Serum amyloid P component (SAP) modulates antidepressant effects through promoting membrane insertion of the serotonin transporter.

Serum amyloid P component (SAP) is a universal constituent of human amyloid deposits including those in Alzheimer's disease. SAP has been observed to be elevated in patients with depression, and higher SAP levels are associated with better response to the antidepressant escitalopram. The mechanisms underlying these clinical observations remain unclear. We examined the effect of SAP on serotonin transporter (SERT) expression and localization using Western blot, confocal microscopy, and positron emission tomography with the radioligand [11C]DASB. We also investigated the effect of SAP on treatment response to escitalopram in mice with the forced swim test (FST), a classical behaviour paradigm to assess antidepressant effects. SAP reduced [11C]DASB binding as an index of SERT levels, consistent with Western blots showing decreased total SAP protein because of increased protein degradation. In conjunction with the global decrease in SERT levels, SAP also promotes VAMP-2 mediated SERT membrane insertion. SAP levels are correlated with behavioural despair and SSRI treatment response in mice with FST. In MDD patients, the SAP and membrane SERT levels are correlated with response to SSRI treatment. SAP has complex effects on SERT levels and localization, thereby modulating the effect of SSRIs, which could partially explain clinical variability in antidepressant treatment response. These results add to our understanding of the mechanism for antidepressant drug action, and with further work could be of clinical utility.

Endosomal recycling and dopamine neurotransmission: Exploring the links between the retromer and Parkinson's disease.

The retromer complex is an evolutionarily conserved protein complex involved in the endosomal recycling of various cargo proteins. It is ubiquitously expressed in all tissue and is found in both invertebrate as well as mammalian nervous systems, where it recycles various synaptic membrane proteins including the dopamine transporter and dopamine D1 receptor, two proteins implicated in dopamine homeostasis and neurotransmission. The involvement of the retromer complex in dopamine neurobiology is further underscored by its links to Parkinson's disease, a neurodegenerative disorder of the dopamine system. In this article, the existing literature linking the retromer complex to synaptic function and dopamine homeostasis is reviewed. Additional possible links are highlighted by exploring the retromer and other Parkinson's disease-associated proteins and possible relationships to synaptic function and dopamine transmission.

Learning to balance on a slackline: Development of coordinated multi-joint synergies.

Previous research has investigated synergies involved in locomotion and balance reactions; however, there is limited insight into the emergence of skilled balance control with practice of challenging tasks. We explored motor learning of tandem and single leg stance on an unstable surface-a slackline. Balance was tested in 10 naïve healthy adults at four time points: baseline, after one slackline practice session, after 1 week of practice, and 1 week following the final practice session. We recorded kinematics of the upper and lower arms bilaterally, trunk, and thigh and foot unilaterally while participants balanced in tandem and single leg stance on a slackline and narrow rigid beam (transfer task). When participants first attempted to stand on the slackline, they exhibited fast and frequent movements across all joints with actions along the frontal plane (particularly the hip) and fell after a short period (~3 seconds). Performance improved rapidly (fewer falls), and this was accompanied by dampened trunk and foot oscillations and the development of coordinated movement patterns with a progressive emphasis on more distal upper body segments. Continuous relative phase angles between joint pairs began to cluster around either 0° (indicating in-phase movement) or 180° (indicating anti-phase movement). Participants also began to demonstrate coordinated upper body synergies and performance improvements (fewer falls) on the transfer task, while a control group (n = 10) did not exhibit similar synergies or performance improvements. Our findings describe the emergence of coordinated movement synergies involving the upper body as healthy adults learn a challenging balance task.

Neuroprotective effects of extracellular DJ-1 on reperfusion injury in SH-SY5Y cells.

Using an in vitro model of ischemic stroke we treated differentiated SH-SY5Y cells to oxygen-glucose deprivation (OGD) followed by a reperfusion period where normal growth conditions were restored. Cells undergoing OGD exhibited significant cell death as measure by propidium iodide staining. However, cells treated with exogenous extracellular DJ-1 during reperfusion exhibited significant rescue from OGD-induced cell death.

