NMDA Receptor Affinity Research Articles

ChemInform Abstract: Stereoselective Synthesis of 2‐Aminoethyl Substituted Tricycles with NMDA Receptor Affinity.

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Stereoselective synthesis of 2-aminoethyl substituted tricycles with NMDA receptor affinity

In this paper the stereoselective synthesis of the novel noncompetitive NMDA antagonist 15 with a k i value of 16.1 μM for the phencyclidine binding site is described. Key steps in the preparation of the tricycle 15 are the regio- and stereoselective formation of the acetal 7 , an intramolecular Heck reaction ( 11 → 12 ) and the stereoselective hydrogenation of the double bond of 12 to give 13 . The stereoselective synthesis of the novel NMDA antagonist 2 and its affinity for the phencyclidine binding site (k i = 16.1 μM) are described.

Modulation of glycine affinity for NMDA receptors by extracellular Ca2+ in trigeminal neurons

Glycine and the divalent cation Ca2+ play key roles in regulating the activity of excitatory amino acid NMDA receptor channels. There is accumulating evidence that the concentration of glycine at the synaptic cleft is below a saturated level. We examined the effect of external Ca2+ on NMDA responses in various concentrations of glycine in isolated trigeminal neurons. We found that external Ca2+ potentiated NMDA responses and this potentiation occurred only when glycine sites were unsaturated. Since single-channel conductance decreases in the high external Ca2+ solution, the observation cannot be explained by an increase in Ca2+ influx through the channels. Studying the dose-response curves for glycine in different Ca2+ solutions, we found that the apparent dissociation constant (EC50) for glycine decreases with increasing external Ca2+ concentrations. Kinetics studies of glycine binding to NMDA receptors indicated that external Ca2+ causes a decrease in the off rate of the glycine binding, while having no effect on the on rate. Our analyses suggest that the apparent glycine affinity increases by about 3.7 times in Ca-containing solution. Thus, external Ca2+ contributes to the unusually high glycine affinity for NMDA receptors and may have a role in regulating the NMDA receptor channel activities during intensive or sustained neuronal stimulation.

Protective effect of MgSO 4 infusion on NMDA receptor binding characteristics during cerebral cortical hypoxia in the newborn piglet

This study tests the hypothesis that magnesium, a selective non-competitive antagonist of the NMDA receptor, will attenuate hypoxia-induced alteration in NMDA receptors and preserve MK-801 binding characteristics during cerebral hypoxia in vivo. Anesthetized, ventilated and instrumented newborn piglets were divided into three groups: normoxic controls were compared to untreated hypoxic and Mg 2+-treated hypoxic piglets. Cerebral hypoxia was induced by lowering the FiO 2 to 5–7% and confirmed biochemically by a decrease in the levels of phosphocreatine (82% lower than control). The Mg 2+-treated group received MgSO 4 600 mg/kg over 30 min followed by 300 mg/kg administered during 60 min of hypoxia. Plasma Mg 2+ concentrations increased from1.6 ± 0.1mg/dl to17.7 ± 3.3mg/dl. 3H-MK-801 binding was used as an index of NMDA receptor modification. The B max in control, hypoxic and Mg 2+-treated hypoxic piglets was1.09 ± 0.17, 0.70 ± 0.25and0.96 ± 0.14pmoles/mg protein, respectively. The K d for the same groups were10.02 ± 2.04, 4.88 ± 1.43and8.71 ± 2.23nM, respectively. The B max and K d in the hypoxic group were significantly lower compared to the control and Mg 2+-treated hypoxic groups, indicating a preservation of NMDA receptor number and affinity for MK-801 during hypoxia with Mg 2+. The activity of Na +, K + ATPase, a marker of neuronal membrane function, was lower in the hypoxic group compared to the control and Mg 2+-treated hypoxic groups. These findings show that MgSO 4 prevents the hypoxia-induced modification of the NMDA receptor and attenuates neuronal membrane dysfunction. We suggest that the administration of Mg 2+ prior to and during hypoxia may be neuroprotective in vivo, possibly by reducing the NMDA receptor-mediated influx of calcium.

