A. Nistri

Pharmacology of a novel effect of γ-aminobutyric acid on the frog optic tectum in vitro

Abstract Excitatory postsynaptic field potentials (termed U1 and U2 waves) were extracellularly recorded from superficial layers of the frog optic tectum in vitro following stimulation of the contralateral optic nerve. γ-Aminobutyric acid (GABA) largely enhanced the U1 and U2 waves, an effect which was progressively diminished by lowering the extracellular Cl− content. Muscimol or 3-aminopropanesulphonic acid (3-APS) was more potent than GABA in enhancing the U1 and U2 waveforms, whereas trans-4-aminocrotonic acid (TACA) was equipotent, its cis isomer (CACA) being less active. The action of GABA, TACA and CACA was relatively insensitive to bicuculline (100 μM). Pentobarbitone increased the U1 and U2 waves and antagonized the action of GABA. Midazolam did not change the GABA-induced responses. Picrotoxin antagonized both pentobarbitone and GABA effects. It is suggested that in the frog optic tectum GABA can activate an atypical receptor mechanism blocked by picrotoxin but comparatively insensitive to bicuculline.

An unusual effect of γ-aminobutyric acid on synaptic transmission of frog tectal neurones in vitro

1 Bath‐applied γ‐aminobutyric acid (GABA) enhanced, in a dose‐dependent fashion, the amplitude of optic nerve‐evoked monosynaptic excitatory responses of the frog optic tectum superfused in vitro at 7°C. 2 Muscimol was more potent than GABA in eliciting similar effects. 3 GABA‐induced responses were antagonized by picrotoxin and were insensitive to bicuculline or strychnine. 4 Raising the bath temperature to 20°C reduced the potency of GABA on these preparations. 5 No significant effect of GABA on the compound action potential of the whole optic nerve was found. 6 These data indicate that GABA can amplify visual inputs to the tectum through bicuculline‐insensitive mechanisms.

Effects of glutamate and glutamic acid diethyl ester on the lobster muscle fibre and the frog spinal cord

The effect of bath applications of glutamic acid diethyl ester (GDE) and glutamate on the lobster muscle fibre and the from spinal cord were examined. In the lobster muscle fibre GDE (2 X 10(-3) M) did not antagonize the depolarizing action of glutamate (10(-4) M). In the frog spinal cord a small reduction in the excitatory effects of glutamate (10(-4)-10(-3) M) after GDE pretreatment was found only if the latter was given in very high doses (over 2 X 10(-3) M) which stimulated the neuronal firing. These observations show that GDE is not a specific antagonist of glutamate in these tissues.

An electrophysiological study of the action of N-methyl-d-aspartate on excitatory synaptic transmission in the optic tectum of the frog in vitro

Excitatory synaptic field potentials, induced by stimulating optic nerve fibers, were recorded from in vitro preparations of the optic tectum of the frog. Bath-applied N-methyl-D-aspartate (NMDA), glutamate or quisqualate elicited transient enhancement in these field potentials, followed by a sustained depression reversible on washout. Responses to glutamate or quisqualate and the amplitude of control synaptic potentials, were not affected by the NMDA receptor antagonists aminophosphonovalerate (APV), 3(2-carboxypiperazin-4-yl)propyl-1-phosphonate (CPP), ketamine, magnesium ions or dizocipiline (MK 801) which, on the other hand, blocked the effects of NMDA. The antagonist dinitroquinoxaline-2,3-dione (DNQX), which is preferential for non-NMDA receptors, blocked the action of glutamate and synaptic transmission. In the presence of strychnine, glycine reversed the block of NMDA-mediated responses caused by magnesium. It is suggested that in the optic tectum of the frog, glutamate is the excitatory transmitter of at least one class of optic nerve fibers and that it acts through non-NMDA receptors. Although this area of the brain contains a well-developed NMDA receptor system, its function in physiological synaptic transmission remains to be elucidated.

Pharmacological characterization of D‐aminophosphonovaleric acid antagonism of amino acid and synaptically evoked excitations on frog motoneurones in vitro: an intracellular study

1 The effect of D‐aminophosphonovaleric acid (D‐APV) on the depolarizations induced by N‐methyl‐D‐aspartate (NMDA), glutamate, aspartate or quisqualate was studied with intracellular recordings from frog motoneurones in vitro. 2 D‐APV (0.5–10 μM) produced a slight hyperpolarization of the motoneuronal membrane without significant changes in input conductance. 3 In control and tetrodotoxin‐containing solutions the depolarizations induced by NMDA were strongly reduced by D‐APV while quisqualate depolarizations were unaffected. Responses to glutamate and aspartate were antagonized to an intermediate level. The relatively small conductance increases evoked by excitatory amino acids were unaltered in solutions containing D‐APV. 4 The amplitude of monosynaptic excitatory postsynaptic potentials (e.p.s.ps) was strongly depressed by D‐APV. The amplitude of polysynaptic e.p.s.ps was little changed but their decay time was reduced. 5 It is suggested that D‐APV is a powerful and selective NMDA receptor antagonist and that an endogenous amino acid acting via NMDA receptors may be the transmitter of monosynaptic e.p.s.ps on frog motoneurones.

