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The goal of these investigations was to study the role of tachykinin NK2 receptors in neonatal spinal cords using the selective NK2 receptor agonist [β-Ala8]neurokinin A-(4–10) and the new NK2 receptor antagonist GR 94800. Experiments were performed with superfused hemisected rat and gerbil spinal cords. Dorsal roots were electrically stimulated and the synaptically elicited responses and the DC-potentials were recorded extracellularly from the corresponding ventral roots. [β-Ala8]neurokinin A-(4–10) depolarized ventral roots (0.01–10 μM) and increased their spontaneous activity in a concentration-dependent manner. These effects of [β-Ala8]neurokinin A-(4–10) were reduced by GR 94800. The action of GR 94800 was selective because the depolarizing effects of similar magnitude evoked by the NK1 receptor agonist [Sar9, Met(O2)11]substance P were not affected by GR 94800. The pA2 values of GR 94800 amounted to 6.0±0.4 in the rat and 5.4±0.3 in the gerbil. The NK2 receptor agonist was more potent in the rat than in the gerbil. The estimated EC50 (means ± S.E.M.) was found to be 3.9 + 6.0/ − 1.3 μM in the rat and 2.4 + 2.9/ − 1.3 μM in the gerbil spinal cord. The NK2 receptor agonist [β-Ala8]neurokinin A-(4–10) potentiated the monosynaptic reflex evoked by dorsal root stimulation. The potentiation manifested itself as an increase in the amplitude of the early component of the response. The receptor type mediating this effect could not be elucidated. The potentiation ranged between 30 ± 27 and 110 ± 36% (0.3 and 10 μM), respectively. Tetrodotoxin reduced the depolarizing action elicited by the NK2 receptor agonist, suggesting that part of the [β-Ala8]neurokinin A-(4–10) effect is probably located presynaptically. In contrast to the NK1 receptor antagonist (±)-CP-96,345, the NK2 receptor anatgonist GR 94800 showed no species differences. In conclusion, NK2 receptors appear to have a physiological role in regulating spinal neuronal activity.

6-Cyano-7-nitroquinoxaline-2,3-dione (CNQX) (10 microM) depressed dorsal root-evoked ventral and dorsal root potentials of the in vitro immature rat spinal cord to 26.3 +/- 5.2 S.E.M. and 40.8 +/- 2.7% of control values respectively. These depressant effects of CNQX were partially reversed by D-serine (EC50 values 39.7 microM +/- 8.7 S.E.M. N = 6 and 34.9 +/- 12.5 microM, N = 5 for ventral root potential and dorsal root potential respectively). Under our experimental conditions, which included the presence of Mg2+ (0.75 mM) in the bathing medium, no measurable potentiation of these synaptic reflexes by D-serine was recorded in the absence of CNQX. These data indicate that CNQX, in addition to its depressant effect at non-NMDA receptors, depresses an NMDA receptor-mediated component of segmental transmission through its action at the glycine site of the NMDA receptor complex.

The novel tachykinin receptor antagonist CGP49823 ((2R,4S)-2-benzyl-1-(3,5-dimethylbenzoyl)-4-(quinolin-4-y lmethylamino)piperidine) has been compared with three other selective non-peptide tachykinin NK1 receptor antagonists. The drugs were tested as antagonists of the depolarization of spinal motoneurones induced by bath application of the selective tachykinin NK1 receptor agonist septide-(6-11) (300 nM) for 120 s at 15 min intervals. The antagonists were bath applied and the depolarizations were recorded from lumbar ventral roots of 7 to 12 day old rat and gerbil hemisected spinal cords in vitro. The gerbil preparation is considered to model the human species variant of the tachykinin NK1 receptor. With the exception of SR140333 ((S)-1-[2-[3-(3,4-dichlorophenyl)-1-[[3-(1-methylethoxy)phenyl]ace tyl]-3-piperidinyl]ethyl]-4-phenyl-1-azoniabicyclo[2.2.2]octane chloride), the antagonists were approximately thirty-fold more potent on gerbil preparations. The respective mean IC50 values from gerbil preparations produced by CP96345 ((2S-cis)-2-(diphenylmethyl)-N-[(2-methoxyphenyl)methyl]-1-azabicy clo[2.2.2]octan-3-amine), CGP49823, SR140333 and CP99994 ((2S-cis)-N-[(2-methoxyphenyl)methyl]-2-phenyl-3-piperidinamine) were, in microM +/- S.E. (n) 0.10 +/- 0.02 (6), 0.22 +/- 0.03 (6), 0.30 +/- 0.10 (5) and 0.38 +/- 0.02 (5) and the corresponding values from the rat preparations were 3.7 +/- 0.4 (5), 7.8 + 1.3 (5), 1.06 +/- 0.16 (6) and 10.5 +/- 2.2 (7). Dominance of tachykinin NK1 receptor activity in the measured responses was confirmed by low potency of the tachykinin NK2-selective antagonist SR48968 ((S)-N-methyl-N[4-(4-acetylamino-4-phenyl piperidino)-2-(3,4-dichlorophenyl)butyl] benzamide) which yielded an IC50 value of 12.0 +/- 2.8 (5) on gerbil preparations and produced less than 50% depression of septide-induced depolarization of rat motoneurones at the highest concentration (100 microM) tested.

