Glycine high affinity uptake and strychnine binding associated with glycine receptors in the frog central nervous system

Accumulation of [3H]glycine into synaptosomal fractions occurs by high affinity systems in cerebral cortex, optic tectum, brain stem and spinal cord of the frog. Specific [3H]strychnine binding which appears associated with postsynaptic glycine receptors is also demonstrable in these regions. By contrast, only very low levels of strychnine binding and high affinity glycine uptake occur in higher centers of the rat central nervous system. The relative potencies of small neutral amino acids in competing for [3H]strychnine binding are similar in frog brain and spinal cord. No evidence for a high affinity accumulation of [3H]taurine by synaptosomal fractions of frog spinal cord can be demonstrated. These observations favor glycine rather than taurine as an inhibitory transmitter in frog spinal cord. Moreover, these findings suggest that glycine may have a synaptic role in higher brain centers in the frog.     

Release of [3H]gamma-aminobutyric acid from glial (Müller) cells of the rat retina: effects of K+, veratridine, and ethylenediamine

In several neural systems, glial cells appear to take up and release gamma-aminobutyric acid (GABA) upon depolarization. We have studied the release of [3H]GABA from Müller (glial) cells in the rat retina by a double isotope-labeling technique in which Müller cells are preloaded with 3H-GABA while a population of neurons is prelabeled with [14C]glycine. By autoradiography, we have confirmed that [3H]GABA is taken up by the radially oriented Müller cells, whereas [3H]glycine is accumulated by a subset of amacrine cells (neurons). Using the double-labeling procedure, we have examined the effects of two depolarizing agents, high K+ and veratridine, and the GABA mimetic, ethylenediamine, on transmitter release from glial cells and neurons simultaneously. We found the following. (1) Depolarization with 56 mM K+ released both [3H]GABA and [14C]glycine. About 70 to 80% of this release was blocked in Ca2+-free medium. (2) Veratridine (10 microM) also released both of the transmitters. This release was strongly inhibited by 100 nM tetrodotoxin or 1mM procaine. Under Ca2+-free conditions, less than 20% isotope release was observed. (3) Ethylenediamine released [3H]GABA readily, whereas little [14C]glycine release was observed. Removal of Ca2+ had no significant effect on transmitter release. Furthermore, in Na+-free medium ethylenediamine failed to induce [3H] GABA or [14C]glycine release. These results suggest that high K+ and veratridine release [3H]GABA from Müller cells by a Ca2+-dependent process. Ethylenediamine, on the other hand, appears to induce [3H]GABA release by a Ca2+-independent, carrier-mediated exchange mechanism.     

Activation of gamma-aminobutyric acid GAT-1 transporters on glutamatergic terminals of mouse spinal cord mediates glutamate release through anion channels and by transporter reversal

The effects of gamma-aminobutyric acid (GABA) on the release of glutamate from mouse spinal cord nerve endings have been studied using superfused synaptosomes. GABA elicited a concentration-dependent release of [3H]D-aspartate ([3H]D-ASP; EC50= 3.76 microM). Neither muscimol nor (-)baclofen mimicked GABA, excluding receptor involvement. The GABA-evoked release was strictly Na+ dependent and was prevented by the GABA transporter inhibitor SKF89976A, suggesting involvement of GAT-1 transporters located on glutamatergic nerve terminals. GABA also potentiated the spontaneous release of endogenous glutamate; an effect sensitive to SKF89976A and low-Na+-containing medium. Confocal microscopy shows that the GABA transporter GAT-1 is coexpressed with the vesicular glutamate transporter vGLUT-1 and with the plasma membrane glutamate transporter EAAT2 in a substantial portion of synaptosomal particles. The GABA effect was external Ca2+ independent and was not decreased when cytosolic Ca2+ ions were chelated by BAPTA. The glutamate transporter blocker DL-TBOA or dihydrokainate inhibited in part (approximately 35%) the GABA (10 microM)-evoked [3H]D-ASP release; this release was strongly reduced by the anion channel blockers niflumic acid and NPPB. GABA, up to 30 microM, was unable to augment significantly the basal release of [3H]glycine from spinal cord synaptosomes, indicating selectivity for glutamatergic transmission. It is concluded that GABA GAT-1 transporters and glutamate transporters coexist on the same spinal cord glutamatergic terminals. Activation of these GABA transporters elicits release of glutamate partially by reversal of glutamate transporters present on glutamatergic terminals and largely through anion channels.     

