Glycine-insensitive desensitization of NMDA responses in cultured mouse embryonic neurons (original) (raw)
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Glycine-dependent activation of NMDA receptors
The Journal of general physiology, 2015
N-methyl-d-aspartate (NMDA) receptors are the only neurotransmitter receptors whose activation requires two distinct agonists. Heterotetramers of two GluN1 and two GluN2 subunits, NMDA receptors are broadly distributed in the central nervous system, where they mediate excitatory currents in response to synaptic glutamate release. Pore opening depends on the concurrent presence of glycine, which modulates the amplitude and time course of the glutamate-elicited response. Gating schemes for fully glutamate- and glycine-bound NMDA receptors have been described in sufficient detail to bridge the gap between microscopic and macroscopic receptor behaviors; for several receptor isoforms, these schemes include glutamate-binding steps. We examined currents recorded from cell-attached patches containing one GluN1/GluN2A receptor in the presence of several glycine-site agonists and used kinetic modeling of these data to develop reaction schemes that include explicit glycine-binding steps. Based...
Interactions between the glycine and glutamate binding sites of the NMDA receptor
The Journal of neuroscience : the official journal of the Society for Neuroscience, 1993
The interactions between the glycine and glutamate binding sites of the NMDA receptor have been studied in outside-out patches and synapses from hippocampal neurons in culture using rapid drug application techniques. Desensitization of NMDA receptor-mediated currents elicited by glutamate in newly excised outside-out patches was reduced in the presence of saturating concentrations of glycine. This suggests that the glutamate and glycine binding sites of the NMDA receptor are allosterically coupled as has been reported in whole-cell preparations. A glycine-insensitive form of desensitization increased rapidly over the first few minutes of recording and largely occluded the glycine concentration-sensitive desensitization in outside-out patches. However, even in old patches that displayed no glycine-sensitive desensitization, the unbinding rate of glycine was increased fourfold by the presence of glutamate, suggesting that the two binding sites were still allosterically coupled. These ...
Glycine uptake governs glycine site occupancy at NMDA receptors of excitatory synapses
Journal of neurophysiology, 1998
Glycine uptake governs glycine site occupancy at NMDA receptors of excitatory synapses. J. Neurophysiol. 80: 3336-3340, 1998. At central synapses occupation of glycine binding sites of N-methyl--aspartate receptors (NMDA-Rs) is a necessary prerequisite for the excitatory neurotransmitter glutamate to activate these receptors. There is conflicting evidence as to whether glycine binding sites normally are saturated. If they are not, then alterations in local glycine concentration could modulate excitatory synaptic transmission. By using an in vitro brain stem slice preparation we investigated whether the glycine site is saturated for synaptically activated NMDA-Rs in neonatal rat hypoglossal motoneurons. We found that the NMDA-R-mediated component of spontaneous miniature excitatory postsynaptic currents could be potentiated by exogenously applied glycine as well as by -serine. The effects of glycine were observed only at concentrations (100 microM or more) two orders of magnitude abo...
Glycine Binding Site of the Synaptic NMDA Receptor in Subpostremal NTS Neurons
Journal of Neurophysiology, 2005
The nucleus of the tractus solitarius (NTS) plays an important role in the control of several autonomic reflex functions and has glutamate and GABA as main neurotransmitters. In this work, we used patch-clamp recordings in transverse slice preparations from rats to study whether the glycine binding site of the N-methyl-d-aspartate (NMDA) receptor is saturated or not in neurons of the subpostremal NTS. Except at hyperpolarized voltages and close to the reversal potential, glycine potentiated the NMDA responses in a concentration-dependent manner. The total charge transferred by glutamatergic currents was enhanced by glycine (500 μM; from 28 ± 13 to 42 ± 18 pC at +50 mV, n = 7, P < 0.05). Glycine increased the conductance of the postsynaptic membrane, without altering its reversal potential, both in the presence (from 2.4 ± 0.06 to 3.4 ± 0.09 nS; n = 7) and absence (from 3.1 ± 0.06 to 4.4 ± 0.10 nS; n = 8) of Mg2+ in the bathing solution. d-serine, in the presence of strychnine, al...
Activation of the glycine site in the NMDA receptor is necessary for the induction of LTP
Neuroscience Letters, 1990
The effects of 7-chlorokynurenate (7-C1KY) were examined on N-methyl-D-aspartate (NMDA) receptor-mediated synaptic mechanisms in the CAI region of rat hippocampal slices. 7-CIKY depressed both the NMDA receptor-mediated component of synaptic transmission recorded in Mg2÷-free medium and the induction of long-term potentiation (LTP) in Mg2+-containing medium. Both of these effects were reversed by D-serine, suggesting that the action of 7-CIKY was at the allosteric glycine site on the NMDA receptor.
