Ming Lei Guo

Cocaine Hijacks σ1 Receptor to Initiate Induction of Activated Leukocyte Cell Adhesion Molecule: Implication for Increased Monocyte Adhesion and Migration in the CNS

Human immunodeficiency virus (HIV)-associated increase in monocyte adhesion and trafficking is exacerbated by cocaine abuse. The underlying mechanisms involve cocaine-mediated upregulation of adhesion molecules with subsequent disruption of the blood–brain barrier (BBB). Recently, a novel activated leukocyte cell adhesion molecule (ALCAM) has been implicated in leukocyte transmigration across the endothelium. We now show that upregulation of ALCAM in the brain endothelium seen in HIV+/cocaine drug abusers paralleled increased CD68 immunostaining compared with HIV+/no cocaine or uninfected controls, suggesting the important role of ALCAM in promoting leukocyte infiltration across the BBB. Furthermore, ALCAM expression was increased in cocaine-treated mice with concomitant increase in monocyte adhesion and transmigration in vivo, which was ameliorated by pretreating with the neutralizing antibody to ALCAM, lending additional support to the role of ALCAM. This new concept was further confirmed by in vitro experiments. Cocaine-mediated induction of ALCAM in human brain microvascular endothelial cells through the translocation of σ receptor to the plasma membrane, followed by phosphorylation of PDGF-β (platelet-derived growth factor-β) receptor. Downstream activation of mitogen-activated protein kinases, Akt, and NF-κB (nuclear factor-κB) pathways resulted in induced expression of ALCAM. Functional implication of upregulated ALCAM was confirmed using cell adhesion and transmigration assays. Neutralizing antibody to ALCAM ameliorated this effect. Together, these findings implicate cocaine-mediated induction of ALCAM as a mediator of increased monocyte adhesion/transmigration into the CNS.

Interplay of endoplasmic reticulum stress and autophagy in neurodegenerative disorders

ABSTRACT The common underlying feature of most neurodegenerative diseases such as Alzheimer disease (AD), prion diseases, Parkinson disease (PD), and amyotrophic lateral sclerosis (ALS) involves accumulation of misfolded proteins leading to initiation of endoplasmic reticulum (ER) stress and stimulation of the unfolded protein response (UPR). Additionally, ER stress more recently has been implicated in the pathogenesis of HIV-associated neurocognitive disorders (HAND). Autophagy plays an essential role in the clearance of aggregated toxic proteins and degradation of the damaged organelles. There is evidence that autophagy ameliorates ER stress by eliminating accumulated misfolded proteins. Both abnormal UPR and impaired autophagy have been implicated as a causative mechanism in the development of various neurodegenerative diseases. This review highlights recent advances in the field on the role of ER stress and autophagy in AD, prion diseases, PD, ALS and HAND with the involvement of key signaling pathways in these processes and implications for future development of therapeutic strategies.

Cocaine and HIV-1 Interplay in CNS: Cellular and Molecular Mechanisms

Although antiretrovirals are the mainstay of therapy against HIV infection, neurological complications associated with the virus continue to hamper quality of life of the infected individuals. Drugs of abuse in the infected individuals further fuel the epidemic. Epidemiological studies have demonstrated that abuse of cocaine resulted in acceleration of HIV infection and the progression of NeuroAIDS. Cocaine has not only been shown to play a crucial role in promoting virus replication, but also has diverse but often deleterious effects on various cell types of the CNS. In the neuronal system, cocaine exposure results in neuronal toxicity and also potentiates gp120-induced neurotoxicity. In the astroglia and microglia, cocaine exposure leads to up-regulation of pro-inflammatory mediators such as cytokines and chemokines. These in turn, can lead to neuroinflammation and transmission of toxic responses to the neurons. Additionally, cocaine exposure can also lead to leakiness of the blood-brain barrier that manifests as enhanced transmigraiton of leukocytes/monocytes into the CNS. Both in vitro and in vivo studies have provided valuable tools in exploring the role of cocaine in mediating HIV-associated neuropathogenesis. This review summarizes previous studies on the mechanism(s) underlying the interplay of cocaine and HIV as it relates to the CNS.

Cocaine-mediated microglial activation involves the ER stress-autophagy axis.