Differential effects of veratridine and potassium depolarization on neuronal and glial gaba release

The effect of veratridine and potassium depolarization on the release of [3H]GABA from neural tissues in which GABA uptake is neuronal (cerebrocortical slices and frog retina) or glial (rat retina, spinal and sympathetic ganglia) was studied. The 'neuronal' but not the 'glial' release of [3H]GABA was greatly increased by KC1 (25 mM) and veratridine (10 microM). The 'neuronal' release of [3H]GABA evoked with KC1 was calcium dependent but the small potassium induced increase in 'glial' release were not reduced in the absence of calcium ions even when additional Mg2+ was included in the medium. Surprisingly, the veratridine induced release of [3H]GABA from cortical slices was greatly potentiated in calcium free medium although the release of [3H]noradrenaline produced by veratridine was calcium dependent. Because of the insensitivity of the glial GABA release mechanism to potassium depolarization, it is concluded that release of endogenous GABA from glial pools in response to physiological increases in extracellular potassium is unlikely to be important.     

Conservation of antigenic epitopes of the inhibitory glycine receptor in rodent and goldfish CNS.

Monoclonal antibodies against the inhibitory glycine receptor of rat spinal cord were used to identify corresponding receptor polypeptides in goldfish CNS. Both Western blot analysis and quantitative receptor immunoassays revealed crossreacting antigens in goldfish brain membranes. A polypeptide of 46 kDa molecular weight is immunologically related to the 48 kDa alpha subunit of the mammalian receptor. Similarly, a large receptor-associated protein of 93 kDa is present both in goldfish and mammals. Throughout the goldfish CNS, glycine-displaceable [3H]strychnine binding codistributes with the alpha subunit protein as determined immunologically. Glycine receptor contents were highest in goldfish medulla oblongata, medium in optic tectum and mesencephalon, whereas little or no receptor was detected in cerebellum, olfactory bulb, and spinal cord. Immunohistochemistry confirmed that the alpha subunit antigen and the 93 kDa protein were located in the plasma membrane of neurons and concentrated in small clusters found on the soma and dendrites. These data indicate that immunological properties and cellular distribution of glycine receptors are conserved from fish to mammals.     

Age-dependent changes in brain glycine concentration and strychnine-induced seizures in the rat

Glycine levels and receptor binding were measured in the medulla and spinal cord of 2-month, 10-month, and 24-month-old Fischer 344 rats. The behavioral response to the administration of the glycine antagonist, strychnine, was also evaluated in 2- and 24-month-old animals to investigate the relevance of these parameters to the susceptibility to seizures. Significant reductions in glycine in both the spinal cord and medulla occurred from 2 to 24 months of age. The glycine precursors, serine and threonine, were decreased only in the spinal cord. [3H]Strychnine binding was also decreased by 38% and 34% in the medulla and spinal cord, respectively, of 24-month-old rats compared to 2-month-olds. [3H]GABA binding was similarly reduced while no age-related changes in [3H]diazepam binding in the spinal cord were detected. Comparison of 2- and 24-month-old animals after systemic injection of 1.75 mg/kg strychnine showed that senescent animals have a higher incidence of seizures and mortality compared to young animals. Decreases in glycinergic neurotransmission may lower strychnine seizure threshold in the aged animal.     

Effect of pentobarbitone on the spontaneous efflux of gamma-amino acids from rabbit retina.

The effect of pentobarbitone on the spontaneous release of radioactivity from rabbit retinas preloaded with [3H]GABA, [3H]DABA, [3H]beta-alanine and [3H]glycine was studied. In high concentration of pentobarbitone (10(-3)M) the rate of spontaneous efflux of neuronal [3H]GABA, [3H]DABA and [3H]beta-alanine was reduced but not that of [3H]glycine. Glial release of [3H]GABA was much less influenced. At lower concentrations (10(-4)-10(-5)M) there was an initial increase in the release of [3H]GABA, [3H]DABA and [oH]beta-alanine followed by a decrease, indicating several components in the release system for the amino acids.     

Pentylenetetrazol and reflex activity of isolated frog spinal cord.

