The effect of two novel dipeptide antagonists of excitatory amino acid neurotransmission on the high pressure neurological syndrome in the rat

Intracerebroventricular injection in the rat of beta-D-aspartyl aminomethylphosphonate (Asp-Amp) 1 mumol, or Y-D-glutamylaminomethylsulphonate (GAMS) 1 mumol, increases the onset pressure for the initial tremor phase of the high pressure neurological syndrome (HPNS) by 50%. Asp-Amp also significantly increases the onset pressures for myoclonus and for tonic-clonic seizures. GAMS did not significantly change the onset pressures for myoclonus or tonic clonic seizures, but it caused the appearance of brief clonic seizures prior to the onset of the HPNS.     

Effects of excitatory amino acid antagonists on social recognition of male rats

The effects of NMDA and non-NMDA glutamate receptor antagonists were evaluated on social recognition of adult male rats. Adult animals were injected (s.c.) with drug or saline immediately after the initial exposure to a juvenile male, 21-24 days old, and re-exposed to the same or to a novel juvenile 30min later. If the time spent by animals in social investigation during re-exposure to the same juvenile was comparable with the time measured during the initial exposure and with the time of animals re-exposed to a novel juvenile, the effect of a drug was designated as amnesia. Such an effect was found in adult animals administered 1 and 1.5mg/kg phencyclidine, 0.1mg/kg dizoclipine, 2.5mg/kg CPP, and 4mg/kg CGS 19755. Amnesic effects were observed at doses not affecting motor performance. No amnesic effect was produced by CNQX and NBQX (2.5, 5 and 10mg/kg). These results show that while recognition capacity in adult male rats is disrupted by several NMDA antagonists, non-NMDA antagonists do not interfere with short-term retention of individual odours. This suggests that NMDA glutamate receptors may be involved in the processing of socially relevant olfactory information.     

Noncompetitive excitatory amino acid receptor antagonists

In the first article in this series, Watkins, Krogsgaard-Larsen and Honoré outlined the structure-activity requirements at the receptor sites for excitatory amino acids in the mammalian CNS. The postsynaptic depolarizing actions of glutamate are thought to be mediated by NMDA, AMPA and kainate receptors. Here David Lodge and Kenneth M. Johnson review some of the recent developments in the pharmacology of other means by which the function of these receptors may be modulated. Divalent cations, phencyclidine-like drugs, glycine analogues and polyamines all modulate NMDA receptors whereas barbiturates and some arthropod toxins reduce channel responses to non-NMDA receptor agonists. Modes of action and implications for physiology and pathophysiology are discussed.     

Gamma-aminobutyric acid and the high pressure neurological syndrome

Sodium valproate, nipecotic acid, diaminobutyric acid (DABA) and beta-alanine are drugs which enhance transmission mediated by gamma-aminobutyric acid (GABA) by a variety of mechanisms. They were used to study the role of GABA in the high pressure neurological syndrome (HPNS) in the rat. Sodium valproate, nipecotic acid and DABA reduced the increase in slow waves seen in the electroencephalogram (EEG) of control rats at pressures above 10-20 ATA; however, only sodium valproate had a beneficial effect on the behavioural signs of the high pressure neurological syndrome (tremor, myoclonus and convulsions). Sodium valproate is also thought to decrease neurotransmission produced by excitatory amino acids; thus, these results suggest that GABA is not one of the major neurotransmitters involved in all aspects of the high pressure neurological syndrome and that changes in excitatory neurotransmission may affect the behavioural signs.     

Non-anaesthetic steroids ameliorate the high pressure neurological syndrome in rats

The hypothesis that non-anaesthetic compounds, structurally related to specific anaesthetics, can protect against the high pressure neurological syndrome was tested. Infusion of two structural analogues of alphaxalone (3α-hydroxy-5α pregnane-11,20 dione) during pressurisation of rats with helium and oxygen gas mixtures (total pressure 80–100 ATA; inspired oxygen partial pressure 0.5 ATA) ameliorated the severe tremors associated with the high pressure neurological syndrome without any shift in tremor frequency (11–14 Hz). The steroid analogues which were selected (Δ16 and 3β-hydroxy-alphaxalone) have no known general anaesthetic effects and present an unexpected structural approach to the pharmacology of the syndrome. It may now be possible to investigate, treat or prevent the syndrome by the use of selective drugs without more generalised anaesthetic effects.     

