Slingshot homolog-1-mediated Nrf2 sequestration tips the balance from neuroprotection to neurodegeneration in Alzheimer's disease.

Oxidative damage in the brain is one of the earliest drivers of pathology in Alzheimer's disease (AD) and related dementias, both preceding and exacerbating clinical symptoms. In response to oxidative stress, nuclear factor erythroid 2-related factor 2 (Nrf2) is normally activated to protect the brain from oxidative damage. However, Nrf2-mediated defense against oxidative stress declines in AD, rendering the brain increasingly vulnerable to oxidative damage. Although this phenomenon has long been recognized, its mechanistic basis has been a mystery. Here, we demonstrate through in vitro and in vivo models, as well as human AD brain tissue, that Slingshot homolog-1 (SSH1) drives this effect by acting as a counterweight to neuroprotective Nrf2 in response to oxidative stress and disease. Specifically, oxidative stress-activated SSH1 suppresses nuclear Nrf2 signaling by sequestering Nrf2 complexes on actin filaments and augmenting Kelch-like ECH-associated protein 1 (Keap1)-Nrf2 interaction, independently of SSH1 phosphatase activity. We also show that Ssh1 elimination in AD models increases Nrf2 activation, which mitigates tau and amyloid-ß accumulation and protects against oxidative injury, neuroinflammation, and neurodegeneration. Furthermore, loss of Ssh1 preserves normal synaptic function and transcriptomic patterns in tauP301S mice. Importantly, we also show that human AD brains exhibit highly elevated interactions of Nrf2 with both SSH1 and Keap1. Thus, we demonstrate here a unique mode of Nrf2 blockade that occurs through SSH1, which drives oxidative damage and ensuing pathogenesis in AD. Strategies to inhibit SSH1-mediated Nrf2 suppression while preserving normal SSH1 catalytic function may provide new neuroprotective therapies for AD and related dementias.

Longitudinal in vivo monitoring of axonal degeneration after brain injury.

Traumatic brain injury (TBI)-induced axonal degeneration leads to acute and chronic neuropsychiatric impairment, neuronal death, and accelerated neurodegenerative diseases of aging, including Alzheimer's and Parkinson's diseases. In laboratory models, axonal degeneration is traditionally studied through comprehensive postmortem histological evaluation of axonal integrity at multiple time points. This requires large numbers of animals to power for statistical significance. Here, we developed a method to longitudinally monitor axonal functional activity before and after injury in vivo in the same animal over an extended period. Specifically, after expressing an axonal-targeting genetically encoded calcium indicator in the mouse dorsolateral geniculate nucleus, we recorded axonal activity patterns in the visual cortex in response to visual stimulation. In vivo aberrant axonal activity patterns after TBI were detectable from 3 days after injury and persisted chronically. This method generates longitudinal same-animal data that substantially reduces the number of required animals for preclinical studies of axonal degeneration.

Reducing acetylated tau is neuroprotective in brain injury.

Traumatic brain injury (TBI) is the largest non-genetic, non-aging related risk factor for Alzheimer's disease (AD). We report here that TBI induces tau acetylation (ac-tau) at sites acetylated also in human AD brain. This is mediated by S-nitrosylated-GAPDH, which simultaneously inactivates Sirtuin1 deacetylase and activates p300/CBP acetyltransferase, increasing neuronal ac-tau. Subsequent tau mislocalization causes neurodegeneration and neurobehavioral impairment, and ac-tau accumulates in the blood. Blocking GAPDH S-nitrosylation, inhibiting p300/CBP, or stimulating Sirtuin1 all protect mice from neurodegeneration, neurobehavioral impairment, and blood and brain accumulation of ac-tau after TBI. Ac-tau is thus a therapeutic target and potential blood biomarker of TBI that may represent pathologic convergence between TBI and AD. Increased ac-tau in human AD brain is further augmented in AD patients with history of TBI, and patients receiving the p300/CBP inhibitors salsalate or diflunisal exhibit decreased incidence of AD and clinically diagnosed TBI.

