N-Methyl-d-aspartate receptors (NMDARs) are ionotropic glutamate receptors that mediate excitatory synaptic transmission and have been implicated in numerous neurological disorders. NMDARs typically comprise two GluN1 and two GluN2 subunits. The four GluN2 subtypes (GluN2A-GluN2D) have distinct functional properties and gene expression patterns, which contribute to diverse functional roles for NMDARs in the brain. Here, we present a series of GluN2C/2D-selective negative allosteric modulators built around a N-aryl benzamide (NAB) core. The prototypical compound, NAB-14, is >800-fold selective for recombinant GluN2C/GluN2D over GluN2A/GluN2B in Xenopus oocytes and has an IC50 value of 580 nM at recombinant GluN2D-containing receptors expressed in mammalian cells. NAB-14 inhibits triheteromeric (GluN1/GluN2A/GluN2C) NMDARs with modestly reduced potency and efficacy compared to diheteromeric (GluN1/GluN2C/GluN2C) receptors. Site-directed mutagenesis suggests that structural determinants for NAB-14 inhibition reside in the GluN2D M1 transmembrane helix. NAB-14 inhibits GluN2D-mediated synaptic currents in rat subthalamic neurons and mouse hippocampal interneurons, but has no effect on synaptic transmission in hippocampal pyramidal neurons, which do not express GluN2C or GluN2D. This series possesses some druglike physical properties and modest brain permeability in rat and mouse. Altogether, this work identifies a new series of negative allosteric modulators that are valuable tools for studying GluN2C- and GluN2D-containing NMDAR function in brain circuits, and suggests that the series has the potential to be developed into therapies for selectively modulating brain circuits involving the GluN2C and GluN2D subunits.
The development of whole exome/genome sequencing technologies has given rise to an unprecedented volume of data linking patient genomic variability to brain disorder phenotypes. A surprising number of variants have been found in the N-methyl-D-aspartate receptor (NMDAR) gene family, with the GRIN2B gene encoding the GluN2B subunit being implicated in many cases of neurodevelopmental disorders, which are psychiatric conditions originating in childhood and include language, motor, and learning disorders, autism spectrum disorder (ASD), attention deficit hyperactivity disorder (ADHD), developmental delay, epilepsy, and schizophrenia. The GRIN2B gene plays a crucial role in normal neuronal development and is important for learning and memory. Mutations in human GRIN2B were distributed throughout the entire gene in a number of patients with various neuropsychiatric and developmental disorders. Studies that provide functional analysis of variants are still lacking, however current analysis of de novo variants that segregate with disease cases such as intellectual disability, developmental delay, ASD or epileptic encephalopathies reveal altered NMDAR function. Here, we summarize the current reports of disease-associated variants in GRIN2B from patients with multiple neurodevelopmental disorders, and discuss implications, highlighting the importance of functional analysis and precision medicine therapies.
We previously demonstrated that mice with reduced expression of the vesicular monoamine transporter 2 (VMAT2 LO) undergo age-related degeneration of the catecholamine-producing neurons of the substantia nigra pars compacta and locus ceruleus and exhibit motor disturbances and depressive-like behavior. In this work, we investigated the effects of reduced vesicular transport on the function and viability of serotonin neurons in these mice. Adult (4-6months of age), VMAT2 LO mice exhibit dramatically reduced (90%) serotonin release capacity, as measured by fast scan cyclic voltammetry. We observed changes in serotonin receptor responsivity in in vivo pharmacological assays. Aged (months) VMAT2 LO mice exhibited abolished 5-HT1A autoreceptor sensitivity, as determined by 8-OH-DPAT (0.1mg/kg) induction of hypothermia. When challenged with the 5HT2 agonist, 2,5-dimethoxy-4-iodoamphetamine (1mg/kg), VMAT2 LO mice exhibited a marked increase (50%) in head twitch responses. We observed sparing of serotonergic terminals in aged mice (18-24months) throughout the forebrain by SERT immunohistochemistry and [3H]-paroxetine binding in striatal homogenates of aged VMAT2 LO mice. In contrast to their loss of catecholamine neurons of the substantia nigra and locus ceruleus, aged VMAT2 LO mice do not exhibit a change in the number of serotonergic (TPH2+) neurons within the dorsal raphe, as measured by unbiased stereology at 26-30 months. Collectively, these data indicate that reduced vesicular monoamine transport significantly disrupts serotonergic signaling, but does not drive degeneration of serotonin neurons.
by
Ana Fernandez-Marmiesse;
Hirofumi Kusumoto;
Saray Rekarte;
Iria Roca;
Jin Zhang;
Scott J. Myers;
Stephen Traynelis;
Ma Luz Couce;
Luis Gutierrez-Solana;
Hongjie Yuan