Lakhbir Singh

The behavioural effects of MK-801: a comparison with antagonists acting non-competitively and competitively at the NMDA receptor

The selective non-competitive NMDA receptor antagonist, MK-801, potently blocked convulsions induced in the mouse by N-methyl-DL-aspartic acid (NMDLA) with an i.v. ED50 dose of 0.2 mg/kg. Similar doses of MK-801 were also effective in blocking seizures induced by pentylenetetrazol (PTZ), electroshock and by sound in audiogenic seizure-prone animals. Other less selective non-competitive NMDA receptor antagonists including phencyclidine (PCP), thienylcyclohexylpiperidine (TCP), (+)-N-allylnormetazocine [+)-NANM, (+)-SKF 10,047) and ketamine also blocked NMDLA-induced seizures with a rank order of potency of MK-801 greater than PCP greater than TCP = (+)-NANM greater than ketamine. The competitive NMDA receptor antagonist, 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP) blocked NMDLA-induced seizures with an ED50 of 4.5 mg/kg, 22- and 560-fold more potently than the competitive antagonists, 2-DL-amino-7-phosphonoheptanoic acid (2-APH) and 2-DL-amino-5-phosphonovaleric acid (2-APV), respectively. MK-801 was the most potent of the non-competitive antagonists to induce a motor syndrome including head weaving, body rolling, increased locomotion and ataxia, characteristic of the behavioural response to PCP in the mouse. The syndrome was also present following injection of the competitive NMDA receptor antagonists, although they were generally less potent (probably a reflection of poor brain penetration) and less efficacious than the non-competitive antagonists. For all compounds except CPP, the anticonvulsant ED50 dose was close to the minimum effective dose to induce motor stimulation: CPP was 5- to 10-fold more potent as an anticonvulsant.(ABSTRACT TRUNCATED AT 250 WORDS)

Modulation of seizure susceptibility in the mouse by the strychnine-insensitive glycine recognition site of the NMDA receptor/ion channel complex.

1 In order to determine whether the strychnine‐insensitive glycine modulatory site on the N‐methyl‐d‐aspartate (NMDA) receptor/ion channel complex is fully activated in vivo, the ability of the selective glycine receptor agonist, d‐serine, to modulate seizure susceptibility in the mouse has been examined. 2 d‐Serine (10–200 μg per mouse, i.c.v.) dose‐dependently increased the potency of NMDLA in inducing seizures in Swiss Webster mice by approximately 3 fold. l‐Serine was without significant effect. 3 The potency of pentylenetetrazol in inducing seizures was also enhanced by d‐, but not l‐serine, although the magnitude of the shift (1.6 fold) was considerably less than for NMDLA. 4 Similar doses of d‐serine were also able to block the anticonvulsant effect of the non‐selective glycine receptor antagonist, kynurenic acid, against seizures induced by NMDLA, but were without effect on the anticonvulsant effect of the competitive NMDA receptor antagonist, 3‐((+)‐2‐carboxypiperazin‐4‐yl)‐propyl‐1‐phosphonic acid (CPP). 5 d‐Serine completely antagonized the protective effect of the selective glycine receptor antagonist, 7‐chlorokynurenic acid, against sound‐induced seizures in DBA/2 mice, but was less effective in this model against the less selective antagonist, kynurenic acid. 6 The results indicate that in vivo, NMDA receptors are not maximally potentiated by endogenous glycine and suggest an important involvement of the glycine modulatory site on the NMDA receptor/ion channel complex in the pathophysiology of epilepsy.

A role for receptors of N-methyl-D-aspartic acid in the discriminative stimulus properties of phencyclidine

Ketamine and (+)-N-allylnormetazocine ((+)-NANM) were found to generalize in a rat operant drug discrimination paradigm to the interoceptive stimulus induced by phencyclidine (PCP). Intraperitoneal administration of the non-competitive antagonist of the N-methyl-D-aspartate (NMDA) receptor, MK-801, and intracerebroventricular injection of the competitive antagonist, 2-DL-amino-7-phosphonoheptanoic acid (2-APH), also resulted in a dose-dependent generalisation to the PCP discriminative stimulus. The results suggest that NMDA receptor antagonism may play an important role in the mediation of the discriminative stimulus properties of PCP. The low potency of MK-801 and 2-APH to displace [3H](+)-NANM binding in vitro argues against an involvement of the haloperidol-sensitive sigma recognition site in the behaviour.