Characterization of the Jet-Flow Overpressure Model of Traumatic Brain Injury in Mice.

The jet-flow overpressure chamber (OPC) has been previously reported as a model of blast-mediated traumatic brain injury (bTBI). However, rigorous characterization of the features of this injury apparatus shows that it fails to recapitulate exposure to an isolated blast wave. Through combined experimental and computational modeling analysis of gas-dynamic flow conditions, we show here that the jet-flow OPC produces a collimated high-speed jet flow with extreme dynamic pressure that delivers a severe compressive impulse. Variable rupture dynamics of the diaphragm through which the jet flow originates also generate a weak and infrequent shock front. In addition, there is a component of acceleration-deceleration injury to the head as it is agitated in the headrest. Although not a faithful model of free-field blast exposure, the jet-flow OPC produces a complex multi-modal model of TBI that can be useful in laboratory investigation of putative TBI therapies and fundamental neurophysiological processes after brain injury.

Maternal P7C3-A20 Treatment Protects Offspring from Neuropsychiatric Sequelae of Prenatal Stress.

Aims: Impaired embryonic cortical interneuron development from prenatal stress is linked to adult neuropsychiatric impairment, stemming in part from excessive generation of reactive oxygen species in the developing embryo. Unfortunately, there are no preventive medicines that mitigate the risk of prenatal stress to the embryo, as the underlying pathophysiologic mechanisms are poorly understood. Our goal was to interrogate the molecular basis of prenatal stress-mediated damage to the embryonic brain to identify a neuroprotective strategy. Results: Chronic prenatal stress in mice dysregulated nicotinamide adenine dinucleotide (NAD+) synthesis enzymes and cortical interneuron development in the embryonic brain, leading to axonal degeneration in the hippocampus, cognitive deficits, and depression-like behavior in adulthood. Offspring were protected from these deleterious effects by concurrent maternal administration of the NAD+-modulating agent P7C3-A20, which crossed the placenta to access the embryonic brain. Prenatal stress also produced axonal degeneration in the adult corpus callosum, which was not prevented by maternal P7C3-A20. Innovation: Prenatal stress dysregulates gene expression of NAD+-synthesis machinery and GABAergic interneuron development in the embryonic brain, which is associated with adult cognitive impairment and depression-like behavior. We establish a maternally directed treatment that protects offspring from these effects of prenatal stress. Conclusion: NAD+-synthesis machinery and GABAergic interneuron development are critical to proper embryonic brain development underlying postnatal neuropsychiatric functioning, and these systems are highly susceptible to prenatal stress. Pharmacologic stabilization of NAD+ in the stressed embryonic brain may provide a neuroprotective strategy that preserves normal embryonic development and protects offspring from neuropsychiatric impairment. Antioxid. Redox Signal. 35, 511-530.

Dopamine D1R-neuron cacna1c deficiency: a new model of extinction therapy-resistant post-traumatic stress.

Post-traumatic stress disorder (PTSD) is characterized by persistent fear memory of remote traumatic events, mental re-experiencing of the trauma, long-term cognitive deficits, and PTSD-associated hippocampal dysfunction. Extinction-based therapeutic approaches acutely reduce fear. However, many patients eventually relapse to the original conditioned fear response. Thus, understanding the underlying molecular mechanisms of this condition is critical to developing new treatments for patients. Mutations in the neuropsychiatric risk gene CACNA1C, which encodes the Cav1.2 isoform of the L-type calcium channel, have been implicated in both PTSD and highly comorbid neuropsychiatric conditions, such as anxiety and depression. Here, we report that male mice with global heterozygous loss of cacna1c exhibit exacerbated contextual fear that persists at remote time points (up to 180 days after shock), despite successful acute extinction training, reminiscent of PTSD patients. Because dopamine has been implicated in contextual fear memory, and Cav1.2 is a downstream target of dopamine D1-receptor (D1R) signaling, we next generated mice with specific deletion of cacna1c from D1R-expressing neurons (D1-cacna1cKO mice). Notably, D1-cacna1cKO mice also show the same exaggerated remote contextual fear, as well as persistently elevated anxiety-like behavior and impaired spatial memory at remote time points, reminiscent of chronic anxiety in treatment-resistant PTSD. We also show that D1-cacna1cKO mice exhibit elevated death of young hippocampal neurons, and that treatment with the neuroprotective agent P7C3-A20 eradicates persistent remote fear. Augmenting survival of young hippocampal neurons may thus provide an effective therapeutic approach for promoting durable remission of PTSD, particularly in patients with CACNA1C mutations or other genetic aberrations that impair calcium signaling or disrupt the survival of young hippocampal neurons.