2-Amino-phosphonoheptanoic acid protects against the high pressure neurological syndrome

In the present study, pretreatment of rats with captopril significantly stimulated prostacyclin (PGI₂) synthesis by their isolated aorta. This effect was maximal at captopril doses of 1.0 μmol/kg body weight. When added directly into the incubation buffer, captopril at final concentrations of 50 and 500 nM also increased the synthesis of PGI₂ by isolated rat aorta. Our results show that captopril stimulates PGI₂ synthesis in vascular tissue and that this effect may be due at least in part to a direct action of this substance.     

Disruption of learning by excitatory amino acid receptor antagonists

Compounds that act as competitive or uncompetitive N-methyl-D-aspartate (NMDA) antagonists, glycine/NMDA-site antagonists, or alpha-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxalzolepropionic acid (AMPA)-receptor antagonists were evaluated for effects on a repeated acquisition of behavioral chains schedule by rats. Responding by rats was maintained by food presentation under a repeated acquisition or a performance procedure. Under the repeated acquisition procedure, subjects acquired a different three-response chain each daily session. Thus, each day a new learning curve could be generated for each animal thereby providing a repeated measure of learning. Food was presented under a second-order fixed-ratio three (FR3) schedule. Under the performance schedule rats responded under the same second-order FR3 schedule of food presentation: however, instead of a new sequence being presented each day, the same sequence of responding was required for each daily session. Both the competitive (CGS 19755) and uncompetitive (dizocilpine) NMDA antagonists distrupted repeated acquisition at doses that did not disrupt performance. In contrast, the glycine/NMDA antagonist MDL 104,653 or the competitive AMPA receptor antagonist LY 293558 did not disrupt acquisition or performance up to doses that suppressed responding. These results suggest there are different roles for various excitatory amino acid receptors, or sites on the NMDA receptor, in the neural bases of learning and that the disruption of acquisition by glutamate antagonists is dependent upon the particular receptor at which they have activity as well as the particular modulatory site of action.     

Interactions of gamma-aminobutyric acid and noradrenaline in the high pressure neurological syndrome

The effects on the high pressure neurological syndrome (HPNS) of reducing brain noradrenaline (NA) levels were studied in adult rats. The onset of tremors and convulsions, which occur as pressure is increased, were used as endpoints for assessing the onset and severity of the HPNS. Neonatal treatment with 6-hydroxydopamine (6-OHDA; 100 mg kg-1 i.p. alternate days from birth for 2 weeks) which depleted brain NA, produced no change in the HPNS as assessed by the appearance of tremors and convulsions. A second series of NA-depleted rats and equivalent controls were treated with a GABA agonist, muscimol, 0.1 microgram intracerebroventricularly. Subsequently the rats were exposed to pressure and the onset and severity of the HPNS was assessed by observation of tremors and convulsions. A combination of NA depletion and intracerebroventricular injection of muscimol significantly raised the onset pressures for tremors and convulsions, i.e. delayed the appearance of the HPNS. These results are consistent with the HPNS being associated with a disturbance in the balance of two or more neurotransmitter systems, rather than simply an increase or reduction in levels of a single transmitter.     

Effect of excitatory amino acid receptor antagonists on apomorphine-, oxytocin- and ACTH-induced penile erection and yawning in male rats.

The effect of excitatory amino acid receptor antagonists, (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclo-hepten-5, 10-imine hydrogen maleate ((+)-MK-801), (+/-)-3-(2-carboxy-piperazin-4-yl)-propyl-1-phosphonic acid (CPP), 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and (+/-)-2-amino-4-phosphonobutanoic acid (AP-4), on penile erection and yawning induced by subcutaneous apomorphine (80 micrograms/kg), intracerebroventricular (i.c.v.) oxytocin (30 ng) and adrenocorticotropin (ACTH)-(1-24) (10 micrograms) was studied in male rats. Intraperitoneal (0.1-0.4 mg/kg) and i.c.v. (10-50 micrograms) (+)-MK-801 prevented dose dependently the penile erection and yawning induced by the three drugs. The (+)-MK-801 effect coincided with the appearance of head weaving, body rolling, hyperlocomotion and ataxia. Haloperidol (0.5 mg/kg i.p.) antagonized the prevention by (+)-MK-801 of oxytocin responses. Penile erection but not yawning was also prevented by high, but not low doses of CPP and CNQX, which impaired motor performance, AP-4 was ineffective at all doses tested. The above compounds were ineffective when injected into the paraventricular nucleus of the hypothalamus, the brain area where apomorphine and oxytocin act to induce penile erection and yawning. The results suggest that excitatory amino acid transmission is not involved in the expression of penile erection and yawning induced by the above compounds.     

