Different oxysterols have opposing actions at N-methyl-d-aspartate receptors

Abstract

Oxysterols have emerged as important biomarkers in disease and as signaling molecules. We recently showed that the oxysterol 24(S)-hydroxycholesterol, the major brain cholesterol metabolite, potently and selectively enhances NMDA receptor function at a site distinct from other modulators. Here we further characterize the pharmacological mechanisms of 24(S)-hydroxycholesterol and its synthetic analogue SGE201. We describe an oxysterol antagonist of this positive allosteric modulation, 25-hydroxycholesterol. We found that 24(S)-hydroxycholesterol and SGE201 primarily increased the efficacy of NMDAR agonists but did not directly gate the channel or increase functional receptor number. Rather than binding to a direct aqueous-accessible site, oxysterols may partition into the plasma membrane to access the NMDAR, likely explaining slow onset and offset kinetics of modulation. Interestingly, oxysterols were ineffective when applied to the cytosolic face of inside-out membrane patches or through a whole-cell pipette solution, suggesting a non-intracellular site. We also found that another natural oxysterol, 25-hydroxycholesterol, although exhibiting slight potentiation on its own, non-competitively and enantioselectively antagonized the effects of 24(S)-hydroxycholesterol analogues. In summary, we suggest two novel allosteric sites on NMDARs that separately modulate channel gating, but together oppose each other.

24(S)-Hydroxycholesterol as a Modulator of Neuronal Signaling and Survival

“…There are some peculiar aspects to 24S-HC modulation of NMDAR function that may yield insights into mechanisms (Linsenbardt and others 2014; Paul and others 2013). Although some positive and negative allosteric modulators exhibit selective effects on receptors containing certain GluN2 subunits (Hedegaard and others 2012), we failed to find any dependence of oxysterol potentiation on subunit composition.…”

LTP-triggered cholesterol redistribution activates Cdc42 and drives AMPA receptor synaptic delivery

“…Therefore, we expected that LTP and synaptic potentiation induced by cholesterol removal may occlude each other. On the other hand, it has also been shown that oxysterols may directly modulate NMDR function (Linsenbardt et al, 2014). We then tested whether cholesterol removal was affecting NMDAR-mediated synaptic responses and, in this manner, may alter LTP induction.…”

“…Thus, AMPAR synaptic potentiation required both Rab11 and Cdc42 activity, suggesting that besides mobilization of recycling endosomes, this process engages additional signaling This result supports the interpretation that cholesterol removal is a necessary step for LTP expression. However, NMDAR function itself may be regulated by cholesterol, as it has been shown for oxysterols (Linsenbardt et al, 2014). To test this possibility, we acutely added MBCD-cholesterol while monitoring NMDAR synaptic responses.…”

Mechanism and properties of positive allosteric modulation of N -methyl- d -aspartate receptors by 6-alkyl 2-naphthoic acid derivatives

“…Elevated agonist concentrations can increase a modulator’s potency and conversely a PAM can increase agonist potency, as seen for SGE201 (Linsenbardt et al, 2014) and GNE-8324 (Hackos et al, 2016). In such cases, saturating agonist concentrations may mask potentiating activity if the primary PAM action is to increase agonist occupation.…”

“…The alkyl-naphthoic acid PAMs characterized here add to the pharmacodynamic diversity of the rapidly expanding list of NMDAR PAMs such as PS (Chopra et al, 2015; Horak et al, 2006; Horak et al, 2004; Jang et al, 2004; Kostakis et al, 2011; Wu et al, 1991), UBP512, UBP646 (Costa et al, 2010), UBP714 (Irvine et al, 2012), CIQ (Mullasseril et al, 2010), PYD106 (Khatri et al, 2014), SGE201 (Linsenbardt et al, 2014; Paul et al, 2013), and GNE6901 (Hackos et al, 2016). These agents differ in their subtype-selectivity, N-terminal insert-sensitivity, pH-sensitivity, use/disuse-dependency, and their effects on agonist potency, efficacy and deactivation.…”