Cocaine abuse leads to neuroinflammation, which, in turn, contributes to the pathogenesis of neurodegeneration associated with advanced HIV-1 infection. Autophagy plays important roles in both innate and adaptive immune responses. However, the possible functional link between cocaine and autophagy has not been explored before. Herein, we demonstrate that cocaine exposure induced autophagy in both BV-2 and primary rat microglial cells as demonstrated by a dose- and time-dependent induction of autophagy-signature proteins such as BECN1/Beclin 1, ATG5, and MAP1LC3B. These findings were validated wherein cocaine treatment of BV-2 cells resulted in increased formation of puncta in cells expressing either endogenous MAP1LC3B or overexpressing GFP-MAP1LC3B. Specificity of cocaine-induced autophagy was confirmed by treating cells with inhibitors of autophagy (3-MA and wortmannin). Intriguingly, cocaine-mediated induction of autophagy involved upstream activation of 2 ER stress pathways (EIF2AK3- and ERN1-dependent), as evidenced by the ability of the ER stress inhibitor salubrinal to ameliorate cocaine-induced autophagy. In vivo validation of these findings demonstrated increased expression of BECN1, ATG5, and MAP1LC3B-II proteins in cocaine-treated mouse brains compared to untreated animals. Increased autophagy contributes to cocaine-mediated activation of microglia since pretreatment of cells with wortmannin resulted in decreased expression and release of inflammatory factors (TNF, IL1B, IL6, and CCL2) in microglial cells. Taken together, our findings suggest that cocaine exposure results in induction of autophagy that is closely linked with neuroinflammation. Targeting autophagic proteins could thus be considered as a therapeutic strategy for the treatment of cocaine-related neuroinflammation diseases.

Post-Translational Modification Biology of Glutamate Receptors and Drug Addiction

Post-translational covalent modifications of glutamate receptors remain a hot topic. Early studies have established that this family of receptors, including almost all ionotropic and metabotropic glutamate receptor subtypes, undergoes active phosphorylation at serine, threonine, or tyrosine residues in their intracellular domains. Recent evidence identifies several glutamate receptor subtypes to be direct substrates for palmitoylation at cysteine residues. Other modifications such as ubiquitination and sumoylation at lysine residues also occur to certain glutamate receptors. These modifications are dynamic and reversible in nature and are regulatable by changing synaptic inputs. The regulated modifications significantly impact the receptor in many ways, including interrelated changes in biochemistry (synthesis, subunit assembling, and protein–protein interactions), subcellular redistribution (trafficking, endocytosis, synaptic delivery, and clustering), and physiology, usually associated with changes in synaptic plasticity. Glutamate receptors are enriched in the striatum and cooperate closely with dopamine to regulate striatal signaling. Emerging evidence shows that modification processes of striatal glutamate receptors are sensitive to addictive drugs, such as psychostimulants (cocaine and amphetamine). Altered modifications are believed to be directly linked to enduring receptor/synaptic plasticity and drug-seeking. This review summarizes several major types of modifications of glutamate receptors and analyzes the role of these modifications in striatal signaling and in the pathogenesis of psychostimulant addiction.

Roles of subunit phosphorylation in regulating glutamate receptor function.

Protein phosphorylation is an important mechanism for regulating ionotropic glutamate receptors (iGluRs). Early studies have established that major iGluR subtypes, including α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors and N-methyl-d-aspartate (NMDA) receptors, are subject to phosphorylation. Multiple serine, threonine, and tyrosine residues predominantly within the C-terminal regions of AMPA receptor and NMDA receptor subunits have been identified as sensitive phosphorylation sites. These distinct sites undergo either constitutive phosphorylation or activity-dependent phosphorylation induced by changing cellular and synaptic inputs. An increasing number of synapse-enriched protein kinases have been found to phosphorylate iGluRs The common kinases include protein kinase A, protein kinase C, Ca(2+)/calmodulin-dependent protein kinase II, Src/Fyn non-receptor tyrosine kinases, and cyclin dependent kinase-5. Regulated phosphorylation plays a well-documented role in modulating the biochemical, biophysical, and functional properties of the receptor. In the future, identifying the precise mechanisms how phosphorylation regulates iGluR activities and finding the link between iGluR phosphorylation and the pathogenesis of various brain diseases, including psychiatric and neurodegenerative diseases, chronic pain, stroke, Alzheimers disease and substance addiction, will be hot topics and could contribute to the development of novel pharmacotherapies, by targeting the defined phosphorylation process, for suppressing iGluR-related disorders.

Cocaine and HIV-1 Interplay: Molecular Mechanisms of Action and Addiction

Human immunodeficiency virus (HIV) infection is now being driven by drug-abusing populations. Epidemiological studies on drug abusers with AIDS link abuse of cocaine, even more than other drugs, to increased incidence of HIV seroprevalence and progression to AIDS. Both cell culture and animal studies demonstrate that cocaine can both potentiate HIV replication and can potentiate HIV proteins to cause enhanced glial cell activation, neurotoxicity, and breakdown of the blood-brain barrier. Based on the ability of both HIV proteins and cocaine to modulate NMDA receptor on neurons, NMDA receptors have been suggested as a common link underlying the crosstalk between drug addiction and HIV infection. While the role of dopamine system as a major target of cocaine cannot be overlooked, recent studies on the role of sigma receptors in mediating the effects of cocaine in both cell and organ systems warrants a deeper understanding of their functional role in the field. In this review, recent findings on the interplay of HIV infection and cocaine abuse and their possible implications in mode of action and/or addiction will be discussed.