The retinoblastoma protein regulates hypoxia-inducible genetic programs, tumor cell invasiveness and neuroendocrine differentiation in prostate cancer cells.

Loss of tumor suppressor proteins, such as the retinoblastoma protein (Rb), results in tumor progression and metastasis. Metastasis is facilitated by low oxygen availability within the tumor that is detected by hypoxia inducible factors (HIFs). The HIF1 complex, HIF1α and dimerization partner the aryl hydrocarbon receptor nuclear translocator (ARNT), is the master regulator of the hypoxic response. Previously, we demonstrated that Rb represses the transcriptional response to hypoxia by virtue of its association with HIF1. In this report, we further characterized the role Rb plays in mediating hypoxia-regulated genetic programs by stably ablating Rb expression with retrovirally-introduced short hairpin RNA in LNCaP and 22Rv1 human prostate cancer cells. DNA microarray analysis revealed that loss of Rb in conjunction with hypoxia leads to aberrant expression of hypoxia-regulated genetic programs that increase cell invasion and promote neuroendocrine differentiation. For the first time, we have established a direct link between hypoxic tumor environments, Rb inactivation and progression to late stage metastatic neuroendocrine prostate cancer. Understanding the molecular pathways responsible for progression of benign prostate tumors to metastasized and lethal forms will aid in the development of more effective prostate cancer therapies.

A Physical Interaction between the Dopamine Transporter and DJ-1 Facilitates Increased Dopamine Reuptake.

The regulation of the dopamine transporter (DAT) impacts extracellular dopamine levels after release from dopaminergic neurons. Furthermore, a variety of protein partners have been identified that can interact with and modulate DAT function. In this study we show that DJ-1 can potentially modulate DAT function. Co-expression of DAT and DJ-1 in HEK-293T cells leads to an increase in [3H] dopamine uptake that does not appear to be mediated by increased total DAT expression but rather through an increase in DAT cell surface localization. In addition, through a series of GST affinity purifications and co-immunoprecipitations, we provide evidence that the DAT can be found in a complex with DJ-1, which involve distinct regions within both DAT and DJ-1. Using in vitro binding experiments we also show that this complex can be formed in part by a direct interaction between DAT and DJ-1. Co-expression of a mini-gene that can disrupt the DAT/DJ-1 complex appears to block the increase in [3H] dopamine uptake by DJ-1. Mutations in DJ-1 have been linked to familial forms of Parkinson's disease, yet the normal physiological function of DJ-1 remains unclear. Our study suggests that DJ-1 may also play a role in regulating dopamine levels by modifying DAT activity.

Delineation of domains within the cannabinoid CB1 and dopamine D2 receptors that mediate the formation of the heterodimer complex.

Both the cannabinoid CB1 receptor (CB1) and dopamine D2 receptor (D2R) are G protein-coupled receptors that are linked to inhibitory Gαi/o protein, whereby activation of the receptor leads to the inhibition of cAMP production. Moreover, previous findings have shown evidence of cross-talk between the dopamine and endocannabinoid systems. In this report, we confirm the interaction of CB1 and D2R with co-immunoprecipitation experiments using human embryonic kidney 293T (HEK-293T) cells co-expressing both receptors. We also generated GST and His-tagged fusion proteins of the D2R and CB1 and conducted affinity purification assays and in vitro binding experiments to show that the CB1-D2R complex can be formed by a direct protein-protein interaction. This interaction is mediated by the carboxyl terminus of the CB1 receptor and the third intracellular loop of the D2 receptor. Co-transfection of an inhibitory mini-gene resulted in decreased levels of the CB1-D2R complex. Using a cAMP biosensor, we show that activation of D2R or CB1 alone in HEK-293T cells co-expressing both receptors leads to an inhibition of forskolin-stimulated cAMP accumulation. However, co-activation of both receptors resulted in a loss of this inhibition on cAMP accumulation. Our findings characterize the physical interaction between CB1 and D2R as well as demonstrate the potential functional outcome of the receptor complex.

Adaptation of gait termination on a slippery surface in Parkinson's disease.

