Extracellular Ca2+ ions reduce NMDA receptor conductance and gating

Maki, Bruce A.; Popescu, Gabriela

doi:10.1085/jgp.201411244

Cited by 27 publications

(30 citation statements)

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“…The findings that Ca 2+ or Cd 2+ binding to the DRPEER motif would favor channel activation (probably more desensitization than opening) may broaden the scope of Ca 2+ -dependent desensitization of the NMDA channel to involve essential conformational changes in the external pore mouth of the NMDA channel, where the activation gate and the glycine-independent desensitization process are very likely also co-localized232453. Together with the findings that the prolonged closed events could always lead into an open event43, and that closed NMDA channels could also desensitize54, it seems that at least a desensitization state is reciprocally interconnected with both the closed and the open state. It may be desirable to explore the conformational changes near the bundle crossing region at the external pore mouth in more detail for further characterization of the molecular nature of NMDA channel opening and desensitization.…”

Section: Discussionmentioning

confidence: 74%

“…In this regard, it is interesting that extracellular Ca 2+ could reduce single NMDA channel conductance, in addition to a decrease in opening probability, which is chiefly ascribable to the increase in prolonged closed events43. If these prolonged closed events, which are in general a few seconds in length, could be viewed as the desensitized state, then these single channel findings would be consistent with the higher affinity of extracellular Cd 2+ toward the desensitized than the open state demonstrated in this study (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…In this regard, an extracellular Ca 2+ binding site, if the occupancy of which could lead to significant gating changes, could then serve to “sense” the extracellular level of Ca 2+ and act as an intramolecular feedback mechanism to regulate Ca 2+ influxes and related physiological consequences. Consistent with this view, a recent work combining single-channel recordings and model simulations suggested a lengthened closed time by extracellularly applied Ca 2+ 43. However, the model does not intend to differentiate the effects of Ca 2+ binding to an extracellular site from a site in the conduction pathway (pore).…”

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confidence: 99%

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Modulation of NMDA channel gating by Ca2+ and Cd2+ binding to the external pore mouth

Yang

Kuo

2016

Sci Rep

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NMDA receptor channels are characterized by high Ca2+ permeability. It remains unclear whether extracellular Ca2+ could directly modulate channel gating and control Ca2+ influxes. We demonstrate a pore-blocking site external to the activation gate for extracellular Ca2+ and Cd2+, which has the same charge and radius as Ca2+ but is impermeable to the channel. The apparent affinity of Cd2+ or Ca2+ is higher toward the activated (a steady-state mixture of the open and desensitized, probably chiefly the latter) than the closed states. The blocking effect of Cd2+ is well correlated with the number of charges in the DRPEER motif at the external pore mouth, with coupling coefficients close to 1 in double mutant cycle analyses. The effect of Ca2+ and especially Cd2+ could be allosterically affected by T647A mutation located just inside the activation gate. A prominent “hook” also develops after wash-off of Cd2+ or Ca2+, suggesting faster unbinding rates of Cd2+ and Ca2+ with the mutation. We conclude that extracellular Ca2+ or Cd2+ directly binds to the DRPEER motif to modify NMDA channel activation (opening as well as desensitization), which seems to involve essential regional conformational changes centered at the bundle crossing point A652 (GluN1)/A651(GluN2).

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Section: Discussionmentioning

confidence: 74%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Modulation of NMDA channel gating by Ca2+ and Cd2+ binding to the external pore mouth

Yang

Kuo

2016

Sci Rep

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“…Specifically, Ca 2þ levels affect NMDA receptor currents through several distinct mechanisms. External Ca 2þ ions bind directly to residues in the outer vestibule of the obligatory GluN1 subunit to reduce channel conductance and gating (1)(2)(3), whereas internal Ca 2þ ions act indirectly through slow and fast mechanisms. Slow, minute-scale processes initiate Ca 2þ -dependent changes in receptor phosphorylation by kinases (e.g., protein kinase A, protein kinase C, and CaMKII) and/or phos-phatases (e.g., calcineurin) to affect channel gating and/or permeation (4)(5)(6)(7), whereas a fast, millisecond-scale process, known as Ca 2þ -dependent inactivation (CDI), which requires calmodulin (CaM) binding to intracellular portions of the GluN1 subunit (8,9), reduces the NMDA receptor current by increasing its macroscopic desensitization (10,11).…”

Section: Introductionmentioning

confidence: 99%

Resident Calmodulin Primes NMDA Receptors for Ca2+-Dependent Inactivation

Iacobucci

Popescu

2017

Biophysical Journal

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N-methyl-d-aspartate (NMDA) receptors are glutamate- and glycine-gated channels that flux Na and Ca into postsynaptic neurons during synaptic transmission. The resulting intracellular Ca transient is essential to physiological and pathological processes related to synaptic development, plasticity, and apoptosis. It also engages calmodulin (CaM) to reduce subsequent NMDA receptor activity in a process known as Ca-dependent inactivation (CDI). Here, we used whole-cell electrophysiology to measure CDI and computational modeling to dissect the sequence of events that underlies it. With these approaches, we estimate that CaM senses NMDA receptor Ca influx at ∼9 nm from the channel pore. Further, when we controlled the frequency of Ca influx through individual channels, we found that a kinetic model where apoCaM associates with channels before their activation best predicts the measured CDI. These results provide, to our knowledge, novel functional evidence for CaM preassociation to NMDA receptors in living cells. This particular mechanism for autoinhibitory feedback reveals strategies and challenges for Ca regulation in neurons during physiological synaptic activity and disease.

