The association of spinophilin with disks large-associated protein 3 (SAPAP3) is regulated by metabotropic glutamate receptor (mGluR) 5

Spinophilin is the most abundant protein phosphatase 1 targeting protein in the postsynaptic density of dendritic spines. Spinophilin associates with myriad synaptic proteins to regulate normal synaptic communication; however, the full complement of spinophilin interacting proteins and mechanisms regulating spinophilin interactions are unclear. Here we validate an association between spinophilin and the scaffolding protein, disks large-associated protein 3 (SAP90/PSD-95 associated protein 3; SAPAP3). Loss of SAPAP3 leads to obsessive-compulsive disorder (OCD)-like behaviors due to alterations in metabotropic glutamate receptor (mGluR) signaling. Here we report that spinophilin associates with SAPAP3 in the brain and in a heterologous cell system. Moreover, we have found that expression or activation of group I mGluRs along with activation of the mGluR-dependent kinase, protein kinase C β, enhances this interaction. Functionally, global loss of spinophilin attenuates amphetamine-induced hyperlocomotion, a striatal behavior associated with dopamine dysregulation and OCD. Together, these data delineate a novel link between mGluR signaling, spinophilin, and SAPAP3 in striatal pathophysiology

Estrus cyclicity of spinogenesis: underlying mechanisms

Hippocampal spine density varies with the estrus cycle. The cyclic change in estradiol levels in serum was hypothesized to underlie this phenomenon, since treatment of ovariectomized animals with estradiol induced an increase in spine density in hippocampal dendrites of rats, as compared to ovariectomized controls. In contrast, application of estradiol to hippocampal slice cultures did not promote spinogenesis. In addressing this discrepancy, we found that hippocampal neurons themselves are capable of synthesizing estradiol de novo. Estradiol synthesis can be suppressed by aromatase inhibitors and by knock-down of Steroid Acute Regulatory Protein (StAR) and enhanced by substrates of steroidogenesis. Expression of estrogen receptors (ERs) and synaptic proteins, synaptogenesis, and long-term potentiation (LTP) correlated positively with aromatase activity in hippocampal cultures without any difference between genders. All effects due to inhibition of aromatase activity were rescued by application of estradiol to the cultures. Most importantly, gonadotropin-releasing hormone (GnRH) increased estradiol synthesis dose-dependently via an aromatase-mediated mechanism and consistently increased spine synapse density and spinophilin expression. As a consequence, our data suggest that cyclic fluctuations in spine synapse density result from pulsative release of GnRH from the hypothalamus and its effect on hippocampal estradiol synthesis, rather than from varying levels of serum estradiol. This hypothesis is further supported by higher GnRH receptor (GnRH-R) density in the hippocampus than in the cortex and hypothalamus and the specificity of estrus cyclicity of spinogenesis in the hippocampus, as compared to the cortex

Modulation of dopamine D1 receptors via histamine H3 receptors is a novel therapeutic target for Huntington's disease

Early Huntington's disease (HD) include over-activation of dopamine D1 receptors (D1R), producing an imbalance in dopaminergic neurotransmission and cell death. To reduce D1R over-activation, we present a strategy based on targeting complexes of D1R and histamine H3 receptors (H3R). Using an HD mouse striatal cell model and HD mouse organotypic brain slices we found that D1R-induced cell death signaling and neuronal degeneration, are mitigated by an H3R antagonist. We demonstrate that the D1R-H3R heteromer is expressed in HD mice at early but not late stages of HD, correlating with HD progression. In accordance, we found this target expressed in human control subjects and low-grade HD patients. Finally, treatment of HD mice with an H3R antagonist prevented cognitive and motor learning deficits and the loss of heteromer expression. Taken together, our results indicate that D1R - H3R heteromers play a pivotal role in dopamine signaling and represent novel targets for treating HD

Abnormal social behavior, hyperactivity, impaired remote spatial memory, and increased D1-mediated dopaminergic signaling in neuronal nitric oxide synthase knockout mice

