on mechanisms underlying the dysregulated Ca2+ homeostasis in the mPFC. Thus, this bold research will vertically elevate our understanding of the mechanisms that may contribute to the neuropathogenesis of HAND and neuropsychiatric disorders associated with HIV+ and cocaine abuse. This new knowledge will also lead to a R01 application in which novel therapeutic strategies will be developed to reduce such pathology. PUBLIC HEALTH RELEVANCE: The combination of cocaine abuse and AIDS accelerates the brain damage that can accompany either condition alone. Unfortunately, the frequency of this combination, or comorbidity, it is increasing in our society. This project will determine how toxic viral proteins produced by HIV in the rat brain, with or without cocaine self- administration, alter the function of neurons in a bran region (the medial prefrontal cortex) which, in humans, is vulnerable to both HIV infection and cocaine abuse. We propose that the detrimental effects of HIV toxins will be potentiated by cocaine on specific proteins that are critical for normal neuronal firing. By identifying common molecular targets of cocaine and HIV toxins, outcomes from this project should prove to be critically important in developing therapeutic means to slow the devastation of HIV/AIDS comorbidity with cocaine abuse, as well as to promote drug abstinence in these individuals.

DESCRIPTION (provided by applicant): Despite antiretroviral therapy, approximately 50% of HIV+ patients in the USA are diagnosed with HIV-associated neurocognitive disorders (HAND). HIV alters two key regulators of cognition and psychomotor activity, the medial prefrontal cortex (mPFC) and caudate-putamen (CPu; a.k.a. dorsal striatum). Excessive Ca2+ influx (partly via NMDA receptor, NMDAR) is critical in neuronal excitotoxicity, but the mechanisms underlying HIV neuropathogenesis are not entirely clear. Our published and pilot data point to an additional mechanism in HIV-mediated neuronal hyper-excitability which involves the voltage-gated L-type Ca2+ channel, independent of NMDAR. We showed that HIV-1 transactivator of transcription (Tat) increases Ca2+ influx by upregulating L-channels in mPFC pyramidal neurons, rendering these cells more susceptible and vulnerable to hyper-excitability. Tat also potentiates the interaction between NMDAR and L-channel, and neurons showed similar over-excitation as aged or modeled using HIV-1 transgenic (Tg) rats. Together, these findings point to a unique mechanism by which HIV induces excessive neuroexcitation. Given that neither an L-channel blocker nor a NMDAR antagonist alone was able to treat HAND at late stage, understanding the intricate interplay between HIV and the L-channels may provide novel therapeutic strategies for HAND. Our central hypothesis is that HIV infection of the brain renders mPFC pyramidal neurons and CPu medium spiny neurons (MSNs) more susceptible and vulnerable to excitatory stimuli via over-activating the L-channels; and that combined treatments of an L-channel and NMDAR blocker will ameliorate the mPFC/CPu neuropathophysiology, more effectively in adolescent than in older brain. We will test this hypothesis via three aims using integrated electro- physiological, optic/fluorescence imaging and immuno/biochemical approaches. In Aim 1 we will define the cellular/molecular mechanism(s) by which HIV mediates L-channel over-activation and consequently neuronal hyper-excitation in the mPFC/CPu of adolescent Tg rats. We expect that HIV-1 proteins released by infected cells mediate L-channel over-activation in part via engaging dopamine D1 receptor (D1R), and will reveal the domain of Tat that mediates the Tat effects on L-channels. In Aim 2 we will evaluate the interplay between NMDAR and L-channel in mediating neuronal hyper-excitability. We hypothesize that NMDA-evoked Ca2+ signal is over-amplified and relayed by LVA-L channels to other ion channels that ultimately control firing. In Aim 3 we will ascertain age (a significant risk factor for HAND)-associated exacerbation in L-channel over-activation and mPFC/CPu neuropathophysiology in older Tg rats. We hypothesize that mPFC/CPu neuropathophysiology will be more severe in aging Tg rats than in adolescent Tg rats (and aging non-Tg rats). Lately, we expect that combined treatments of L-channel/NMDAR blockers will diminish/ameliorate HIV-mediated neuronal hyper-excitability. Collectively, our studies will establish a novel paradigm in HIV-mediated neuropathogenesis and identify novel targets for future therapeutic intervention for HIV-mediated neuropathology.

ABSTRACT Despite long-term combined antiretroviral therapy (cART), HIV+ drug abusers experience more severe and rapid progression of HIV-associated neurocognitive disorders (HAND). Methamphetamine (Meth) is a potent stimulant, which is highly abused in the USA, and associated with chronic systemic inflammation. Understanding the mechanism(s) by which Meth and HIV induce pathologies in the brain, whether and how chronic cART alters astrocyte/neuron function in the CNS, and whether Math (or other stimulants) affects cART, is critical for improving current, and developing new effective therapeutic strategies for treating Meth addiction and its comorbid conditions with HIV/AIDS. The medial prefrontal cortex (mPFC) is one of the key regulators of cognition and addiction; but it is profoundly altered following chronic Meth and HIV exposure in vivo with disrupted dopamine (DA) neurotransmission and immune activation of astrocytes. Both human and laboratory animal studies demonstrate that the mPFC dysregulation is significantly implicated in chronic Meth-induced brain dysfunction, associated with euphoria, craving, drug-taking and relapse. Cumulating evidence also indicates that both astrocytes and glutamatergic pyramidal neurons in the PFC are altered after chronic Meth or HIV exposure. Chronic Meth in vivo induces astrocytic and neuronal plasticity that differs from changes induced by acute Meth in vitro; but the underlying mechanism(s) of either one is not fully understood. In this proposed study, we will elucidate a novel, trace amine-associated receptor 1 (TAAR1)-mediated mechanism, by which acute Meth in vitro alters astrocyte activity; and determine alterations in this mechanism after Meth self-administration (Meth-SA) followed by a withdrawal (that elicits drug-seeking), in HIV-1 transgenic (Tg) or non-Tg rats. Moreover, we will also determine whether and how chronic cART in vivo affects functional activity of mPFC astrocytes and neurons; and whether such cART effects are altered by Meth-SA in the context of neuroHIV. Specifically, we will first define the mechanism by which chronic Meth in vivo alters astrocyte function (Aim1). We then will determine the mechanism by which Meth/HIV impacting on mPFC astrocytes/ neurons (Aim2). Finally, we will identify chronic effects of cART (using a fixed-dose combination of abacavir, dolutegravir and lamivudine) in vivo on mPFC astrocytes/neurons, and whether chronic Meth-SA alters them (Aim3). To prove the fundamental base of this proposed research, we provide justification, related citations, and preliminary data to support the Scientific Premise, Scientific Rigor, Relevant Biological Variables, and Resource Authentication for this proposal. Outcomes from the proposed research will reveal the novel, TAAR1- mediated mechanism that plays a key role in underlying the comorbid Meth/HIV impact, and chronic cART effects, on altering astrocytes and pyramidal neurons in the mPFC. This will advance our understanding for interactive effects of Meth, HIV and cART in the CNS, and therefore will induce a strong impact to the field.