Parkinson's disease (PD) causes instability and difficulty adapting to changing environmental and task demands. We examined the effects of PD on the adaptation of gait termination (GT) on a slippery surface under unexpected and cued circumstances. An unexpected slip perturbation during GT was followed by a slip perturbation during GT under two conditions: planned over multiple steps and cued one step prior to GT. Feed forward and feedback-based responses to the perturbation were compared to determine (1) how PD affects the ability to integrate adaptive feed forward and feedback-based GT strategies on a slippery surface, (2) if adaptations can be implemented when GT is required within one step, and (3) if behaviour changes with repeated exposure. Similar to the control group (n=10), the PD group (n=8) adapted and integrated feed forward and feedback-based components of GT under both stop conditions. Feed forward adaptations included a shorter, wider step, and appropriate stability margin modifications. Feedback-based adaptations included a longer, wider subsequent step. When cued to stop quickly, both groups maintained most of these adaptations: foot angle at contact increased in the first cued stop but adapted with practice. The group with PD differed in their ability to adapt GT with slower, wider steps and less stability.

Direct interaction between GluR2 and GAPDH regulates AMPAR-mediated excitotoxicity.

Over-activation of AMPARs (α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid subtype glutamate receptors) is implicated in excitotoxic neuronal death associated with acute brain insults, such as ischemic stroke. However, the specific molecular mechanism by which AMPARs, especially the calcium-impermeable AMPARs, induce neuronal death remains poorly understood. Here we report the identification of a previously unrecognized molecular pathway involving a direct protein-protein interaction that underlies GluR2-containing AMPAR-mediated excitotoxicity. Agonist stimulation of AMPARs promotes GluR2/GAPDH (glyceraldehyde-3-phosphate dehydrogenase) complex formation and subsequent internalization. Disruption of GluR2/GAPDH interaction by administration of an interfering peptide prevents AMPAR-mediated excitotoxicity and protects against damage induced by oxygen-glucose deprivation (OGD), an in vitro model of brain ischemia.

Uncoupling the D1-N-methyl-D-aspartate (NMDA) receptor complex promotes NMDA-dependent long-term potentiation and working memory.

BACKGROUND: Although dopamine D1 receptors are involved in working memory, how D1 receptors contribute to this process remains unclear. Numerous studies have shown that D1 receptors have extensive functional interaction with N-methyl-D-aspartate (NMDA) receptor. Our group previously demonstrated that D1 receptors were able to regulate NMDA receptor functions through direct protein-protein interactions involving the carboxyl terminals of D1 receptors and NMDA receptor NR1a and NR2A subunits respectively. In this study, we explored the effects of the D1-NR1 interaction on NMDA receptor-dependent long-term potentiation (LTP) and working memory by using the TAT-conjugated interfering peptide (TAT-D1-t2). METHODS: Miniature excitatory postsynaptic currents are recorded in rat hippocampal primary cultures. Coimmunoprecipitation and calcium/calmodulin-dependent protein kinase II (CaMKII) activity are measured in hippocampal slices and hippocampal neurons under the specified experimental conditions, respectively. Working memory was assessed using a delayed match-to-place protocol in the Morris Water Maze following administration of the TAT-D1-t2 peptide. RESULTS: Electrophysiology experiments showed that activation of D1 receptor upregulates NMDA receptor-mediated LTP in a CaMKII-dependent manner. Furthermore, D1 receptor agonist stimulation promotes the NR1-CaMKII coupling and enhances the CaMKII activity; and the D1 receptor-mediated effects can be blocked by the application of the TAT-D1-t2 peptide. Interestingly, animals injected with TAT-D1-t2 peptide exhibited significantly impaired working memory. CONCLUSIONS: Our study showed a critical role of NMDA-D1 direct protein-protein interaction in NMDA receptor-mediated LTP and working memory and implicated the involvement of CaMKII in this process.

NCS-1 in the dentate gyrus promotes exploration, synaptic plasticity, and rapid acquisition of spatial memory.