Different forms of traumatic brain injuries cause different tactile hypersensitivity profiles.

ABSTRACT: Chronic complications of traumatic brain injury represent one of the greatest financial burdens and sources of suffering in the society today. A substantial number of these patients suffer from posttraumatic headache (PTH), which is typically associated with tactile allodynia. Unfortunately, this phenomenon has been understudied, in large part because of the lack of well-characterized laboratory animal models. We have addressed this gap in the field by characterizing the tactile sensory profile of 2 nonpenetrating models of PTH. We show that multimodal traumatic brain injury, administered by a jet-flow overpressure chamber that delivers a severe compressive impulse accompanied by a variable shock front and acceleration-deceleration insult, produces long-term tactile hypersensitivity and widespread sensitization. These are phenotypes reminiscent of PTH in patients, in both cephalic and extracephalic regions. By contrast, closed head injury induces only transient cephalic tactile hypersensitivity, with no extracephalic consequences. Both models show a more severe phenotype with repetitive daily injury for 3 days, compared with either 1 or 3 successive injuries in a single day, providing new insight into patterns of injury that may place patients at a greater risk of developing PTH. After recovery from transient cephalic tactile hypersensitivity, mice subjected to closed head injury demonstrate persistent hypersensitivity to established migraine triggers, including calcitonin gene-related peptide and sodium nitroprusside, a nitric oxide donor. Our results offer the field new tools for studying PTH and preclinical support for a pathophysiologic role of calcitonin gene-related peptide in this condition.

P7C3-A20 treatment one year after TBI in mice repairs the blood-brain barrier, arrests chronic neurodegeneration, and restores cognition.

Chronic neurodegeneration in survivors of traumatic brain injury (TBI) is a major cause of morbidity, with no effective therapies to mitigate this progressive and debilitating form of nerve cell death. Here, we report that pharmacologic restoration of the blood-brain barrier (BBB), 12 mo after murine TBI, is associated with arrested axonal neurodegeneration and cognitive recovery, benefits that persisted for months after treatment cessation. Recovery was achieved by 30 d of once-daily administration of P7C3-A20, a compound that stabilizes cellular energy levels. Four months after P7C3-A20, electron microscopy revealed full repair of TBI-induced breaks in cortical and hippocampal BBB endothelium. Immunohistochemical staining identified additional benefits of P7C3-A20, including restoration of normal BBB endothelium length, increased brain capillary pericyte density, increased expression of BBB tight junction proteins, reduced brain infiltration of immunoglobulin, and attenuated neuroinflammation. These changes were accompanied by cessation of TBI-induced chronic axonal degeneration. Specificity for P7C3-A20 action on the endothelium was confirmed by protection of cultured human brain microvascular endothelial cells from hydrogen peroxide-induced cell death, as well as preservation of BBB integrity in mice after exposure to toxic levels of lipopolysaccharide. P7C3-A20 also protected mice from BBB degradation after acute TBI. Collectively, our results provide insights into the pathophysiologic mechanisms behind chronic neurodegeneration after TBI, along with a putative treatment strategy. Because TBI increases the risks of other forms of neurodegeneration involving BBB deterioration (e.g., Alzheimer's disease, Parkinson's disease, vascular dementia, chronic traumatic encephalopathy), P7C3-A20 may have widespread clinical utility in the setting of neurodegenerative conditions.