Neurotransmitters and shaking behaviour — more than a ‘gut-bath’ for the brain?

Abstract Drug-induced shakes and twitches, once regarded as a pharmacological curiosity, have considerable potential as a quantitative probe on in vivo receptor function and as a means of unravelling transmitter interactions argue Sheila Handley and Lakhbir Singh . An example is the ability of β-adrenoceptor agonist to enhance shaking due to 5-HT agonists and precursors. This potentiation involves the locus coeruleus and is reduced by chronic antidepressants. The meaning of such interactions will only become clear when we understand more about the control of this misleadingly simple behaviour which can also be evoked by analogues of GABA, glutamate, acetylcholine, opiates and hormones.

Evidence that a proconvulsant action of lithium is mediated by inhibition of myo-inositol phosphatase in mouse brain.

Lithium inhibits myo-inositol mono- and polyphosphatase activity in brain at concentrations similar to those optimal for the treatment of manic depressive psychosis. A consequence of this inhibition is the possibility that the availability of myo-inositol for the regeneration of polyphosphoinositides involved in cellular signalling mechanisms may be reduced. While there are no good models of manic depressive disorders in rodents, lithium is known to alter their behavioural responsiveness to a number of neurotransmitter receptor agonists, but the role of the phosphatidylinositol second messenger system in these effects is unknown. Consistent with the myo-inositol depletion hypothesis, when injected directly into the CNS, myo-inositol, but not its biologically inactive epimer, scyllo-inositol or D-mannitol, has been found to reverse a proconvulsant action of lithium in mice given the muscarinic receptor agonist, pilocarpine.

The discriminative stimulus properties of (+)-HA-966, an antagonist at the glycine/N-methyl-D-aspartate receptor

Using a two-lever operant drug discrimination paradigm, rats have been trained to discriminate between the administration of saline and R-(+)-HA-966 (R-(+)-3-amino-1-hydroxypyrrolid-2-one, 30 mg/kg i.p.) an antagonist at the glycine modulatory site on the N-methyl-D-aspartate (NMDA) receptor/ion channel complex. Drug-appropriate responding was not induced in stimulus generalisation experiments when the non-competitive NMDA receptor antagonist, phencyclidine (PCP, 1-8 mg/kg i.p.) was substituted for (+)-HA-966. Similarly, (+)-HA-966 (6-50 mg/kg i.p.) did not induce drug-appropriate responding in animals trained to discriminate PCP (3 mg/kg i.p.) from saline. The results suggest that the behavioural profile of compounds attenuating the actions of NMDA via blockade of the glycine modulatory site may be substantially different from those acting at the ion channel of the NMDA receptor complex.

In vivo labelling of the NMDA receptor channel complex by [3H]MK-801

An in vivo radioligand binding assay for the N-methyl-D-aspartate (NMDA) receptor channel complex in the mouse brain has been developed using the non-competitive NMDA receptor antagonist [3H]MK-801. In vivo binding of [3H]MK-801 was displaced by MK-801 (ED50 = 0.17 mg/kg i.p.), (-)-MK-801 (1.0 mg/kg), thienylcyclohexylpiperidine (1.8 mg/kg), etoxadrol (5.1 mg/kg) and (+)-SKF 10,047 (34.5 mg/kg). The potency of these drugs in this in vivo binding assay was highly correlated (r = 0.97) with their functional effects as antagonists of N-methyl-DL-aspartate-induced tonic convulsions.

