Molecular basis of positive allosteric modulation of GluN2B NMDA receptors by polyamines

NMDA receptors (NMDARs) form glutamate‐gated ion channels that have central roles in neuronal communication and plasticity throughout the brain. Dysfunctions of NMDARs are involved in several central nervous system disorders, including stroke, chronic pain and schizophrenia. One hallmark of NMDARs is that their activity can be allosterically regulated by a variety of extracellular small ligands. While much has been learned recently regarding allosteric inhibition of NMDARs, the structural determinants underlying positive allosteric modulation of these receptors remain poorly defined. Here, we show that polyamines, naturally occurring polycations that selectively enhance NMDARs containing the GluN2B subunit, bind at a dimer interface between GluN1 and GluN2B subunit N‐terminal domains (NTDs). Polyamines act by shielding negative charges present on GluN1 and GluN2B NTD lower lobes, allowing their close apposition, an effect that in turn prevents NTD clamshell closure. Our work reveals the mechanistic basis for positive allosteric modulation of NMDARs. It provides the first example of an intersubunit binding site in this class of receptors, a discovery that holds promise for future drug interventions.NMDA powder

Allosteric modulation of membrane receptors is widely viewed as a particularly promising strategy in the quest for novel treatments against disorders of the central nervous system (CNS). It relies on the observation that most receptors involved in neurotransmission, be it ionotropic or metabotropic, harbour binding sites for small ligands or ions distinct from the agonist binding sites and the occupancy of which alters receptor activity (orthosteric versus allosteric sites; see Bertrand and Gopalakrishnan, 2007; Niswender and Conn, 2010). The functional outcome of the binding of an allosteric ligand can be either a decrease or increase of the agonist‐evoked response (negative versus positive allosteric modulation) depending on whether the receptors are stabilized in one of the inactive or active states. There are several advantages of targeting allosteric, rather than orthosteric, sites (Bertrand and Gopalakrishnan, 2007; Pin and Prezeau, 2007): first, they do not interfere with the biological patterns of receptor activity; second, allosteric modulators may not compete with a natural ligand as orthosteric ligands with the physiological agonist; third, they allow for subunit‐specific modulation, something which is usually difficult to achieve at orthosteric sites given their high degree of conservation between receptor subtypes. For all these reasons, allosteric modulators represent pharmacological and therapeutic tools of great interest. This is the case in particular for compounds that interact with ionotropic glutamate receptors, a family of glutamate‐gated ion channels that mediate excitatory synaptic transmission in the vertebrate CNS. Molecules capable of enhancing the activity of AMPA or NMDA receptors (NMDARs), the two principal classes of ionotropic glutamate receptors, have potential benefits for the treatment of cognitive deficits caused by neurodegenerative diseases, depression or schizophrenia (Lynch, 2004; Traynelis et al, 2010). However, while the molecular mechanisms of positive allosteric modulation of AMPA receptors have been dissected in much detail (Sun et al, 2002; Lynch, 2004; Jin et al, 2005), at NMDARs these mechanisms have yet to be elucidated.

The first discovered and best‐characterized positive allosteric modulators of NMDARs are polyamines (Rock and Macdonald, 1995; Williams, 1997). Polyamines, such as spermine and spermidine, are polybasic aliphatic amines that are widely distributed throughout the body (Igarashi and Kashiwagi, 2010). They are found at high levels in the intracellular compartment where they regulate several cellular functions. In the CNS, there is also evidence that polyamines can be released into the extracellular medium in an activity‐dependent manner. Once in the extracellular space, polyamines have the potential to modulate neuronal excitability by acting on various ion channels and receptors, including calcium channels and NMDARs (Rock and Macdonald, 1995; Williams, 1997; Mott et al, 2003). Extracellular polyamines have multiple effects on NMDAR responses including a voltage‐dependent pore blockade, an increase in the apparent affinity for the coagonist glycine and a voltage‐independent and glycine‐independent potentiation that proceeds through a reduction of tonic proton inhibition and results in an enhancement of NMDAR responses recorded in saturating concentrations of agonists (McGurk et al, 1990; Lerma, 1992; Rock and MacDonald, 1992; Benveniste and Mayer, 1993; Williams et al, 1994; Traynelis et al, 1995). This latter effect of polyamines (hereafter named ‘polyamine potentiation’ for simplicity) has been most studied because of its unique subunit selectivity. NMDARs form heterotetrameric complexes usually consisting of two GluN1 and two GluN2 subunits, of which there are four subtypes (Paoletti and Neyton, 2007; Traynelis et al, 2010). Only NMDARs containing the GluN2B subunit display polyamine potentiation (Williams et al, 1994; Zhang et al, 1994; Traynelis et al, 1995). Moreover, when the GluN1 subunit contains the exon 5 insert (GluN1‐1b subunit), spermine potentiation is strongly diminished (Zhang et al, 1994; Traynelis et al, 1995).