Identification of chronic brain protein changes and protein targets of serum auto-antibodies after blast-mediated traumatic brain injury.

In addition to needing acute emergency management, blast-mediated traumatic brain injury (TBI) is also a chronic disorder with delayed-onset symptoms that manifest and progress over time. While the immediate consequences of acute blast injuries are readily apparent, chronic sequelae are harder to recognize. Indeed, the identification of individuals with mild-TBI or TBI-induced symptoms is greatly impaired in large part due to the lack of objective and robust biomarkers. The purpose of this study was to address these need by identifying candidates for serum-based biomarkers of blast TBI, and also to identify unique or differentially regulated protein expression in the thalamus in C57BL/6J mice exposed to blast using high throughput qualitative screens of protein expression. To identify thalamic proteins differentially or uniquely associated with blast exposure, we utilized an antibody-based affinity-capture strategy (referred to as "proteomics-based analysis of depletomes"; PAD) to deplete thalamic lysates from blast-treated mice of endogenous thalamic proteins also found in control mice. Analysis of this "depletome" detected 75 unique proteins, many with associations to the myelin sheath. To identify blast-associated proteins eliciting production of circulating autoantibodies, serum antibodies of blast-treated mice were immobilized, and their immunogens subsequently identified by proteomic analysis of proteins specifically captured following incubation with thalamic lysates (a variant of a strategy referred to as "proteomics-based expression library screening"; PELS). This analysis identified 46 blast-associated immunogenic proteins, including 6 shared in common with the PAD analysis (ALDOA, PHKB, HBA-A1, DPYSL2, SYN1, and CKB). These proteins and their autoantibodies are appropriate for further consideration as biomarkers of blast-mediated TBI.

Neuroprotective Efficacy of a Sigma 2 Receptor/TMEM97 Modulator (DKR-1677) after Traumatic Brain Injury.

Compounds targeting the sigma 2 receptor, which we recently cloned and showed to be identical with transmembrane protein 97 (σ2R/TMEM97), are broadly applicable therapeutic agents currently in clinical trials for imaging in breast cancer and for treatment of Alzheimer's disease and schizophrenia. These promising applications coupled with our previous observation that the σ2R/TMEM97 modulator SAS-0132 has neuroprotective attributes and improves cognition in wild-type mice suggests that modulating σ2R/TMEM97 may also have therapeutic benefits in other neurodegenerative conditions such as traumatic brain injury (TBI). Herein, we report that DKR-1677, a novel derivative of SAS-0132 with increased affinity and selectivity for σ2R/Tmem97 ( Ki = 5.1 nM), is neuroprotective after blast-induced and controlled cortical impact (CCI) TBI in mice. Specifically, we discovered that treatment with DKR-1677 decreases axonal degeneration after blast-induced TBI and enhances survival of cortical neurons and oligodendrocytes after CCI injury. Furthermore, treatment with DKR-1677 preserves cognition in the Morris water maze after blast TBI. Our results support an increasingly broad role for σ2R/Tmem97 modulation in neuroprotection and suggest a new approach for treating patients suffering from TBI.

Neuroprotective efficacy of P7C3 compounds in primate hippocampus.