The superfused in vitro frog spinal cord preparation was used to investigate the effects of pentylenetetrazol (PTZ) on the spinal cord. PTZ depressed monosynaptic, but augmented polysynaptic reflexes, and decreased primary afferent deplorization. Concurrently, in Ringer's solution containing sufficient magnesium or cobalt ions to block synaptic transmission, PTZ antagonized the hyperpolarizing effects on motoneurons and the depolarizing effects on primary afferent fibers of the inhibitory amino acids GABA, beta-alanine, taurine, and glycine. PTZ did not affect responses to the excitatory amino acids glutamic acid and aspartic acid. Furthermore, PTZ did not alter high affinity uptake by cord slices, K+ -evoked release of [3H]GABA from them, or the spinal concentration of GABA. These data suggest that PTZ may produce its excitatory effects by postsynaptic blockade of inhibitory processes mediated by GABA (and possibly by other amino acids).     

In vivo release of newly synthesized [H]GABA in the substantia nigra of the rat: relative contribution of GABA striato-pallido-nigral afferents and nigral GABA neurons

The release of [3H]gamma-aminobutyric acid ([3H]GABA) continuously formed from [3H]glutamine has been measured with a push-pull cannula implanted in the substantia nigra of the rat anesthetized with ketamine. Consistent with the high density of GABA terminals coming from both the striato-pallido-nigral afferents, and from GABA nigrofugal neurons, our results showed that a large amount of [3H]GABA was spontaneously released in the reticulata, about 4 times higher than in the compacta. In the absence of calcium the spontaneous [3H]GABA release was reduced (-30%), as well as the K(+)-induced release of [3H]GABA (-66%). Bicuculline (10(-4) M) did not affect the K(+)-evoked release of [3H]GABA, suggesting that autoreceptors on GABA afferent fibers are distinct from the GABAA subtype. Partial lesions of striato- and pallido-nigral GABA neurons with kainic acid (1.2 micrograms) decrease by 40% the glutamic acid decarboxylase (GAD) activity in the ipsilateral SN without decreasing the spontaneous release of [3H]GABA; even following extensive lesions with kainic acid (2.5 micrograms), GAD activity (-72%) and spontaneous [3H]GABA release (-83%) were not completely abolished. These results suggest that a non-negligible contribution of GABA nigral neurons accounts for the spontaneous GABA release measured in the substantia nigra. This is further supported by the decrease (-20%), and the increase (+40%) of [3H]GABA release produced by the local application of glycine (10(-6) M), and bicuculline (10(-4) M), which respectively, inhibits and activates the nigral neuron activity. The contribution of nigral GABA neurons to the amount of [3H]GABA release from the substantia nigra, is likely linked to their high spontaneous firing rate.     

Release of exogenous glycine in the pigeon optic tectum during stimulation of a midbrain nucleus

A pathway having an affinity for glycine has been investigated in the pigeon optic lobe; it originates in the nucleus isthmi pars parvocellularis (Ipc) and terminates in the tectum. In an attempt to obtain evidence that glycine plays a role as a transmitter in this system, the effect of electrical stimulation on release of labeled substances previously injected in the tectum was tested. By perfusing the upper strata of the optic tectum with a push-pull cannula the release of radioactive glycine was shown to be markedly increased by electrical stimulation of Ipc, but not by stimulation of other sites. Ipc stimulation did not affect the efflux of exogenous leucine or urea, whereas a GABA release was observed. With K⁺ (40 mM) stimulation all amino acids tested were released. It is suggested that the Ipc neuron terminals in the tectum take up glycine and release it upon stimulation of the Ipc nucleus.     