The intrathecal administration of excitatory amino acid receptor antagonists selectively attenuated carrageenan-induced behavioral hyperalgesia in rats.

A single unilateral injection of carrageenan (4.5-6.0 mg in 0.15-0.20 ml saline) into the rat hindpaw induced behavioral hyperalgesia as evidenced by a significant reduction in hindpaw withdrawal latency to a noxious thermal stimulus. The involvement of N-methyl-D-aspartate (NMDA) receptors in this model of hyperalgesia was examined by intrathecal administration of the selective excitatory amino acid (EAA) receptor antagonists: (+/-)-2-amino-5-phosphonopentanoic acid (AP-5), (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), ketamine hydrochloride (ketamine), 7-chlorokynurenic acid (7-Cl kynurenic acid), and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). The effects of dizocilpine maleate (MK-801) were studied under the same conditions and published previously (Ren et al., 1992) and the data are presented for comparison. While the withdrawal latencies of the non-injected paws and of the paws of naive rats were not significantly affected by application of the EAA receptor antagonists at doses tested, the paw withdrawal latencies of the carrageenan-injected paws were elevated dose dependently. The rank order of potency of these agents to reduce hyperalgesia was: MK-801 greater than or equal to AP-5 greater than or equal to CPP = 7-Cl kynurenic acid = ketamine much greater than CNQX greater than 0. In contrast, intrathecal injection of the opioid receptor agonists, [D-Ala2,MePhe4,Gly-ol5]enkephalin (DAMGO, mu-selective) and [D-Pen2,D-Pen5] enkephalin (DPDPE, delta-selective), produced antinociception in both injected and non-injected paws. DAMGO was much more potent, while DPDPE was less potent, than MK-801.(ABSTRACT TRUNCATED AT 250 WORDS)     

Behavioural and electroencephalographic effects of excitatory amino acid antagonists and sigma opiate/phencyclidine-like compounds in rats

The present work was a comparative study of the electrocortical and behavioural effects of phencyclidine-like drugs and sigma-opiate drugs and of some excitatory amino acid antagonists. Phencyclidine-like drugs and sigma-opiate drugs elicited three dose-dependent electrocortical patterns: 1. Increase of periods of cortical desynchronization 2. Increase of the amplitude of fast (20-30 Hz) cortical waves; 3. Appearance of typical cortical complexes consisting of slow-wave (0.5-2 Hz) sharp-wave complexes. These effects were accompanied by excitatory motor symptoms, such as stereotyped movements of the neck and limbs. The NMDA competitive antagonist D,L-2-amino-5-phosphonovaleric acid (D,L-AP5), administered intracerebroventricularly (0.25-2 mumol), elicited phencyclidine-like stereotyped behaviour and cortical desynchronization, but failed to elicit/sigma typical cortical complexes. The non-selective amino acid antagonist cis-2,3 piperidine dicarboxilic acid (cis 2,3-PDA), administered intraperitoneally (3.3 mmol) failed to elicit both stereotyped behaviour and typical phencyclidine/sigma cortical complexes. The results strongly suggest the hypothesis that a reduction of excitatory amino acid neurotransmission, utilizing N-methyl-d-aspartate (NMDA)-preferring receptors, may be involved in the genesis of phencyclidine/sigma-induced stereotyped behaviour.     

Structure-activity relationships in the development of excitatory amino acid receptor agonists and competitive antagonists

Development of new selective ligands for excitatory amino acid receptors has been fundamental in supporting this rapidly developing field. Some of the most important ligands have come from the laboratories of Jeff Watkins, Povl Krogsgaard-Larsen and Tage Honoré, who collaborate in this double-length review to describe the chemical features and SARs of agonists and antagonists, particularly those features associated with subtype selectivity.     