Phosphorylation and Feedback Regulation of Metabotropic Glutamate Receptor 1 by Calcium/Calmodulin-Dependent Protein Kinase II

The metabotropic glutamate receptor 1 (mGluR1) is a Gαq-protein-coupled receptor and is distributed in broad regions of the mammalian brain. As a key element in excitatory synaptic transmission, the receptor regulates a wide range of cellular and synaptic activities. In addition to regulating its targets, the receptor itself is believed to be actively regulated by intracellular signals, although underlying mechanisms are essentially unknown. Here we found that a synapse-enriched protein kinase, Ca2+/calmodulin-dependent protein kinase IIα (CaMKIIα), directly binds to the intracellular C terminus (CT) of mGluR1a. This binding is augmented by Ca2+ in vitro. The direct interaction promotes CaMKIIα to phosphorylate mGluR1a at a specific threonine site (T871). In rat striatal neurons, the mGluR1 agonist triggers the receptor-associated phosphoinositide signaling pathway to induce Ca2+-dependent recruitment of CaMKIIα to mGluR1a–CT. This enables the kinase to inhibit the response of the receptor to subsequent agonist exposure. Our data identify an agonist-induced and Ca2+-dependent protein–protein interaction between a synaptic kinase and mGluR1, which constitutes a feedback loop facilitating desensitization of mGluR1a.

CaMKIIα interacts with M4 muscarinic receptors to control receptor and psychomotor function

Muscarinic acetylcholine receptors (mAChRs) are widely expressed in the mammalian brain and are essential for neuronal functions. These receptors are believed to be actively regulated by intracellular signals, although the underlying mechanisms are largely unknown. In this study, we show that Ca2+/calmodulin‐dependent protein kinase II (CaMKII) binds directly and selectively to one of five mAChR subtypes, M4 receptors (M4Rs), at their C‐terminal regions of second intracellular loops. This binding relies on Ca2+ activation of the kinase and leads to the phosphorylation of M4Rs at a specific threonine site (Thr145). Complementary in vivo studies in rat striatal neurons enriched with M4Rs confirm that rising Ca2+ recruits CaMKIIα to M4Rs to potentiate receptor signalling, which controls behavioural sensitivity to dopamine stimulation in an activity‐dependent manner. Our data identify a new model of protein–protein interactions. In a Ca2+‐sensitive manner, CaMKIIα regulates M4R efficacy and controls the acetylcholine–dopamine balance in the basal ganglia and also the dynamics of movement.

Cocaine induces astrocytosis through ER stress-mediated activation of autophagy

ABSTRACT Cocaine is known to induce inflammation, thereby contributing in part, to the pathogenesis of neurodegeneration. A recent study from our lab has revealed a link between macroautophagy/autophagy and microglial activation. The current study was aimed at investigating whether cocaine could also mediate activation of astrocytes and, whether this process involved induction of autophagy. Our findings demonstrated that cocaine mediated the activation of astrocytes by altering the levels of autophagy markers, such as BECN1, ATG5, MAP1LC3B-II, and SQSTM1 in both human A172 astrocytoma cells and primary human astrocytes. Furthermore, cocaine treatment resulted in increased formation of endogenous MAP1LC3B puncta in human astrocytes. Additionally, astrocytes transfected with the GFP-MAP1LC3B plasmid also demonstrated cocaine-mediated upregulation of the green fluorescent MAP1LC3B puncta. Cocaine-mediated induction of autophagy involved upstream activation of ER stress proteins such as EIF2AK3, ERN1, ATF6 since blockage of autophagy using either pharmacological or gene-silencing approaches, had no effect on cocaine-mediated induction of ER stress. Using both pharmacological and gene-silencing approaches to block either ER stress or autophagy, our findings demonstrated that cocaine-induced activation of astrocytes (measured by increased levels of GFAP) involved sequential activation of ER stress and autophagy. Cocaine-mediated-increased upregulation of GFAP correlated with increased expression of proinflammatory mediators such as TNF, IL1B, and IL6. In conclusion, these findings reveal an association between ER stress-mediated autophagy and astrogliosis in cocaine-treated astrocytes. Intervention of ER stress and/or autophagy signaling would thus be promising therapeutic targets for abrogating cocaine-mediated neuroinflammation.