There is a critical need for translating basic science discoveries into new therapeutics for patients suffering from difficult to treat neuropsychiatric and neurodegenerative conditions. Previously, a target-agnostic in vivo screen in mice identified P7C3 aminopropyl carbazole as capable of enhancing the net magnitude of postnatal neurogenesis by protecting young neurons from death. Subsequently, neuroprotective efficacy of P7C3 compounds in a broad spectrum of preclinical rodent models has also been observed. An important next step in translating this work to patients is to determine whether P7C3 compounds exhibit similar efficacy in primates. Adult male rhesus monkeys received daily oral P7C3-A20 or vehicle for 38 weeks. During weeks 2-11, monkeys received weekly injection of 5'-bromo-2-deoxyuridine (BrdU) to label newborn cells, the majority of which would normally die over the following 27 weeks. BrdU+ cells were quantified using unbiased stereology. Separately in mice, the proneurogenic efficacy of P7C3-A20 was compared to that of NSI-189, a proneurogenic drug currently in clinical trials for patients with major depression. Orally-administered P7C3-A20 provided sustained plasma exposure, was well-tolerated, and elevated the survival of hippocampal BrdU+ cells in nonhuman primates without adverse central or peripheral tissue effects. In mice, NSI-189 was shown to be pro-proliferative, and P7C3-A20 elevated the net magnitude of hippocampal neurogenesis to a greater degree than NSI-189 through its distinct mechanism of promoting neuronal survival. This pilot study provides evidence that P7C3-A20 safely protects neurons in nonhuman primates, suggesting that the neuroprotective efficacy of P7C3 compounds is likely to translate to humans as well.

Ca2+-Binding Protein 1 Regulates Hippocampal-dependent Memory and Synaptic Plasticity.

Ca2+-binding protein 1 (CaBP1) is a Ca2+-sensing protein similar to calmodulin that potently regulates voltage-gated Ca2+ channels. Unlike calmodulin, however, CaBP1 is mainly expressed in neuronal cell-types and enriched in the hippocampus, where its function is unknown. Here, we investigated the role of CaBP1 in hippocampal-dependent behaviors using mice lacking expression of CaBP1 (C-KO). By western blot, the largest CaBP1 splice variant, caldendrin, was detected in hippocampal lysates from wild-type (WT) but not C-KO mice. Compared to WT mice, C-KO mice exhibited mild deficits in spatial learning and memory in both the Barnes maze and in Morris water maze reversal learning. In contextual but not cued fear-conditioning assays, C-KO mice showed greater freezing responses than WT mice. In addition, the number of adult-born neurons in the hippocampus of C-KO mice was ∼40% of that in WT mice, as measured by bromodeoxyuridine labeling. Moreover, hippocampal long-term potentiation was significantly reduced in C-KO mice. We conclude that CaBP1 is required for cellular mechanisms underlying optimal encoding of hippocampal-dependent spatial and fear-related memories.

Treatment with Uric Acid Reduces Infarct and Improves Neurologic Function in Female Mice After Transient Cerebral Ischemia.

BACKGROUND: Exogenous administration of uric acid, a naturally occurring antioxidant that scavenges reactive oxygen species in vasculature, has shown protective efficacy in both rodent models of stroke and human stroke patients in Spain as an adjuvant treatment to mechanical thrombectomy. Before clinical trials can be initiated in the United States, however, confirmation of efficacy in alternative preclinical models is required in accordance with stroke therapy academic industry roundtable-RIGOR criteria. To date, preclinical efficacy has only been established in the acute setting in male rodents. METHODS: To address this need, we subjected 7- to 9-week old ovariectomized female mice to filament-induced right middle cerebral artery ischemia and reperfusion, an established preclinical model of mechanical thrombectomy. Fidelity of the procedure was monitored by laser Doppler flowmetry. A separate lab randomly assigned animals to vehicle versus uric acid infusion, which was initiated immediately after 45 minutes of reperfusion. Poststroke analysis of infarction size and neurologic function were conducted by investigators blind to treatment group, with a 7-day primary endpoint and a 3-day intermediary analysis at 1and. RESULTS: Infarct size and neurologic function at 7 days poststroke were significantly improved in uric acid-treated animals, relative to vehicle. CONCLUSION: Efficacy of uric acid in preclinical models of stroke is now expanded to include female mice analyzed at a later time point than has been investigated previously. These results support stroke therapy academic industry roundtable-RIGOR driven determination of the suitability of acute administration of uric acid as an adjuvant to mechanical thrombectomy in clinical trials for patients with stroke.

