Blunted type-5 metabotropic glutamate receptor-mediated polyphosphoinositide hydrolysis in two mouse models of monogenic autism.

The involvement of the mGlu5 receptors in the pathophysiology of several forms of monogenic autism has been supported by numerous studies following the seminal observation that mGlu5 receptor-dependent long-term depression was enhanced in the hippocampus of mice modeling the fragile-X syndrome (FXS). Surprisingly, there are no studies examining the canonical signal transduction pathway activated by mGlu5 receptors (i.e. polyphosphoinositide - PI - hydrolysis) in mouse models of autism. We have developed a method for in vivo assessment of PI hydrolysis based on systemic injection of lithium chloride followed by treatment with the selective mGlu5 receptor PAM, VU0360172, and measurement of endogenous inositolmonophosphate (InsP) in brain tissue. Here, we report that mGlu5 receptor-mediated PI hydrolysis was blunted in the cerebral cortex, hippocampus, and corpus striatum of Ube3am-/p+ mice modeling Angelman syndrome (AS), and in the cerebral cortex and hippocampus of Fmr1 knockout mice modeling FXS. In vivo mGlu5 receptor-mediated stimulation of Akt on threonine 308 was also blunted in the hippocampus of FXS mice. These changes were associated with a significant increase in cortical and striatal Homer1 levels and striatal mGlu5 receptor and Gαq levels in AS mice, and with a reduction in cortical mGlu5 receptor and hippocampal Gαq levels, and an increase in cortical phospholipase-Cß and hippocampal Homer1 levels in FXS mice. This is the first evidence that the canonical transduction pathway activated by mGlu5 receptors is down-regulated in brain regions of mice modeling monogenic autism.

Aberrant mitochondrial bioenergetics in the cerebral cortex of the Fmr1 knockout mouse model of fragile X syndrome.

Impaired energy metabolism may play a role in the pathogenesis of neurodevelopmental disorders including fragile X syndrome (FXS). We checked brain energy status and some aspects of cell bioenergetics, namely the activity of key glycolytic enzymes, glycerol-3-phosphate shuttle and mitochondrial respiratory chain (MRC) complexes, in the cerebral cortex of the Fmr1 knockout (KO) mouse model of FXS. We found that, despite a hyperactivation of MRC complexes, adenosine triphosphate (ATP) production via mitochondrial oxidative phosphorylation (OXPHOS) is compromised, resulting in brain energy impairment in juvenile and late-adult Fmr1 KO mice. Thus, an altered mitochondrial energy metabolism may contribute to neurological impairment in FXS.

Activation of Serotonin 5-HT7 Receptors Modulates Hippocampal Synaptic Plasticity by Stimulation of Adenylate Cyclases and Rescues Learning and Behavior in a Mouse Model of Fragile X Syndrome.

We have previously demonstrated that activation of serotonin 5-HT7 receptors (5-HT7R) reverses metabotropic glutamate receptor-mediated long term depression (mGluR-LTD) in the hippocampus of wild-type (WT) and Fmr1 Knockout (KO) mice, a model of Fragile X Syndrome (FXS) in which mGluR-LTD is abnormally enhanced. Here, we have investigated intracellular mechanisms underlying the effect of 5-HT7R activation using patch clamp on hippocampal slices. Furthermore, we have tested whether in vivo administration of LP-211, a selective 5-HT7R agonist, can rescue learning and behavior in Fmr1 KO mice. In the presence of an adenylate cyclase blocker, mGluR-LTD was slightly enhanced in WT and therefore the difference between mGluR-LTD in WT and Fmr1 KO slices was no longer present. Conversely, activation of adenylate cyclase by either forskolin or Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) completely reversed mGluR-LTD in WT and Fmr1 KO. 5-HT7R activation reversed mGluR-LTD in WT and corrected exaggerated mGluR-LTD in Fmr1 KO; this effect was abolished by blockade of either adenylate cyclase or protein kinase A (PKA). Exposure of hippocampal slices to LP-211 caused an increased phosphorylation of extracellular signal regulated kinase (ERK), an intracellular effector involved in mGluR-LTD, in WT mice. Conversely, this effect was barely detectable in Fmr1 KO mice, suggesting that 5-HT7R-mediated reversal of mGluR-LTD does not require ERK stimulation. Finally, an acute in vivo administration of LP-211 improved novel object recognition (NOR) performance in WT and Fmr1 KO mice and reduced stereotyped behavior in Fmr1 KO mice. Our results indicate that mGluR-LTD in WT and Fmr1 KO slices is bidirectionally modulated in conditions of either reduced or enhanced cAMP formation. Activation of 5-HT7 receptors reverses mGluR-LTD by activation of the cAMP/PKA intracellular pathway. Importantly, a systemic administration of a 5-HT7R agonist to Fmr1 KO mice corrected learning deficits and repetitive behavior. We suggest that selective 5-HT7R agonists might become novel pharmacological tools for FXS therapy.