Adenosine outflow from hippocampal slices evoked by ischemic-like conditions: Effect of the excitatory amino acid antagonists

The effect of amino acid receptor antagonists on adenosine and inosine outflow evoked by 5 or 10 min ischemia-like conditions “in vitro” (glucose-free Krebs solution gassed with 95% N₂/5% CO₂) was investigated on rat hippocampal slices. The viability of the slices after ischemia was evaluated by extracellular recording of the evoked synaptic responses in the CA1 region. The NMDA receptor antagonist D(?)-2-amino-7-phosphonoheptanoic acid (D-AP7 100 μM) and the non-NMDA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX 10 μM) did not significantly decrease the adenosine and inosine outflow evoked by ischemia. The glutamate uptake blocker dihydrokainate (500 μM) also did not affect the outflow of adenosine and inosine. It is concluded that ischemia-like conditions “In vitro” evoke and adenosine and inosine outflow, through a mechanism which does not involve excitatory amino acids. © 1993 Wiley-Liss, Inc.     

Drugs that increase gamma-aminobutyric acid transmission protect against the high pressure neurological syndrome

1 The effects on the high pressure neurological syndrome (HPNS) of drugs which facilitate gamma-aminobutyric acid (GABA) transmission were investigated. Threshold pressures for the onset of the behavioural signs of the HPNS in mice--tremors and convulsions were established. 2 Flurazepam hydrochloride 20 and 10 mg/kg and sodium valproate 800 and 400 mg/kg substantially raised the threshold pressures for both tremor and convulsions. 3 Amino-oxyacetic acid 35 and 25 mg/kg and diaminobutyric acid 600 mg/kg also significantly increased the thresholds. Muscimol 1 mg/kg (and 150 ng i.c.v.) was ineffective at non-toxic doses. 4 These effects paralleled the drugs' ability to raise the convulsion threshold to intravenous infusion of bicuculline in mice. 5 These results demonstrate that drugs with actions more selective than those of the general anaesthetics are effective against the HPNS. It is also possible that there is a GABAergic component to the effects of general anaesthetics on the HPNS.     

Brain monoamines and the high pressure neurological syndrome

The effects of high pressure on the whole brain concentrations of 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), dopamine (DA), and noradrenaline (NA) were determined in the mouse. Application of high pressure (over 100atm) elevated the brain 5-HIAA concentrations but had little or no influence on the concentrations of 5-HT, DA, or NA. The pressure-induced increase in 5-HIAA was still present after the administration of the monoamine-depleting drugs, parachlorophenylalanine (PCPA), α-methyl-p-tyrosine (α-MPT), reserpine, and FLA-63. Reserpine (4 mg/kg) and FLA-63 (50mg/kg) lowered the pressure at which tremors, convulsions, and death occurred. In contrast, PCPA (3 doses of 300 mg/kg) and α-MPT (2 doses of 250 mg/kg) did not alter the threshold pressures for the behavioural end-points in the high pressure neurological syndrome. Thus, the depletions of 5-HT or of DA and NA alone do not seem to affect the response to pressure, and it may be that alterations in tremor and convulsion threshold pressures are dependent upon a balance of all the monoamine neurotransmitter systems. The involvement of monoamines in the high pressure neurological syndrome resembles to some extent their postulated roles in electroshock, pentylenetetrazol and auditory convulsions.     

Comparison of sigma- and kappa-opiate receptor ligands as excitatory amino acid antagonists

Using the technique of microelectrophoresis in pentobarbitone-anaesthetized cats and rats, the effects of benzomorphans, with known actions at sigma- and kappa- opioid receptors, were tested on responses of spinal neurones to amino acids and acetylcholine. The racemic mixture and both enantiomers of the sigma opiate receptor agonist, N-allylnormetazocine (SKF 10, 047), and the dissociative anaesthetic, ketamine, reduced or abolished excitation evoked by N-methyl-aspartate (NMA) with only small and variable effects on responses to quisqualate or kainate. (+)-SKF 10, 047 was 1.2 +/- 0.7 times more potent than the (-)-enantiomer in antagonizing NMA. On Renshaw cells, (+)-SKF 10, 047 enhanced responses to acetylcholine whereas the (-) enantiomer produced only a small reduction. The kappa- opiate receptor agonist, ethylketocyclazocine, had no selective effects on responses to amino acids or to acetylcholine. We conclude that actions at sigma- but not kappa-, opiate receptors are responsible for the NMA antagonism observed with benzomorphans.     