<p>Abstract</p> <p>Background</p> <p>Neuronal nitric oxide synthase (nNOS) is involved in the regulation of a diverse population of intracellular messenger systems in the brain. In humans, abnormal NOS/nitric oxide metabolism is suggested to contribute to the pathogenesis and pathophysiology of some neuropsychiatric disorders, such as schizophrenia and bipolar disorder. Mice with targeted disruption of the nNOS gene exhibit abnormal behaviors. Here, we subjected nNOS knockout (KO) mice to a battery of behavioral tests to further investigate the role of nNOS in neuropsychiatric functions. We also examined the role of nNOS in dopamine/DARPP-32 signaling in striatal slices from nNOS KO mice and the effects of the administration of a dopamine D1 receptor agonist on behavior in nNOS KO mice.</p> <p>Results</p> <p>nNOS KO mice showed hyperlocomotor activity in a novel environment, increased social interaction in their home cage, decreased depression-related behavior, and impaired spatial memory retention. In striatal slices from nNOS KO mice, the effects of a dopamine D1 receptor agonist, SKF81297, on the phosphorylation of DARPP-32 and AMPA receptor subunit GluR1 at protein kinase A sites were enhanced. Consistent with the biochemical results, intraperitoneal injection of a low dose of SKF81297 significantly decreased prepulse inhibition in nNOS KO mice, but not in wild-type mice.</p> <p>Conclusion</p> <p>These findings indicate that nNOS KO upregulates dopamine D1 receptor signaling, and induces abnormal social behavior, hyperactivity and impaired remote spatial memory. nNOS KO mice may serve as a unique animal model of psychiatric disorders.</p

The Effects of Altered Prenatal Melatonin Signaling on Adult Behavior and Hippocampal Gene Expression of the Male Rat: A Circadioneuroendocrine-Axis Hypothesis of Psychopathology

Disturbances in melatonin - the neurohormone that signals environmental darkness as part of the circadian circuit of mammals - have been implicated in various psychopathologies in humans. At present, experimental evidence linking prenatal melatonin signaling to adult physiology, behavior, and gene expression is lacking. We hypothesized that administration of melatonin (5 mg/kg) or the melatonin receptor antagonist luzindole (5 mg/kg) to rats in utero would permanently alter the circadian circuit to produce differential growth, adult behavior, and hippocampal gene expressionin the male rat. Prenatal treatment was found to increase growth in melatonin-treated animals. In addition, subjects exposed to melatonin prenatally displayed increased rearing in the open field test and an increased right turn preference in the elevated plusmaze. Rats administered luzindole prenatally, however, displayed greater freezing and grooming behavior in the open field test and improved learning in the Morris water maze. Analysis of relative adult hippocampal gene expression with RT-PCR revealed increasedexpression of brain-derived neurotrophic factor (BDNF) with a trend toward increased expression of melatonin 1A (MEL1A) receptors in melatonin-exposed animals whereas overall prenatal treatment had a significant effect on microtubule-associated protein 2(MAP2) expression. Our data support the conclusion that the manipulation of maternal melatonin levels alters brain development and leads to physiological and behavioral abnormalities in adult offspring. We designate the term circadioneuroendocrine (CNE)axis and propose the CNE-axis hypothesis of psychopathology

Neurabin Contributes to Hippocampal Long-Term Potentiation and Contextual Fear Memory

Neurabin is a scaffolding protein that interacts with actin and protein phosphatase-1. Highly enriched in the dendritic spine, neurabin is important for spine morphogenesis and synaptic formation. However, less is known about the role of neurabin in hippocampal plasticity and its possible effect on behavioral functions. Using neurabin knockout (KO) mice, here we studied the function of neurabin in hippocampal synaptic transmission, plasticity and behavioral memory. We demonstrated that neurabin KO mice showed a deficit in contextual fear memory but not auditory fear memory. Whole-cell patch clamp recordings in the hippocampal CA1 neurons showed that long-term potentiation (LTP) was significantly reduced, whereas long-term depression (LTD) was unaltered in neurabin KO mice. Moreover, increased AMPA receptor but not NMDA receptor-mediated synaptic transmission was found in neurabin KO mice, and is accompanied by decreased phosphorylation of GluR1 at the PKA site (Ser845) but no change at the CaMKII/PKC site (Ser831). Pre-conditioning with LTD induction rescued the following LTP in neurabin KO mice, suggesting the loss of LTP may be due to the saturated synaptic transmission. Our results indicate that neurabin regulates contextual fear memory and LTP in hippocampal CA1 pyramidal neurons

Abnormal COX2/PGE2 Signalling in the Developing Cerebellum - A Link to Autism Spectrum Disorders