Viability of olfactory ensheathing cells after hypoxia and serum deprivation: Implication for therapeutic transplantation.

Olfactory ensheathing cells (OECs) represent glial cells supporting neuronal turnover in the olfactory system. In vitro, OECs promote axonal growth as a source of neurotrophic growth factors; in vivo, they produce myelin, promoting remyelination of damaged axons. Consequently, OEC transplantation appears to be a promising treatment for spinal cord injury, although the functional recovery is limited. This might be ascribed to the microenvironment at the lesion site, lacking growth factors (GFs), nutrients, and oxygen. To mimic this condition, we used an in vitro approach by growing primary neonatal mouse OECs under hypoxic conditions and/or serum deprivation. In addition, we compared OECs survival/proliferation with that of primary cultures of Schwann cells (SCs) and astrocytes under the same experimental conditions. Cultures were analyzed by immunocytochemistry, and cell viability was evaluated by MTT assay. Different GFs, such as NGF, bFGF, and GDNF, and their combination were used to rescue cells from serum and/or oxygen deprivation. We show that the cell types were differently sensitive to the tested stress conditions and that OECs were the most sensitive among them. Moreover, OEC viability was rescued by bFGF under serum-deprived or hypoxic condition but not under conditions of drastic serum deprivation and hypoxia. bFGF was effective also for the other cell types, whereas the effect of the other GFs was negligible. This model suggests that administration of bFGF might be considered useful to sustain cell survival/proliferation after transplantation of OECs either alone or in combination with other glial cell types.

Metabotropic glutamate receptors in glial cells.

Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system (CNS) and exerts its actions via a number of ionotropic glutamate receptors/channels and metabotropic glutamate (mGlu) receptors. In addition to being expressed in neurons, glutamate receptors are expressed in different types of glial cells including astrocytes, oligodendrocytes, and microglia. Astrocytes are now recognized as dynamic signaling elements actively integrating neuronal inputs. Synaptic activity can evoke calcium signals in astrocytes, resulting in the release of gliotransmitters, such as glutamate, ATP, and D-serine, which in turn modulate neuronal excitability and synaptic transmission. In addition, astrocytes, and microglia may play an important role in pathology such as brain trauma and neurodegeneration, limiting or amplifying the pathologic process leading to neuronal death. The present review will focus on recent advances on the role of mGlu receptors expressed in glial cells under physiologic and pathologic conditions.

Group I metabotropic glutamate receptors: a role in neurodevelopmental disorders?

Group I metabotropic glutamate receptors (mGlu1 and mGlu5) are coupled to polyphosphoinositide hydrolysis and are involved in activity-dependent forms of synaptic plasticity, both during development and in the adult life. Group I mGlu receptors can also regulate proliferation, differentiation, and survival of neural stem/progenitor cells, which further support their role in brain development. An exaggerated response to activation of mGlu5 receptors may underlie synaptic dysfunction in Fragile X syndrome, the most common inherited form of mental retardation. In addition, group I mGlu receptors are overexpressed in dysplastic neurons of focal cortical dysplasia and hemimegaloencephaly, which are disorders of cortical development associated with chronic epilepsy. Drugs that block the activity of group I mGlu receptors (in particular, mGlu5 receptors) are potentially helpful for the treatment of Fragile X syndrome and perhaps other neurodevelopmental disorders.

A reduced number of metabotropic glutamate subtype 5 receptors are associated with constitutive homer proteins in a mouse model of fragile X syndrome.

Fragile X (FRAX) syndrome is a common inherited form of mental retardation resulting from the lack of fragile X mental retardation protein (FMRP) expression. The consequences of FMRP absence in the mechanism underlying mental retardation are unknown. Here, we tested the hypothesis that glutamate receptor (GluR) expression might be altered in FRAX syndrome. Initial in situ hybridization and Western blotting experiments did not reveal differences in mRNA levels and protein expression of AMPA and NMDA subunits and metabotropic glutamate subtype 5 (mGlu5) receptors between control and Fmr1 knock-out (KO) mice during postnatal development. However, a detergent treatment (1% Triton X-100) revealed a selective reduction of mGlu5 receptor expression in the detergent-insoluble fraction of synaptic plasma membranes (SPMs) from KO mice, with no difference in the expression of NR2A, GluR1, GluR2/3, GluR4, and Homer proteins. mGlu5 receptor expression was also lower in Homer immunoprecipitates from Fmr1 KO SPMs. Homer, but not NR2A, mGlu5, and GluR1, was found to be less tyrosine phosphorylated in Fmr1 KO than control mice. Our data indicate that, in FRAX syndrome, a reduced number of mGlu5 receptors are tightly linked to the constituents of postsynaptic density and, in particular, to the constitutive forms of Homer proteins, with possible consequent alterations in synaptic plasticity.