Inhibitory influence of excitatory amino acid antagonists on penicillin-induced epileptiform bursting in rat hippocampal slices

The inhibitory influence of excitatory amino acid (E.A.A.) antagonists such as kynurenic acid, 2-amino-5-phosphonopentanoic acid (AP5), cis-2,3-piperidine dicarboxylic acid (cis-2,3 PDA) and (+)-5-methyl-10,11,-dihydro-5H-dibenzo(a,d)cyclo-hepten-5,10-imine maleate (MK 801), has been studied on the epileptiform activity elicited in rat hippocampal slices, bathed in penicillin (1 mM). The rank of the inhibitory potency was: MK 801 greater than kynurenic acid greater than cis 2,3 PDA greater than AP5. Moreover, only MK 801 was able to block the last population spike of the penicillin-induced epileptiform bursting in 100% of the experiments. The data indicate that the antiepileptic activity of E.A.A. antagonists on the penicillin epileptiform bursting in CA1 pyramidal cells is low and limited, indicating that the hippocampal area is not the primary site of the anticonvulsant activity of E.A.A. antagonists.     

Agonists, antagonists and modulators of excitatory amino acid receptors in the guinea-pig myenteric plexus

1. The receptors for glutamic acid (L-Glu) present in the guinea-pig myenteric plexus-ileal longitudinal muscle preparation have been studied by measuring the muscle contraction induced by numerous putative endogenous agonists acting at these receptors. Furthermore, the actions of different concentrations of antagonists, glycine, Mg2+ and Ca2+ on the ileal contractions induced by L-Glu have been evaluated. 2. The EC50 values of the most common putative endogenous agonists of these receptors were: L-Glu 1.9 X 10(-5) M; L-aspartate 8 X 10(-5) M; quinolinate 5 X 10(-4) M; L-homocysteate 1.4 X 10(-4) M; the dipeptide aspartyl-glutamate 8 X 10(-5) M, while N-acetyl-aspartyl-glutamate was inactive. Among the molecules used to classify excitatory amino acid receptors, N-methyl-D-aspartate (NMDA) was the most potent (EC50 5 X 10(-4) M). Kainic and quisqualic acids were almost completely inactive. 3. The responses to L-Glu were competitively antagonized by 2-amino-5-phosphonovaleric acid. They were, also, prevented by hyoscine (10(-7) M) and by tetrodotoxin (3 X 10(-7) M), suggesting that the L-Glu-induced ileal contraction was in some way dependent upon an action on the myenteric cholinergic neurones. Kynurenic acid was a non-competitive antagonist, gamma-D-glutamyl-taurine (10(-4) M) and aminophosphonobutyric acid (10(-4) M) did not modify the L-Glu-induced contractions. 4. Glycine (10(-5) M) significantly potentiated the effects of glutamate especially when the ionic composition of the superfusion medium contained concentrations of Ca2+ in the range of 0.6-1.2 mM. Strychnine 3 X 10(-5) M did not modify the actions of glycine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Perspectives for the use of excitatory amino acid ionotropic receptor antagonists as antiepileptic drugs

Numerous antagonists of glutamate, belonging to N-methyl-D-aspartate (NMDA) and non-NMDA receptor antagonists, were demonstrated to display anticonvulsant activity in various models of experimental epilepsy. Also, the glutamate antagonists enhanced the protective activity of conventional antiepileptic drugs against maximal electroshock-induced seizures in mice. However, some combinations of conventional antiepileptics with NMDA receptor antagonists resulted in profound adverse effects. In contrast, the antagonists for the non-NMDA receptor group potentiated the anticonvulsant activity of conventional antiepileptics, with no side-effects being observed. In some cases, the combined treatments were superior to the administration of an antiepileptic drugs alone, as regards adverse effects. In no case, glutamate antagonists affected the plasma levels of antiepileptic drugs. These data seem to indicate that non-NMDA receptor antagonists bear more important clinical potential as adjuvant antiepileptic drugs.