Aspartate and glutamate mediate excitatory synaptic transmission in area CA1 of the hippocampus

We examined whether L-aspartate (ASP) and L-glutamate (GLU) both function as endogenous neurotransmitters in area CA1 of the rat hippocampus. Radioligand displacement experiments using 3H-DL-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (3H-AMPA) to label AMPA/kainate receptors and 3H-cis-4-phosphonomethyl-2-piperidine carboxylic acid (3H-CGS-19755) to label NMDA receptors confirmed that GLU (Ki approximately 500 nM) but not ASP (Ki > 1 mM) has high affinity for AMPA/kainate receptors whereas GLU (Ki approximately 250 nM) and ASP (Ki approximately 1.3 microM) both have high affinity for NMDA receptors. Elevating extracellular potassium concentration (50 mM, 1 min) evoked the calcium-dependent release of both ASP (approximately 50% increase) and GLU (approximately 200% increase) from hippocampal slices and from minislices of area CA1. Reducing extracellular glucose concentration (0.2 mM) reduced GLU release, enhanced ASP release, and reduced AMPA/kainate receptor-mediated responses more than NMDA receptor-mediated responses (to 7% and 34% of control, respectively). Fiber volleys, antidromic population spikes, membrane potential, input resistance, and ATP content all were not affected by glucose reduction. Unlike low glucose, the inhibitory neuromodulator adenosine (5 microM), which reduces ASP and GLU release to a similar extent, reduced AMPA/kainate and NMDA receptor-mediated population EPSPs similarly (to 11% and 12% of control, respectively). AMPA/kainate and NMDA receptor-mediated population EPSPs were also similarly reduced by 0.4 microM TTX (to 32% and 22% of control, respectively) and similarly enhanced by 10 microM 4-aminopyridine (to 206% and 248% of control, respectively). Finally, NMDA receptor-mediated EPSCs measured by whole-cell recording decayed faster in low glucose (73 msec vs 54 msec) but not in adenosine (73 msec vs 78 msec). Together, these results confirm that ASP and GLU are both involved in excitatory synaptic transmission at the Schaffer collateral-commissural terminals in area CA1 of the rat hippocampus.

Potent quinoxaline-spaced phosphono alpha-amino acids of the AP-6 type as competitive NMDA antagonists: synthesis and biological evaluation.

A series of alpha-amino-3-(phosphonoalkyl)-2-quinoxalinepropanoic acids was synthesized and evaluated for NMDA receptor affinity using a [3H] CPP binding assay. Functional antagonism of the NMDA receptor complex was evaluated in vitro using a stimulated [3H]TCP binding assay and in vivo by employing an NMDA-induced seizure model. Some analogues also were evaluated in the [3H]-glycine binding assay. Several compounds of the AP-6 type show potent and selective NMDA antagonistic activity both in vitro and in vivo. In particular alpha-amino-7-chloro-3-(phosphonomethyl)-2-quinoxalinepropanoic acid (1) displayed an ED50 of 1.1 mg/kg ip in the NMDA lethality model. Noteworthy is alpha-amino-6,7-dichloro-3-(phosphonomethyl)-2-quinoxalinepropanoic++ + acid (3) with a unique dual activity, displaying in the NMDA receptor binding assay an IC50 of 3.4 nM and in the glycine binding assay an IC50 of 0.61 microM.

Heterocyclic excitatory amino acids. Synthesis and biological activity of novel analogues of AMPA.

The novel acidic amino acids 6a-c, 7, and 8 have been synthesized via 1,3-dipolar cycloadditions, using nitrile oxides and alkynes. The prepared compounds are heterocyclic analogues of glutamic acid with differing chain lengths. One of these compounds, (RS)-2-amino-3-(3-carboxy-5-methyl-4- isoxazolyl)propionic acid (ACPA, 8), was shown in [3H]AMPA binding studies to be more active than AMPA itself (IC50 = 20 nM compared to IC50 = 79 nM for AMPA). No affinity for NMDA receptors (NMDA-sensitive [3H]glutamic acid binding) was found, and only weak affinity in [3H]kainic acid binding (IC50 = 6.3 microM) was detected. The excitatory activity in rat cortical wedge also showed that ACPA was more potent than AMPA (EC50 = 1.0 microM compared to EC50 = 3.5 microM for AMPA). The depolarizing effect of ACPA could be fully antagonized by the selective non-NMDA antagonist 6-cyano-7-nitro-quinoxazoline-2,3-dione (CNQX), but was unaffected by the selective NMDA antagonist D-2-amino-5-phosphonovaleric acid (AP5).