Glutamate release induced by activation of glycine and GABA transporters in spinal cord is enhanced in a mouse model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis is a progressive and fatal neurodegenerative disease, involving both upper and lower motor neurons, the cause of which is obscure, although glutamate (GLU)-induced excitotoxicity has been suggested to play a major role. We studied the release of [3H]d-aspartate ([3H]d-ASP) and endogenous glutamate evoked by glycine (GLY) or GABA from spinal cord synaptosomes in mice expressing a mutant form of human SOD1 with a Gly93Ala substitution ([SOD1-G93A(+)]), a transgenic model of amyotrophic lateral sclerosis, in mice expressing the non-mutated form of human SOD1 [SOD1+], and in non-transgenic littermates [SOD1(-)/G93A(-)]. In parallel experiments, we also studied the release of [3H]GABA evoked by GLY and that of [3H]GLY evoked by GABA. Mutant mice were killed at advanced phase of pathology or during the pre-symptomatic period. In SOD1(-)/G93A(-) or SOD1(+) mice GLY evoked [3H]d-ASP and [3H]GABA release, while GABA caused [3H]d-ASP, but not [3H]GLY, release. The GLY-evoked release of [3H]d-ASP, but not that of [3H]GABA, and the GABA-evoked [3H]d-ASP release, but not that of [3H]GLY, were more pronounced in SOD1-G93A(+) than in SOD1(+) or SOD1(-)/G93A(-) mice. Furthermore, the excessive potentiation of [3H]d-ASP by GLY or GABA was already present in asymptomatic 30-40 day-old SOD1-G93A(+) mice. The releases of endogenous glutamate and GABA also were enhanced by GLY and the GLY-evoked release of endogenous glutamate, but not of endogenous GABA, was higher in SOD1-G93A(+) than in control animals. Potentiation of the spontaneous amino acid release is likely to be mediated by activation of a GLY or a GABA transporter, since the effect of GLY was counteracted by the GLY transporter blocker glycyldodecylamide but not by the GLY receptor antagonists strychnine and 5,7-dichlorokynurenate while the effect of GABA was diminished by the GABA transporter blocker SKF89976-A but not by the GABA receptor antagonists SR9531 and CGP52432. It is concluded that the glutamate release machinery seems excessively functional in SOD1-G93A(+) animals.     

Quantitative autoradiographic analysis of the distribution of [3H]muscimol binding to GABA receptors in chick brain

Quantitative receptor autoradiography was used to investigate the distribution of high-affinity GABA receptors (GABAA) in left and right hemispheres of the brains of 3-week-old chicks. The receptors were labelled with the potent GABA agonist [3H]muscimol. High levels of [3H]muscimol labelling were found throughout the fore-, mid-, and hindbrain, though considerable variation was found in different regions. In the telencephalon the highest concentration of specific binding was found in the hyperstriatum ventrale followed by the neostriatum, and then the lobus parolfactorius of the paleostriatal complex, whilst in the diencephalon highest levels of labelling were present in the infundibulum. In the midbrain distinct lamination was observed in the high levels of [3H]muscimol binding in the optic tectum and in the hind brain the highest density of labelling occurred in the granular layers of the cerebellum. Levels of labelling were generally low in the brainstem regions. The distribution of [3H]muscimol binding in the optic tectum and in the hind brain the highest density of labelling occurred in the granular layers of the cerebellum. Levels of labelling were generally low in the brainstem regions. The distribution of [3H]muscimol binding sites is in good agreement with our previous work on the distribution of GABA-immunoreactivity in the chick brain.     

Biochemical correlates of GABA function in rat cortical neurons in culture

Serial biochemical studies of a rat cortical tissue culture system in which synapses regularly form showed that γ-aminobutyric acid (GABA) is present in the cultures and increases with their maturation. The tissue GABA concentration in mature cultures is similar to that of adult rat cortex in vivo. The synthetic enzyme, glutamate decar☐ylase, also increases with age as does high affinity GABA uptake. GABA uptake was blocked by l-2,4-diaminobutyrate (DABA) and had the properties of neuronal GABA uptake. Specific release by depolarizing media of both exogenous [³H]GABA and GABA synthesized from d-[U-¹⁴C]glucose was demonstrated. The GABA released by high potassium media had higher specific activity and a greater contribution from glucose (as compared to acetate) than GABA found in the medium in the absence of depolarization. Calcium dependency of evoked GABA release could be shown only after pretreatment of cultures with ethyleneglycol-bis-(β-aminoethyl ether)-N,N′-tetraacetic acid or EGTA. Synaptosomes may exhibit greater calcium dependence of evoked transmitter release than intact cells in culture because their intracellular calcium stores are depleted during preparation. Glycine uptake by the cultures was much less in amount than was GABA uptake, and specific release of glycine could not be demonstrated.Specific binding of both a GABA agonist ([³H]muscimol) and an antagonist ([³H]bicuculline) was shown by membranes prepared from the cultures. By contrast, when [³H]muscimol binding to intact cells was studied, essentially all binding was sodium dependent and had the properties of GABA uptake binding. We conclude that the use of [³H]muscimol for receptor studies is valid only after the elimination of GABA uptake systems.Biochemical data from these studies support the concept that GABA is the transmitter for many cortical synapses. Glycine and taurine are not likely to be transmitters in these cortical cultures. When considered together with physiological data from the preceding paper, we have satisfied Werman's criteria (see ref. 36) for accepting GABA as the major inhibitory transmitter in the cortical culture system.     