A START-domain-containing protein is a novel marker of nervous system components of the sea cucumber Holothuria glaberrima.

One of the main challenges faced by investigators studying the nervous system of members of the phylum Echinodermata is the lack of markers to identify nerve cells and plexi. Previous studies have utilized an antibody, RN1, that labels most of the nervous system structures of the sea cucumber Holothuria glaberrima and other echinoderms. However, the antigen recognized by RN1 remained unknown. In the present work, the antigen has been characterized by immunoprecipitation, tandem mass spectrometry, and cDNA cloning. The RN1 antigen contains a START lipid-binding domain found in Steroidogenic Acute Regulatory (StAR) proteins and other lipid-binding proteins. Phylogenetic tree assembly showed that the START domain is highly conserved among echinoderms. We have named this antigen HgSTARD10 for its high sequence similarity to the vertebrate orthologs. Gene and protein expression analyses revealed an abundance of HgSTARD10 in most H. glaberrima tissues including radial nerve, intestine, muscle, esophagus, mesentery, hemal system, gonads and respiratory tree. Molecular cloning of HgSTARD10, consequent protein expression and polyclonal antibody production revealed the STARD10 ortholog as the antigen recognized by the RN1 antibody. Further characterization into this START domain-containing protein will provide important insights for the biochemistry, physiology and evolution of deuterostomes.

Protective efficacy of P7C3-S243 in the 6-hydroxydopamine model of Parkinson's disease.

BACKGROUND: There are currently no therapeutic options for patients with Parkinson's disease that prevent or slow the death of dopaminergic neurons. We have recently identified the novel P7C3 class of neuroprotective molecules that blocks neuron cell death. AIMS: The aim of this study was to determine whether treatment with highly active members of the P7C3 series blocks dopaminergic neuron cell death and associated behavioral and neurochemical deficits in the rat 6-hydroxydopamine (6-OHDA) model of Parkinson's disease. METHODS: After unilateral injection of 6-OHDA into the median forebrain bundle, rats were assessed for behavioral function in the open field, cylinder test, and amphetamine-induced circling test. Thereafter, their brains were subjected to neurochemical and immunohistochemical analysis of dopaminergic neuron survival. Analysis was conducted as a function of treatment with P7C3 compounds, with administration initiated either before or after 6-OHDA exposure. RESULTS: Animals administered P7C3-A20 or P7C3-S243, two of the most advanced agents in the P7C3 series of neuroprotective compounds, both before and after 6-OHDA exposure showed evidence of protective efficacy in all measures. When P7C3-S243 administration was initiated after 6-OHDA exposure, rats also showed protective efficacy in all measures, which included blocking dopaminergic neuron cell death in ipsilateral substantia nigra pars compacta, preservation of dopamine and its metabolites in ipsilateral striatum, and preservation of normal motor behavior. CONCLUSIONS: The P7C3 series of compounds may form the basis for developing new therapeutic agents for slowing or preventing progression of Parkinson's disease.

Cdk5 phosphorylation of EFhd2 at S74 affects its calcium binding activity.

EFhd2 is a calcium binding protein, which is highly expressed in the central nervous system and associated with pathological forms of tau proteins in tauopathies. Previous phosphoproteomics studies and bioinformatics analysis suggest that EFhd2 may be phosphorylated. Here, we determine whether Cdk5, a hyperactivated kinase in tauopathies, phosphorylates EFhd2 and influence its known molecular activities. The results indicated that EFhd2 is phosphorylated by brain extract of the transgenic mouse CK-p25, which overexpresses the Cdk5 constitutive activator p25. Consistently, in vitro kinase assays demonstrated that Cdk5, but not GSK3ß, directly phosphorylates EFhd2. Biomass, tandem mass spectrometry, and mutagenesis analyses indicated that Cdk5 monophosphorylates EFhd2 at S74, but not the adjacent S76. Furthermore, Cdk5-mediated phosphorylation of EFhd2 affected its calcium binding activity. Finally, a phospho-specific antibody was generated against EFhd2 phosphorylated at S74 and was used to detect this phosphorylation event in postmortem brain tissue from Alzheimer's disease and normal-aging control cases. Results demonstrated that EFhd2 is phosphorylated in vivo at S74. These results imply that EFhd2's physiological and/or pathological function could be regulated by its phosphorylation state.