Autism spectrum disorders (ASDs) include a group of neurodevelopmental conditions that are characterized by deficits in social interaction and communication, increases in repetitive/restricted often stereotyped behaviour, and increases in anxiety. The heterogeneous nature of ASDs with regards to symptoms but also genetic profiles of ASD individuals, make understanding factors contributing to the disorder complex. However, literature suggests that ASDs arise from a combination of genetic and environmental factors. Clinical studies have suggested that abnormal lipid signalling, as a result of environmental insults can contribute to the etiology of ASDs. The phospholipid membrane of cells within the can be metabolized into lipid signalling molecules, including prostaglandins. Prostaglandin E2 (PGE2) is one of the most utilized lipid signalling molecules in the brain, involved in developmental processes such as synaptogenesis, migration, and differentiation of neuronal stem cells. Abnormal levels of PGE2, as well as COX-1 and COX-2, the rate-limiting enzymes in PGE2 synthesis have been linked to ASD. Furthermore, various environmental risk factors including exposure to heavy metals, infection/inflammation in pregnancy, exposure to pesticides, fragrances, and the use of over-the-counter medications such as aspirin and acetaminophen can affect PGE2 levels and are linked to ASD. The exact mechanisms that link abnormal COX2/PGE2 signalling to ASD are still unclear. To help address the lack of information, in this dissertation we first examine the effect of exposure to PGE2 on differentiated neuroectodermal (NE-4C) stem cells. Further, Studies have demonstrated that the cerebellum may be important in the etiology of ASDs. Interestingly there is evidence that PGE2 can affect postnatal development of the cerebellum. We examine the effect of increases (in a maternal PGE2 injection model) and decreases (in a COX-2--KI model) in PGE2 levels on prenatal neurodevelopment. We specifically examine the effects of these increases and decreases on cytoskeletal-dependent morphology through dendritic morphology within the cerebellum. Additionally, we examine the effect of prenatal PGE2-exposure on cerebellar-dependent motor function postnatally. Given the importance of sex as a factor in examining neurodevelopmental disorders such as ASD that have a large sex bias towards males, all of our in vivo studies address the modulation of the PGE2 effect by sex. These studies demonstrate that abnormal COX2/PGE2 signalling can affect important neurodevelopmental processes in vitro and development of the cerebellum in vivo. We observed disruptions in cytoskeletal dynamics, and changes in the expression of cytoskeletal proteins corresponding to abnormal COX2/PGE2 signalling. In PGE2-exposed mice, the changes in dendritic morphology in the cerebellum, corresponded to deficits in cerebellar motor function. Further, we found that the disruption of COX2/PGE2 affected development in a sex-dependent manner. The findings strengthen the involvement of COX2/PGE2 signalling in normal development of the brain and further suggest that abnormal COX2/PGE2 signalling as a result of exposure to environmental factors can result in neuropathologies including those found in ASD

Spinophilin-Dependent Regulation of the Phosphorylation, Protein Interactions, and Function of the GluN2B Subunit of the NMDAR and its Implications in Neuronal Cell Death

Indiana University-Purdue University Indianapolis (IUPUI)Excitotoxicity, a major hallmark of neurodegeneration associated with cerebral ischemia, is a result of accumulation of extracellular glutamate. This excess glutamate leads to hyperactivation of glutamate receptors such as the N-methyl-D-asparate (NMDA) receptors (NMDARs) following the activation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor (AMPARs). Excessive activation of NMDARs causes an influx of calcium, which can eventually activate apoptotic pathways and lead to death of neurons. Regulation of NMDAR subunit composition, localization, surface expression, and activity can balance cell survival via activation of either pro-death or pro-survival pathways after a course of an ischemic insult. Specifically, phosphorylation of different NMDAR subunits defines their activity and downstream signaling pathways. NMDARs are phosphorylated by multiple kinases and dephosphorylated by different phosphatases. Besides phosphatases and kinases, per se, phosphorylation of synaptic proteins that regulate kinase or phosphatase targeting and activity also mediate NMDAR phosphorylation. Spinophilin, a major synaptic scaffolding and protein phosphatase 1 (PP1) targeting protein, mediates substrate phosphorylation via its ability to bind PP1. Our studies focus on delineating the role of spinophilin in the regulation of phosphorylation and function of the GluN2B subunit of the NMDA receptor as well as the role of spinophilin in modulating glutamate-induced neurotoxicity. Interestingly, our data demonstrate that spinophilin sequesters PP1 away from GluN2B thereby enhancing phosphorylation of GluN2B at Ser-1284. These changes impact GluN2B protein interactions, subcellular localization, and surface expression, leading to alterations in the amount of calcium entering the neuron via GluN2B-containing NMDARs. Our data show that spinophilin biphasically regulates GluN2B function. Specifically, Ser-1284 phosphorylation enhances calcium influx through GluN2B containing NMDA receptors, but spinophilin leads to dramatic decreases in the surface expression of the receptor independent of Ser-1284 phosphorylation. Moreover, in spinophilin knockout mice, we observe less PP1 binding to GluN2B and less phosphorylation of Ser-1284, but more surface expression of GluN2B and greater levels of caspase activity. Together, these observations suggest a potential neuroprotective role for spinophilin by decreasing GluN2B-containing NMDA receptor-dependent surface expression and thereby decreasing intracellular calcium and neuronal cell death