Biochemical and morphological studies on GABA neurons in reaggregate culture

Dissociated cells from the 14-day fetal mouse corpus striatum (CS), rostral mesencephalic tegmentum (RMT) and tectum were reaggregated in rotation-mediated cultures in the following combinations: CS alone, RMT-CS, RMT-tectum, and tectum alone. Reaggregates were cultured for 1-4 weeks. An even distribution of gamma-aminobutyric acid (GABA)-positive cells was observed in all the reaggregate combinations that were cultured for 1 and 2 weeks. With increasing time in culture, cellular staining was decreased while positive fiber staining increased. CS and RMT-CS co-cultures increased their capacity to take up [3H]GABA with time in culture. All reaggregates reached a maximum uptake/accumulation capacity of 30-40 pmol/mg protein/30 min, by 4 weeks in vitro. There were no significant differences between the various co-aggregate combinations in the accumulation capacity. [3H]GABA accumulation in the reaggregates was largely blocked by the putative neuronal GABA uptake inhibitor, diaminobutyric acid, and was inhibited to a much lesser degree by the putative glial uptake inhibitor beta-alanine. All reaggregates released [3H]GABA to 70 mM potassium depolarization, in a calcium-dependent manner. One-three week CS reaggregates released more [3H]GABA in response to the potassium-induced depolarization than RMT-CS co-cultures. Since nigral dopamine neurons within the RMT proliferate processes and actively release dopamine only when co-cultured with CS target cells, it is suggested that these dopamine neurons might chronically inhibit striatal GABA neurons in the RMT-CS co-cultures, thereby depressing stimulated release of [3H]GABA.     

Effect of pentobarbitone on the spontaneous efflux of γ-amino acids from rabbit retina

The effect of pentobarbitone on the spontaneous release of radioactivity from rabbit retinas preloaded with [³H]GABA, [³H]DABA, [³H]β-alanine and [³H]glycine was studied. In high concentration of pentobarbitone (10⁻³ M) the rate of spontaneous efflux of neuronal [³H]GABA, [³H]DABA and [³H]β-alanine was reduced but not that of [³H]glycine. Glial release of [³H]GABA was much less influenced. At lower concentrations (10⁻⁴-10⁻⁵ M) there was an initial increase in the release of [³H]GABA, [³H]DABA and [³H]β-alanine followed by a decrease, indicating several components in the release systems for the amino acids.     

Descending fibre-mediated release of endogenous glutamate and glycine from the perfused cat spinal cord in vivo

The release of endogenous amino acids into the perfused central canal of the cat spinal cord in vivo was studied using a [3H]dansyl chloride assay procedure. Perfusion of 0.1 mM p-chloromercuriphenyl-sulphonate (pCMS), an uptake inhibitor, augmented the efflux of most amino acids studied. The large increases in glycine, glutamate and GABA efflux (9.9, 5.3 and 4.1-fold, respectively) suggest that the membrane fluxes of these amino acids were particularly high, a finding consistent with their putative neurotransmitter functions. Subsequent electrical stimulation of descending spinal tracts increased the efflux of glycine, glutamate, GABA and leucine (1.8, 2.2, 1.9 and 1.3-fold, respectively), although only the release of glycine and glutamate was clearly independent of the concomitant rise in blood pressure. The evoked amino acid release, which was accompanied by a substantial ventral root discharge, was not observed in the absence of pCMS, nor in the presence of a low concentration (0.01 mM) of this compound. These results have been discussed in relation to the putative neurotransmitter roles for amine acids in the spinal cord.     

