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1 The pharmacological properties of CGP 37849 (DL-(E)-2-amino-4-methyl-5-phosphono-3-pentenoic acid; 4-methyl-APPA) and its carboxyethylester, CGP 39551, novel unsaturated analogues of the Nmethyl-D-aspartate (NMDA) receptor antagonist, 2-amino-5-phosphonopentanoate (AP5), were evaluated in rodent brain in vitro and in vivo. 2 Radioligand binding experiments demonstrated that CGP 37849 potently (K1 220 nM) and competitively inhibited NMDA-sensitive L-[3H]-glutamate binding to postsynaptic density (PSD) fractions from rat brain. It inhibited the binding of the selective NMDA receptor antagonist, [3H]-( ±)-342-carboxypiperazin-4-yl)propyl-1-phosphonate (CPP), with a Ki of 35 nm, and was 4, 5 and 7 fold more potent than the antagonists ((±)-cis-4-phosphonomethylpiperidine-2-carboxylic acid) (CGS 19755), CPP and D-AP5, respectively. Inhibitory activity was associated exclusively with the trans configuration of the APPA molecule and with the D-stereoisomer. CGP 39551 showed weaker activity at NMDA receptor recognition sites and both compounds were weak or inactive at 18 other receptor binding sites. 3 CGP 37849 and CGP 39551 were inactive as inhibitors of L-[3H]-glutamate uptake into rat brain synaptosomes and had no effect on the release of endogenous glutamate from rat hippocampal slices evoked by electrical field stimulation. 4 In the hippocampal slice in vitro, CGP 37849 selectively and reversibly antagonized NMDA-evoked increases in CAI pyramidal cell firing rate. In slices bathed in medium containing low Mg2+ levels, concentrations of CGP 37849 up to 10pM suppressed burst firing evoked in CAl neurones by stimulation of Schaffer collateral-commissural fibres without affecting the magnitude of the initial population spike; CGP 39551 exerted the same effect but was weaker. In vivo, oral administration to rats of either CGP 37849 or CGP 39551 selectively blocked firing in hippocampal neurones induced by ionophoreticallyapplied NMDA, without affecting the responses to quisqualate or kainate. al., 1988). Many studies have demonstrated that activation of the NMDA receptor is involved in the generation of epileptiform activity and in hypoxic-ischaemic neuronal damage, and conversely that NMDA receptor antagonists are anticonvulsant and cerebroprotective in animal models of epilepsy and stroke (see reviews by Meldrum, 1985;Rothman & Olney, 1986;Choi, 1988;Patel et al., 1988;Iversen et al., 1989;Albers et al., 1989 (Meldrum, 1985;Rothman & Olney, 1986;Albers et al., 1989;.Antagonism of NMDA receptor mechanisms potentially may be achieved by a number of different approaches. In addition to the transmitter recognition site, the NMDA receptor complex comprises an allosteric regulatory site and a channel binding domain (Foster & Fagg, 1987b), and possibly also sites defined by the actions of Zn2", polyamines, tricyclic antidepressants, ifenprodil and CGP 31358 (see Lodge, 1989;Baud et al., 1989). At present, however, the two most well characterized sites are (1) the transmitter recognition site, at which substances such as ...

The undecapeptide substance P is a putative neurotransmitter in the mammalian central nervous system (CNS), and may be associated with pain fibres in the spinal cord. Radiolabelled derivatives of other neuropeptides have been used to demonstrate specific interactions with receptor sites on brain membranes, and this approach has now been explored with substance P. We have now prepared [4-3H-Phe8]-substance P and we find that it binds reversibly to a saturable population of sites in rat brain particulate fractions. Scatchard analysis of concentration-dependent saturation of binding indicates a single population of non-interacting sites with a high affinity (Kd=0.38 nM) and a low density (Bmax=27.2 fmol per mg protein). Kinetic analyses indicate an apparent dissociation equilibrium constant of 0.46 nM. A variety of neurotransmitter amines and amino acids, and other peptides do not compete at the substance P sites, but structurally related peptides or shorter C-terminal fragments of substance P are active. The rank order of potency of these substance P-related peptides agrees with that reported for their effects in depolarizing spinal cord neurones. The regional distribution of the specific binding sites for 3H-substance P parallels that of substance P immunoreactivity, being high in the hypothalamus and low in the cerebellum and cerebral cortex. The characteristics of the 3H-substance P binding sites are consistent with those expected for substance P receptors.

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