The effects of some central depressant drugs on amino acid-induced depolarization of motoneurons have been determined in the isolated hemisected frog spinal cord. Motoneuronsdepolarization was recorded from ventral roots and measurements were made in the presence or absence of procaine or tetrodotoxin to minimize indirect effects of both drugs and amino acids. Chlorpromazine (0.05–0.1 mM) and diazepam (0.5 mM) produced a similar differential pattern of depression of amino acid-induced depolarizations. Responses induced by L-homocysteate were markedly antagonized by these drugs, while responses to quisqualate were unaffected. L-Aspartate-induced responses were antagonized more than L-glutamate-induced responses. This pattern of antagonism resembles that previously described for Mg2+. In contrast, pentobarbital (0.1 or 0.3 mM), and the inhibitory amino acids GABA and β-alanine (0.5–1.0 mM), depressed amino acid-induced responses in a more uniform manner. The differential effects observed with chlorpromazine and diazepam provide further support for the possibility that responses to excitant amino acids structurally related to L-glutamate may have different underlying mechanisms.

The specificity of the neurodepressant actions of D-alpha-aminoadipate, alpha,epsilon-diaminopimelic acid, HA-966 (HAP) and Mg2+ has been investigated. On the isolated spinal cord of the neonatal rat, ventral root depolarizations produced by kainate, substance P, carbachol and noradrenaline were relatively unaffected by the same concentrations (0.25--1 mM) of the agents as those which reduced synaptic activity and ventral root depolarizations produced by N-methyl-D-aspartate (especially), L-aspartate and L-glutamate. The same or higher concentrations of the agents did not affect excitatory transmission in the isolated rat superior cervical ganglion. It is proposed that the agents specifically block synaptic transmission mediated by an excitatory amino acid.

Neurotransmission in isolated hemisected spinal cord preparations from immature rats was depressed by micromolar levels of baclofen (threshold 0.5 μM). The depressant action of baclofen was not antagonised by bicuculline and baclofen, unlike GABA, did not depolarize primary afferent fibres. Neurotransmission in isolated vas deferens, anococcygeus muscle and superior cervical ganglion of the rat was unaffected by baclofen (0.1–1 mM). Depolarization of motoneurones, as recorded in ventral roots of tetrodotoxin-blocked spinal cord preparations, induced by excitant amino acids, substance P, noradrenaline or carbachol was unaffected by baclofen (250 μM or higher). The depressant action of baclofen on spinal cord preparations was similar to that produced by the excitant amino acid antagonist α,ϵ-diaminopimelic acid. A structure-activity study showed that the (−)-isomer of baclofen was over 20 times more potent than the (+)-isomer as a spinal depressant. Also the position and nature of the halogen substitutent in the ring is critical with baclofen giving optimal activity. It is concluded that the depressant action of baclofen results from depression of the presynaptic release of excitant amino acid transmitter(s).

Three N-methyl-D-aspartate (NMDA) antagonists (±)2-amino-5-phosphonopentanoate (AP5), 3-((±)-2-carboxypiperazin-4-yl)prophyl-1- phosphonic acid (CPP) and ((±)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine maleate) (MK-801) have been tested for selectivity against depolarization of motoneurones induced by carbachol, 5-hydroxytryptamine, noradrenaline and substance P in isolated immature rat spinal cord preparations. AP5 (400 μM) and CPP (50 μM) gave mean dose-ratios, for antagonism against NMDA, of 103 ± 14.9 S.E.M. (eight preparations) and 34.1 ± 1.9 S.E.M. (14 preparations). MK-801 (1 and 10 μM) was the most potent of the three antagonists yielding dose ratios greater than 100 after 120 min treatment. MK-801 potentiated responses induced by 5-hydroxytryptamine and noradrenaline given dose-ratios of 0.22 ± 0.16 S.E.M. and 0.20 ± 0.06 S.E.M., respectively (four preparations). The three antagonists produced no significant antagonism of the non-amino acid agonists (four preparations for each agonist) when dose-ratios against NMDA were at least 40. The observations support the use of these antagonists as tools to identify sites of identify sites of excitatory amino acid-mediated transmission.