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“…For all of the outside-out patch experiments, we used an external solution containing (in mM): 150 NaCl, 10 HEPES, 0.05 EDTA, pH 8.0 (NaOH). Although this external solution does not mimic what occurs at synapses, we used it since it removes the complications of proton (high pH) and divalent (Zn 2+ ) inhibitory effects (Popescu & Auerbach, 2003) and the short- (Maki & Popescu, 2014) and long- (Legendre et al, 1993) term effects of Ca 2+ on NMDARs. We applied 1 s pulses of either 1 mM glutamate or 0.1 mM glycine through a piezo-driven double-barrel application pipette system (10-90% rise time of 400-600 µs).…”

Section: Outside-out Single Channel Recordingsmentioning

confidence: 99%

NMDA receptors require multiple pre-opening gating steps for efficient synaptic activity

Amin

Gochman

et al. 2020

Preprint

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NMDA receptors (NMDAR) are glutamate-gated ion channels that mediate the majority of fast excitatory synaptic transmission in the nervous system. A central feature of NMDAR physiology is the opening of the ion channel driven by presynaptically-released glutamate. Using glutamate applications to outside-out patches containing a single NMDAR in the continuous presence of the co-agonist glycine, we find that agonist-bound receptors transition to the open state via two conformations, an 'unconstrained pre-active' state that can rapidly transition to the open state and contributes to synaptic events, and a 'constrained pre-active' state that requires more energy and hence time to open and does not contribute to fast signaling. To define how agonist binding might drive these conformations, we decoupled the ligand-binding domains from specific transmembrane segments for the GluN1 and GluN2A subunits. Displacements of the central pore-forming M3 segments define the energy of fast channel opening. However, to enter the unconstrained conformation and contribute to fast signaling, a peripheral helix, the GluN2 pre-M1, must be displaced before the M3 segments move. This pre-M1 displacement is facilitated by the flexibility of another nearby peripheral element, the GluN1 and GluN2A S2-M4. We conclude that peripheral structural elements -pre-M1 and S2-M4 -work in concert to remove constraints and prime the channel for rapid opening, thus facilitating fast synaptic transmission. 214 words. McDaniel et al., 2020). The S2-M4 linker/M4 segment also regulates gating (Amin et al., 2017;Yelshanskaya et al., 2017;Shi et al., 2019). Still, the energetics and timing of these non-pore forming elements to receptor gating remains incomplete.Here, we address the gating mechanism in NMDARs. At most synapses, NMDARs are obligate hetero-tetramers composed of two GluN1 and two GluN2 (A-D) subunits. To define conformational changes between agonist binding and pore opening, we rapidly applied glutamate to outside-out patches containing single NMDARs and assessed the time between when glutamate was first applied to when the channel first opened ('latency to 1 st opening', see Materials and Methods) (Aldrich et al., 1983). With this approach, we identified that wild type GluN1/GluN2A NMDARs, in the transition from agonist-bound closed to the open state, transitions through one of two pre-active states: an 'unconstrained' state that can rapidly transition to the open state and contributes to fast synaptic signaling, and a 'constrained' state that slowly transitions to the open state, presumably due to higher energy requirements, and does not contribute to fast signaling. Further, by decoupling the LBD from the TMD for each specific LBD-TMD linker (S1-M1, M3-S2, and S2-M4, Figure 1B & 1C) (Niu et al., 2004;Kazi et al., 2014), we show that the non-pore forming linkers uniquely influence pore-opening by altering the stability of these pre-active conformations. Thus, our results highlight the temporal and coordinated movements from agonist binding to io...

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Extracellular Ca2+ ions reduce NMDA receptor conductance and gating

Abstract: NMDA receptor conductance and gating is directly affected by fluctuations in extracellular calcium concentration.

Cited by 27 publications

References 64 publications

Modulation of NMDA channel gating by Ca2+ and Cd2+ binding to the external pore mouth

Modulation of NMDA channel gating by Ca2+ and Cd2+ binding to the external pore mouth

Resident Calmodulin Primes NMDA Receptors for Ca2+-Dependent Inactivation

NMDA receptors require multiple pre-opening gating steps for efficient synaptic activity

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