Glycine site associated with the NMDA receptor modulates long-term potentiation
Synapse, 1990
Recent work has shown that kynurenic acid and several quinoxaline derivatives act as non-competitive NMDA receptor antagonists by binding to the glycine site associated with this receptor. In this study, we have tested the effect of the most potent and selective of these compounds, 7-chlorokynurenic acid (C1-Kyn), on the induction of long-term potentiation, an event know to involve activation of NMDA receptors. It was found that 30 m̈M C1-Kyn reversibly abolished the development of both short-term and long-term potentiation in the CA1 region of hippocampal slices. The effectiveness of C1-Kyn matched its ability to inhibit 3H-glycine binding and the association of 3H-TCP with the NDMA receptor in binding experiments (Ki 0.7–1 μM). Weak interactions of C1-Kyn with AMPA receptor sites were observed and may account for a partial, reversible reduction in the epsp. However, blockade of long-term potentiation by C1-Kyn was completely reversed by simultaneous application of the glycine site agonist D-serine and thus must be attributed to its interaction with the glycine site. These results indicate that the glycine site coupled to the NMDA receptor potently modulates channel function during physiological events related to synaptic activation.
The glycine coagonist site of the NMDA receptor
Advances in experimental medicine and biology, 1990
The discovery of the glycine binding site on the NMDA receptor (Johnson and Ascher, 1987) is recognized as a highly significa;1t event by those interested in receptor-channel mechanisms. It also offers a new target site in drug development for the treatment of neuropa-thologies associated with NMDA receptor activation. The list of neural dysfunctions mediated in part by synaptic activation of NMDA receptors very likely includes epileptiform seizures (Croucher et ai., 1982) and brain damage induced by ischemia or hypoxia in some models (Simon et ai., 1984). Growing evidence indicates however that NMDA receptor activation is important also for establishing certain neuronal connections during development (Tsumoto et ai., 1987; Kleinschmidt et ai., 1987; Lincoln et ai., 1988) and has been implicated in some types of learning or memory (Morris et ai., 1986). Thus both agonists and antagonists of the NMDA receptor may find clinical utility in the future. Currently available NMDA receptor blockers, acting either at the glutamate binding site or the open ion channel, either have difficulty penetrating the blood-brain barrier or have unacceptable side effects such as psychosis. The glycine site appears to present a pharmacology sufficiently different from that of the other binding sites on the NMDA receptor to offer a novel target for new drug discovery. The preparation we hilve used is the Xenopus oocyte injected with rat brain mRNA. It is a large (1 mm diameter), hardy cell that ciln be cultured easily for a week or more after injection, during which time transliltion of the foreign mRNA, ilssembly of multisubunit receptors and right-side-out insertion into the plasma membrane occurs. Receptor properties arc then studied under well-defined voltage cl<Jmp conditions. The reproducibility of this preparation offers a reliable bioassilY for the quantitiltive study of receptor-coupled ion channel properties. We have shown previously that NMDA and kainilte/quisqualate receptors with properties indistinguishable from those in neurons are expressed by oocytes (
The Journal of physiology, 1992
1. The characteristics of the activation of the N-methyl-D-aspartate (NMDA) response by glycine were studied using whole-cell and outside-out patch clamp recording techniques. 2. Glycine concentration-response (C-R) curves were measured in the presence of 10 microM-NMDA and fitted with the Hill equation modified to account for the response to NMDA observed in the absence of added glycine. The mean value of the apparent dissociation constant (KD) was 150 nM, and the mean value of the Hill coefficient (nH) was 1.1. When the KD was corrected for the concentration of contaminating glycine in nominally glycine-free solutions, estimated assuming that there is no response in the absence of glycine, the value was 130 nM. 3. The question of how many glycine binding sites there are on each NMDA receptor-channel complex was addressed by examining the curvature at the foot of the glycine C-R curve. An equation that allowed estimation of both the concentration of contaminating glycine and of the...
receptorsreaction mechanism of neuronal NMDA
2015
The activation mechanisms of recombinant N-methyl-D-aspartate receptors (NRs) have been established in sufficient detail to account for their single channel and macroscopic responses; however, the reaction mechanism of native NRs remains uncertain due to indetermination of the isoforms expressed and possible neuron-specific factors. To delineate the activation mechanism of native NRs, we examined the kinetic properties of currents generated by individual channels located at the soma of cultured rat neurons. Cells were dissociated from the embryonic cerebral cortex or hippocampus, and on-cell single channel recordings were done between 4 and 50 days in vitro (DIV). We observed two types of kinetics that correlated with the age of the culture. When we segregated recordings by culture age, we found that receptors recorded from early (4-33 DIV) and late (25-50 DIV) cultures had smaller unitary conductances but had kinetic profiles that matched closely those of recombinant 2B-or 2A-containing receptors, respectively. In addition, we examined the effects of cotransfection with postsynaptic density protein 95 or neuropilin tolloid-like protein 1 on recombinant receptors expressed in human embryonic kidney-293 cells. Our results add support to the view that neuronal cultures recapitulate the developmental patterns of receptor expression observed in the intact animal and demonstrate that the activation mechanism of somatic neuronal NRs is similar to that described for recombinant receptors of defined subunit composition.