The molecular underpinnings of exploration and its link to learning and memory remain poorly understood. Here we show that inducible, modest overexpression of neuronal calcium sensor 1 (Ncs1) selectively in the adult murine dentate gyrus (DG) promotes a specific form of exploratory behavior. The mice also display a selective facilitation of long-term potentiation (LTP) in the medial perforant path and a selective enhancement in rapid-acquisition spatial memory, phenotypes that are reversed by direct application of a cell-permeant peptide (DNIP) designed to interfere with NCS-1 binding to the dopamine type-2 receptor (D2R). Moreover, the DNIP and the D2R-selective antagonist L-741,626 attenuated exploratory behavior, DG LTP, and spatial memory in control mice. These data demonstrate a role for NCS-1 and D2R in DG plasticity and provide insight for understanding how the DG contributes to the origin of exploration and spatial memory acquisition.

New measurement of the K+-->pi+ nunu branching ratio.

Three events for the decay K+-->pi+ nunu have been observed in the pion momentum region below the K+-->pi+pi0 peak, 140 < Ppi < 199 MeV/c, with an estimated background of 0.93+/-0.17(stat.) -0.24+0.32(syst.) events. Combining this observation with previously reported results yields a branching ratio of B(K+-->pi+ nunu) = (1.73(-1.05)+1.15) x 10(-10) consistent with the standard model prediction.

FMRP acts as a key messenger for dopamine modulation in the forebrain.

The fragile X mental retardation protein (FMRP) is an RNA-binding protein that controls translational efficiency and regulates synaptic plasticity. Here, we report that FMRP is involved in dopamine (DA) modulation of synaptic potentiation. AMPA glutamate receptor subtype 1 (GluR1) surface expression and phosphorylation in response to D1 receptor stimulation were reduced in cultured Fmr1(-/-) prefrontal cortex (PFC) neurons. Furthermore, D1 receptor signaling was impaired, accompanied by D1 receptor hyperphosphorylation at serine sites and subcellular redistribution of G protein-coupled receptor kinase 2 (GRK2) in both PFC and striatum of Fmr1(-/-) mice. FMRP interacted with GRK2, and pharmacological inhibition of GRK2 rescued D1 receptor signaling in Fmr1(-/-) neurons. Finally, D1 receptor agonist partially rescued hyperactivity and enhanced the motor function of Fmr1(-/-) mice. Our study has identified FMRP as a key messenger for DA modulation in the forebrain and may provide insights into the cellular and molecular mechanisms underlying fragile X syndrome.

Dopamine receptor interacting proteins (DRIPs) of dopamine D1-like receptors in the central nervous system.

Dopamine is a major neurotransmitter in the mammalian central nervous system (CNS) that regulates neuroendocrine functions, locomotor activity, cognition and emotion. The dopamine system has been extensively studied because dysfunction of this system is linked to various pathological conditions including Parkinson's disease, schizophrenia, Tourette's syndrome, and drug addiction. Accordingly, intense efforts to delineate the full complement of signaling pathways mediated by individual receptor subtypes have been pursued. Dopamine D1-like receptors are of particular interest because they are the most abundant dopamine receptors in CNS. Recent work suggests that dopamine signaling could be regulated via dopamine receptor interacting proteins (DRIPs). Unraveling these DRIPs involved in the dopamine system may provide a better understanding of the mechanisms underlying CNS disorders related to dopamine system dysfunction and may help identify novel therapeutic targets.

Genetic factors involved in the pathogenesis of Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disease characterized by a loss of nigrostriatal dopaminergic neurons. Recently, PD research has been stimulated by the identification of genes that are implicated in rare familial forms of PD. However, despite these discoveries, the primary cause of PD is still unclear. Various pathogenic mechanisms may be involved including mitochondrial dysfunction, proteasomal dysfunction/protein aggregation, oxidative damage, environmental factors and genetic disposition. Furthermore, dopamine has also been implicated in contributing to the pathogenesis of PD. This review will focus on the genes that have been identified to be associated with PD and how they may impair dopamine metabolism. Understanding the role of these PD-related genes in dopamine neurobiology may provide insight into the underpinning pathogenic mechanisms of PD.

Compensatory postural adaptations during continuous, variable amplitude perturbations reveal generalized rather than sequence-specific learning.