Solution structure of the tandem acyl carrier protein domains from a polyunsaturated fatty acid synthase reveals beads-on-a-string configuration.

The polyunsaturated fatty acid (PUFA) synthases from deep-sea bacteria invariably contain multiple acyl carrier protein (ACP) domains in tandem. This conserved tandem arrangement has been implicated in both amplification of fatty acid production (additive effect) and in structural stabilization of the multidomain protein (synergistic effect). While the more accepted model is one in which domains act independently, recent reports suggest that ACP domains may form higher oligomers. Elucidating the three-dimensional structure of tandem arrangements may therefore give important insights into the functional relevance of these structures, and hence guide bioengineering strategies. In an effort to elucidate the three-dimensional structure of tandem repeats from deep-sea anaerobic bacteria, we have expressed and purified a fragment consisting of five tandem ACP domains from the PUFA synthase from Photobacterium profundum. Analysis of the tandem ACP fragment by analytical gel filtration chromatography showed a retention time suggestive of a multimeric protein. However, small angle X-ray scattering (SAXS) revealed that the multi-ACP fragment is an elongated monomer which does not form a globular unit. Stokes radii calculated from atomic monomeric SAXS models were comparable to those measured by analytical gel filtration chromatography, showing that in the gel filtration experiment, the molecular weight was overestimated due to the elongated protein shape. Thermal denaturation monitored by circular dichroism showed that unfolding of the tandem construct was not cooperative, and that the tandem arrangement did not stabilize the protein. Taken together, these data are consistent with an elongated beads-on-a-string arrangement of the tandem ACP domains in PUFA synthases, and speak against synergistic biocatalytic effects promoted by quaternary structuring. Thus, it is possible to envision bioengineering strategies which simply involve the artificial linking of multiple ACP domains for increasing the yield of fatty acids in bacterial cultures.

Up-regulation of the neuronal nicotinic receptor α7 by HIV glycoprotein 120: potential implications for HIV-associated neurocognitive disorder.

Approximately 30-50% of the >30 million HIV-infected subjects develop neurological complications ranging from mild symptoms to dementia. HIV does not infect neurons, and the molecular mechanisms behind HIV-associated neurocognitive decline are not understood. There are several hypotheses to explain the development of dementia in HIV(+) individuals, including neuroinflammation mediated by infected microglia and neuronal toxicity by HIV proteins. A key protein associated with the neurological complications of HIV, gp120, forms part of the viral envelope and can be found in the CSF of infected individuals. HIV-1-gp120 interacts with several receptors including CD4, CCR5, CXCR4, and nicotinic acetylcholine receptors (nAChRs). However, the role of nAChRs in HIV-associated neurocognitive disorder has not been investigated. We studied the effects of gp120(IIIB) on the expression and function of the nicotinic receptor α7 (α7-nAChR). Our results show that gp120, through activation of the CXCR4 chemokine receptor, induces a functional up-regulation of α7-nAChRs. Because α7-nAChRs have a high permeability to Ca(2+), we performed TUNEL staining to investigate the effects of receptor up-regulation on cell viability. Our data revealed an increase in cell death, which was blocked by the selective antagonist α-bungarotoxin. The in vitro data are supported by RT-PCR and Western blot analysis, confirming a remarkable up-regulation of the α7-nAChR in gp120-transgenic mice brains. Specifically, α7-nAChR up-regulation is observed in mouse striatum, a region severely affected in HIV(+) patients. In summary, CXCR4 activation induces up-regulation of α7-nAChR, causing cell death, suggesting that α7-nAChR is a previously unrecognized contributor to the neurotoxicity associated with HIV infection.