Excitatory amino acid-induced responses of frog motoneurones bathed in low Na+ media: an intracellular study

Motoneurones of the frog spinal cord slice preparation were impaled with microelectrodes and superfused at 7 degrees C with the excitatory amino acids glutamate, quisqualate or N-methyl-D-aspartate. The role of Na+ in the action of these amino acids was studied by comparing amplitude matched depolarizations obtained in control Ringer solution with the responses recorded from the same cells after replacing (86-100%) Na+ by choline or glucosamine. Effective replacement of extracellular Na+ proved to be a rather slow process requiring 30-60 min. In glucosamine solution depolarizations evoked by glutamate, N-methyl-D-aspartate or quisqualate were abolished or strongly reduced with recovery following return to control Ringer. In choline solution, glutamate and N-methyl-D-aspartate effects were blocked whereas the quisqualate response was surprisingly unaffected. Mn2+ (2 mM) added to choline solution strongly diminished the action of quisqualate. These results suggest that Na+ was important in mediating amino acid responses and that quisqualate activated an additional conductance mechanism (perhaps to Ca2+) unmasked only in choline-containing solution.

Complex effects of baclofen on synaptic transmission of the frog optic tectum in vitro

In the frog optic tectum in vitro, low concentrations of the GABAB agonist (-)-baclofen reversibly depressed field potentials evoked by optic nerve stimulation. This effect, which was dose-dependent and stereo-specific, differed from the potent facilitatory action of bath-applied GABA. A transient enhancement in field potential amplitude often preceded the more consistently observed depression. Both phases of the baclofen response were insensitive to the GABAA antagonist picrotoxin. Since the effects of baclofen and GABA were different, it is suggested that GABAB receptors were unlikely to mediate the action of GABA on synaptic transmission in the optic tectum.

Biphasic effects of glycine on synaptic responses of the frog optic tectum in vitro

Electrophysiological recordings from the in vitro amphibian optic tectum were used to study the action of bath-applied glycine (Gly) on synaptic transmission between optic tract fibres and tectal neurones. Gly enhanced synaptic field potentials (ED50 = 0.3 mM) and, when applied in concentrations greater than or equal to 1 mM subsequently blocked them. All these effects of Gly were antagonized by 0.2-1 microM strychnine. Recordings from optic tract afferent fibres indicated that Gly increased the amplitude of small submaximal compound action potentials while depressing those which were near maximal. It is suggested that Gly exerted a novel modulatory effect on synaptic transmission via a combination of pre- and postsynaptic sites of action.

The excitation of frog motoneurones in vitro by the glutamate analogue, dl-α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA), and the effect of amino acid antagonists

Intracellular recordings were made from motoneurones of the frog spinal cord in vitro and the excitatory effects of the glutamate analogue DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) examined. AMPA (25-100 microM) depolarized motoneurones with an increase in input conductance and an increase in spontaneous cell firing. The time-course of the AMPA response was prolonged compared to L-glutamate. D-Aminophosphonovalerate (1 microM) and D-aminoadipate (100 microM) did not antagonize AMPA-induced excitations while depressing L-glutamate responses. Glutamate diethylester up to 1 mM was not found to be a useful amino acid antagonist since it did not significantly reduce the depolarizations to quisqualate, L-glutamate and AMPA. The results are compatible with an action of AMPA on receptors distinct from those to N-methyl-D-aspartate and shared by quisqualate.

A study of amino acid-activated currents recorded from frog motoneurones in vitro

Superfusion of the excitatory amino acids glutamate (1-2 mM), quisqualate (15-30 microM) and N-methyl-D-aspartate (NMDA: 15-30 microM) induced inward currents in voltage-clamped motoneurones, in vitro. Typically the NMDA and quisqualate currents had prolonged time courses relative to glutamate currents. No desensitization was apparent during repeated agonist application. D-2-Amino-5-phosphonovalerate (10 microM) selectively antagonized the NMDA current without affecting the quisqualate current; the glutamate current was partially reduced reflecting its mixed agonist properties.