We examined changes in the motor organization of postural control in response to continuous, variable amplitude oscillations evoked by a translating platform and explored whether these changes reflected implicit sequence learning. The platform underwent random amplitude (maximum +/- 15 cm) and constant frequency (0.5 Hz) oscillations. Each trial was composed of three 15-s segments containing seemingly random oscillations. Unbeknownst to participants, the middle segment was repeated in each of 42 trials on the first day of testing and in an additional seven trials completed approximately 24 h later. Kinematic data were used to determine spatial and temporal components of total body centre of mass (COM) and joint segment coordination. Results showed that with repeated trials, participants reduced their magnitude of COM displacement, shifted from a COM phase lag to a phase lead relative to platform motion and increased correlations between ankle/platform motion and hip/platform motion as they shifted from an ankle strategy to a multi-segment control strategy involving the ankle and hip. Maintenance of these changes across days provided evidence for learning. Similar improvements for the random and repeated segments, indicated that participants did not exploit the sequence of perturbations to improve balance control. Rather, the central nervous system may have been tuning into more general features of platform motion. These findings provide important insight into the generalizabilty of improved compensatory balance control with training.

Parkin disrupts the alpha-synuclein/dopamine transporter interaction: consequences toward dopamine-induced toxicity.

Parkinson's disease is characterized by progressive neuronal degeneration of dopaminergic neurons in the substantia nigra. Many factors are thought to contribute to the neuronal cell death that occurs in Parkinson's disease, including alpha-synuclein-mediated toxicity. Previously, we have reported that alpha-synuclein directly couples to the carboxyl tail of the dopamine transporter (DAT) and that the alpha-synuclein/DAT protein complex formation accelerates DAT-mediated cellular dopamine (DA) uptake and DA-induced cellular apoptosis. In the present study, we report that parkin, an E2-dependent E3 protein ubiquitin ligase associated with recessive early onset Parkinson's disease, exerts a protective effect against DA-induced alpha-synuclein-dependent cell toxicity. Parkin impairs the alpha-synuclein/DAT coupling by interacting with the carboxyl-terminus of the DAT and blocks the alpha-synuclein-induced enhancement in both DAT cell surface expression and DAT-mediated DA uptake. Moreover, we have found that parkin protects against DA-induced cell toxicity in dopaminergic SK-N-SH cells. These findings will help identify the role of these proteins in the etiology and/or maintenance of Parkinson's disease.

Dopamine transporter cell surface localization facilitated by a direct interaction with the dopamine D2 receptor.

Altered synaptic dopamine levels have been implicated in several neurological/neuropsychiatric disorders, including drug addiction and schizophrenia. However, it is unclear what precipitates these changes in synaptic dopamine levels. One of the key presynaptic components involved in regulating dopaminergic tone is the dopamine transporter (DAT). Here, we report that the DAT is also regulated by the dopamine D2 receptor through a direct protein-protein interaction involving the DAT amino-terminus and the third intracellular loop of the D2 receptor. This physical coupling facilitates the recruitment of intracellular DAT to the plasma membrane and leads to enhanced dopamine reuptake. Moreover, mice injected with peptides that disrupt D2-DAT interaction exhibit decreased synaptosomal dopamine uptake and significantly increased locomotor activity, reminiscent of DAT knockout mice. Our data highlight a novel mechanism through which neurotransmitter receptors can functionally modulate neurotransmitter transporters, an interaction that can affect the synaptic neurotransmitter levels in the brain.

Direct receptor cross-talk can mediate the modulation of excitatory and inhibitory neurotransmission by dopamine.

Dopamine is a neurotransmitter that can regulate both excitatory and inhibitory fast synaptic transmission. The overlapping dopaminergic, glutamatergic, and GABAergic systems provide a basis for the interaction between these three neurotransmitters. Although there is considerable evidence for the involvement of second-messenger systems to mediate receptor cross-talk between these receptor systems, there is emerging evidence that receptors can interact through direct protein-protein interactions. The functional implications and overall significance of the dopamine/glutamate/GABA interactions will be examined.