The uptake and radioautographical localization in the frog retina of [H](±)-aminocyclohexane carboxylic acid, a selective inhibitor of neuronal gaba transport

The accumulation of [3H](+/-)-cis-3-aminocyclohexane carboxylic acid ([3H] ACHC) in frog retinae in vitro was highly localized in horizontal cells and their processes. [3H]GABA was also mainly accumulated within horizontal cells, but [3H]L-2,4-diaminobutyric acid ([3H]DABA) was taken up predominantly by the neuroglial Müller cells, whilst [3H]beta-alanine was localised largely within the photoreceptors. The uptake of [3H]ACHC (4.2 microM) was almost linear for 30 min and after 60 min a tissue/medium ratio of 5.25 was achieved. The uptake process was temperature sensitive highly dependent on sodium ions, and appeared to be mediated by a saturable transport process with an IC50 value of 0.83 mM. The accumulation of [3H]ACHC was inhibited by GABA and DABA (IC50 = 0.32 mM and 0.23 mM, respectively) whilst beta-alanine was a relatively weak inhibitor (IC50 = 9 mM) of ACHC uptake. In agreement with these results, the efflux of [3H]ACHC from the retina was increased by exposure to ACHC, GABA and DABA but not beta-alanine. In contrast, the efflux of [3H]DABA from the retina was not increased by GABA or ACHC, although DABA itself and potassium depolarization stimulated the release of [3H]DABA. These results strongly suggest that ACHC is accumulated in the frog retina by the same neuronal transport process as GABA. In contrast, the high affinity sites for DABA are localized mainly in glia, although inhibitor and release studies suggest that, at high concentration, DABA also interacts with the neuronal GABA (ACHC) transport process.     

Comparison of binding affinities and capacities of gamma-aminobutyric acid and glycine in synaptosome-enriched fractions of rat cerebral cortex and spinal cord.

The “binding” affinities and capacities of [³H]-γ-aminobutyric acid (³H-GABA) and [¹⁴C]-glycine to synaptosome-enriched fractions of rat cerebral cortex and spinal cord have been compared over a 10^?8 to 10^?3m concentration range in isosmotic sucrose solutions containing 32 mm NaCl. At 0 C, the binding of both GABA and glycine to particles of the cerebral cortex appeared to take place by “triple-affinity” processes. The respective binding affinities (reciprocals of K_B values) of GABA for particles of the cortex were generally greater than those for glycine, whereas maximal binding capacities (B_max values) for GABA were about twice as great as those for glycine. In fractions of spinal cord, at 0 C, the binding of both GABA and glycine occurred by “double-affinity” processes, and though the binding affinities of GABA were about twice those of glycine, B_max values for GABA were smaller than those for glycine. Striking changes occurred when particles were allowed to stand in contact with [³H]-GABA plus [¹⁴C]-glycine at 23 C for 45 min. Although double-affinity mechanisms persisted at the higher temperature for the binding of both amino acids to particles of spinal cord (GABA still being bound with greater affinities than glycine), the binding of both amino acids to particles of cerebral cortex was best resolved as “single-affinity” processes. The differences between B_max values for GABA and glycine in particles of cerebral cortex and spinal cord appeared to reflect their different endogenous concentrations.     

GABAA and GABAB sites in bovine adrenal medulla membranes

The effect of several ligands and Ca2+ ions on [3H]GABA binding to bovine adrenal medulla membranes was investigated. Without any blockade, the [3H]GABA binding showed two components, one of low affinity (Kd = 139 +/- 22 nM and Bmax = 3.2 +/- 0.4 pmol/mg protein) and the other of high affinity (Kd = 41 +/- 6 nM and Bmax = 0.35 +/- 0.26 pmol/mg protein). Muscimol specifically blocked low-affinity sites, and (-)baclofen blocked high-affinity components. Ca2+ ions were strictly necessary for maximum binding to high-affinity sites, whereas they did not significantly affect sites of the lower affinity. These results show that the bovine adrenal medulla has a GABAA receptor population of low affinity together with a GABAB receptor of high affinity.     

Stimulated release of endogenous GABA and glycine from the goldfish retina

The release of endogenous gamma-aminobutyric acid (GABA) and glycine from the isolated goldfish retina, measured by high-pressure liquid chromatography (HPLC), was Ca2+-independent when evoked by L-glutamate or L-aspartate and partially Ca2+-dependent when evoked by 50 mM K+. D-Aspartate potentiated GABA and glycine release evoked by L-glutamate and inhibited that evoked by L-aspartate. These data are similar to those reported for radiolabeled GABA and glycine. However, the relative amount released compared to the total amino acid content in the retina was much less (10%) for the endogenous compounds. We suggest that results obtained with [3H]GABA and [3H]glycine can be generalized in a qualitative manner to their endogenous counterparts in goldfish retina.