The anticonvulsant and behavioural profile of L‐687,414, a partial agonist acting at the glycine modulatory site on the N‐methyl‐D‐aspartate (NMDA) receptor complex

1 The anticonvulsant and behavioural effects of the glycine/NMDA receptor partial agonist, L‐687,414 (R(+)‐cis‐β‐methyl‐3‐amino‐1‐hydroxypyrrolid‐2‐one) have been investigated in rodents. 2 L‐687,414 dose‐dependently antagonized seizures induced by N‐methyl‐D, L‐aspartic acid (NMDLA, ED50 = 19.7 mg kg−1), pentylenetetrazol (PTZ, ED50 = 13.0 mg kg−1) and electroshock (ED50 = 26.1 mg kg−1) when given intravenously 15min before test, in male Swiss Webster mice but was most potent against audiogenic seizures induced by a 120 dB bell in DBA/2 mice (ED50 = 5.1 mg kg−1, i.p., 30min before test). 3 L‐687,414 also induced impairments of performance in a rotarod test in both Swiss Webster and DBA/2 mice and the ratio [rotarod MED:anticonvulsant ED50] varied between 0.9 and 5, depending on the convulsant used. 4 Similar behaviours to those seen after administration of the non‐competitive NMDA receptor antagonist, MK‐801 (head weaving, body rolling, hyperlocomotion) were seen in the mouse after giving L‐687,414, although the peak effect occurred at a dose (100 mg kg−1) which was 5–20 times the anticonvulsant ED50S, depending on the convulsant used. Unlike MK‐801, however, doses of L‐687,414 that were behaviourally stimulant did not increase dopamine turnover in the nucleus accumbens. 5 Consistent with the interaction of L‐687,414 with the glycine/NMDA receptor, the anticonvulsant, ataxic and motor stimulant effects of the compound were significantly attenuated by the glycine/NMDA receptor agonist, D‐serine (10–100 μg per mouse, i.c.v.). 6 The results show that L‐687,414 is a potent, orally active anticonvulsant with a more benign pharmacological profile than antagonists acting at the ion channel of the NMDA receptor complex. The compound is a useful tool with which to probe the functional role of the glycine co‐agonist site in vivo.

Evidence against an involvement of the haloperidol-sensitive σ recognition site in the discriminative stimulus properties of (+)-N-allylnormetazocine ((+)-SKF 10,047)

1 The involvement of the haloperidol‐sensitive, σ recognition site and the N‐methyl‐d‐aspartic acid (NMDA) receptor in the mediation of the discriminative stimulus properties of (+)‐N‐allylnormetazocine ((+)‐NANM, (+)‐SKF 10,047), has been investigated in the rat by use of a two‐lever, operant drug discrimination paradigm. 2 Six compounds with nanamolar affinity for the σ recognition site ((±)‐pentazocine, (+)‐3‐(hydroxyphenyl)‐N‐propylpiperidine ((+)‐3‐PPP), ditolylguanidine (DTG), haloperidol, (−)‐butaclamol and BMY 14802) were investigated for their ability to generalise or antagonise the (+)‐NANM discriminative stimulus. Each drug was tested at doses found in an ex vivo radioligand binding assay to displace [3H]‐DTG from the central σ recognition site by more than 40%. 3 While (±)‐pentazocine (in the presence of naloxone) generalised and (+)‐3‐PPP partially antagonised the (+)‐NANM cue, the other putative σ ligands were ineffective either as agonists or antagonists at doses clearly occupying the σ site in vivo. 4 Dose‐dependent generalisation to the (+)‐NANM cue was seen with the selective non‐competitive NMDA receptor antagonist, MK‐801, a compound devoid of significant affinity for the σ recognition site. 5 (±)‐Pentazocine was found to antagonise seizures induced in the mouse by NMDLA, a model reflecting antagonism of central NMDA receptors, and a strong correlation was found between the rank order of potency of compounds to generalise to the (+)‐NANM discriminative stimulus and their potencies as anticonvulsants. 6 In conclusion, no evidence was found to substantiate the contention that the discriminative stimulus properties of (+)‐NANM are mediated by the haloperidol‐sensitive σ recognition site. On the other hand, the results are consistent with the interoceptive stimulus being mechanistically based in the NMDA receptor complex.

Anticonvulsant activity of glycine-site NMDA antagonists. 1. 2-carboxyl prodrugs of 5,7-dichlorokynurenic acid

Anticonvulsant activity in the DBA/2 mouse audiogenic seizure model has been obtained in water-soluble prodrug esters of the glycine-site NMDA antagonist 5,7-dichlorokynurenic acid (1), leading to the 1-methyl(2-dimethylamino)ethyl ester 10 (ED50 62 mg/kg i.p.), which shows no behavioural stimulation at the anticonvulsant dose.