RESUMO
Complex dynamic cellular networks have been studied in physiological and pathological processes under the light of single-cell calcium imaging (SCCI), a method that correlates functional data based on calcium shifts operated by different intracellular and extracellular mechanisms integrated with their cell phenotypes. From the classic synaptic structure to tripartite astrocytic model or the recent quadripartite microglia added ensemble, as well as other physiological tissues, it is possible to follow how cells signal spatiotemporally to cellular patterns. This methodology has been used broadly due to the universal properties of calcium as a second messenger. In general, at least two types of receptor operate through calcium permeation: a fast-acting ionotropic receptor channel and a slow-activating metabotropic receptor, added to exchangers/transporters/pumps and intracellular Ca2+ release activated by messengers. These prototypes have gained an enormous amount of information in dynamic signaling circuits. SCCI has also been used as a method to associate phenotypic markers during development and stage transitions in progenitors, stem, vascular cells, neuro- and glioblasts, neurons, astrocytes, oligodendrocytes, and microglia that operate through ion channels, transporters, and receptors. Also, cancer cells or inducible cell lines from human organoids characterized by transition stages are currently being used to model diseases or reconfigure healthy cells in terms of the expression of calcium-binding/permeable molecules and shed light on therapy.
RESUMO
Development of progenitors in the embryonic retina is modulated by signaling molecules, and cannabinoid receptors are highly expressed in the early developing retina. Here, we investigated whether the CB1/CB2 receptor agonist WIN 5212-2 (WIN) modulated the proliferation, viability, and calcium responses in chick embryo retinal progenitors in culture. A decline in [3H]-thymidine incorporation was observed when cultures were incubated with 0.5-1.0 µM WIN, an effect that was mimicked by URB602 and URB597, inhibitors of the monoacylglycerol lipase and fatty acid amide hydrolase, respectively. A reduction in the number of proliferating cell nuclear antigen-positive nuclei was also noticed in WIN-treated cultures, suggesting that activation of cannabinoid receptors decreases the proliferation of cultured retinal progenitors. WIN (0.5-5.0 µM), but not capsaicin, decreased retinal cell viability, an effect that was blocked by CB1 and CB2 receptor antagonists and by the P2X7 receptor antagonist A438079, implicating this nucleotide receptor in the cannabinoid-mediated cell death. Treatment with WIN also induced an increase in mitochondrial superoxide and P2X7 receptor-mediated uptake of sulforhodamine B in the cultured cells. While a high proportion of cultured cells responded to glutamate, GABA, and 50 mM KCl with intracellular calcium shifts, very few cells responded to the activation of P2X7 receptors by ATP. Noteworthy, while decreasing the number of cells responding to glutamate, GABA, and KCl, treatment of the cultures with WIN induced a significant increase in the number of cells responding to 1 mM ATP, suggesting that activation of cannabinoid receptors primes P2X7 receptor calcium signaling in retinal progenitors in culture.
Assuntos
Apoptose/efeitos dos fármacos , Canabinoides/farmacologia , Neuroglia/citologia , Receptores Purinérgicos P2X7/metabolismo , Retina/citologia , Transdução de Sinais/efeitos dos fármacos , Células-Tronco/metabolismo , Animais , Benzoxazinas/farmacologia , Cálcio/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Embrião de Galinha , Corantes Fluorescentes/metabolismo , Morfolinas/farmacologia , Naftalenos/farmacologia , Nestina/metabolismo , Fenótipo , Receptor CB1 de Canabinoide/metabolismo , Receptor CB2 de Canabinoide/metabolismo , Células-Tronco/efeitos dos fármacosRESUMO
Neurospheres prepared from multipotent progenitors in the retina obtained from postnatal mice differentiate into neurons and Müller glia (De Melo Reis et al., in Cell Mol Neurobiol 31:835-846, 2011). Here, we investigated whether neurospheres prepared from adult chickens (ciliary marginal zone, CMZ) or (ciliary body) retina could also lead to differentiated neurons and glia. Neurospheres were prepared from post-hatched chickens or from adult mice after 7 days in the presence of mitogenic factors (FGFb, insulin, and EGF), generating neurons and glial cells. In addition, Müller (2M6 or glutamine synthetase positive cells) derived from post-hatch chicken CMZ neurospheres displayed the dopaminergic phenotype. Furthermore, we observed that Müller cells derived from adult chickens and mice retina neurospheres released significant amounts of dopamine as well as of its metabolites. Taken together, our data lead us to conclude that as for embryonic (chick) or newborn (mouse), the dopaminergic phenotype is a default condition of Müller glial cells obtained from neurospheres prepared from mature retina. Our data raise the possibility that Müller cells from differentiated tissue could be used to ameliorate neurodegenerative diseases involving dopaminergic dysfunction as in Parkinson's disease as shown previously (Stutz et al., in J Neurochem 128:829-840, 2014).
Assuntos
Envelhecimento/metabolismo , Dopamina/metabolismo , Células Ependimogliais/citologia , Retina/citologia , Esferoides Celulares/citologia , Animais , Animais Recém-Nascidos , Separação Celular , Células Cultivadas , Galinhas , Células Ependimogliais/metabolismo , Metaboloma , Camundongos Endogâmicos C57BL , Fenótipo , Esferoides Celulares/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
BACKGROUND: The endosymbiosis in trypanosomatids is characterized by co-evolution between one bacterium and its host protozoan in a mutualistic relationship, thus constituting an excellent model to study organelle origin in the eukaryotic cell. In this association, an intense metabolic exchange is observed between both partners: the host provides energetic molecules and a stable environment to a reduced wall symbiont, while the bacterium is able to interfere in host metabolism by enhancing phospholipid production and completing essential biosynthesis pathways, such as amino acids and hemin production. The bacterium envelope presents a reduced cell wall which is mainly composed of cardiolipin and phosphatidylcholine, being the latter only common in intracellular prokaryotes. Phosphatidylinositol (PI) is also present in the symbiont and host cell membranes. This phospholipid is usually related to cellular signaling and to anchor surface molecules, which represents important events for cellular interactions. METHODS: In order to investigate the production of PI and its derivatives in symbiont bearing trypanosomatids, aposymbiotic and wild type strains of Angomonas deanei, as well as isolated symbionts, were incubated with [(3)H]myo-inositol and the incorporation of this tracer was analyzed into inositol-containing molecules, mainly phosphoinositides and lipoproteins. Gene searches and their phylogenies were also performed in order to investigate the PI synthesis in symbiontbearing trypanosomatids. RESULTS: Our results showed that the bacterium did not incorporate the tracer and that both strains produced similar quantities of PI and its derivatives, indicating that the symbiont does not influence the production of these metabolites. Gene searches related to PI synthesis revealed that the trypanosomatid genome contains an inositol transporter, PI synthase and the myo-inositol synthase. Thus, the host is able to produce PI either from exogenous myo-inositol (inositol transporter) or from myo-inositol synthesized de novo. Phylogenetic analysis using other organisms as references indicated that, in trypanosomatids, the genes involved in PI synthesis have a monophyletic origin. In accordance with experimental data, sequences for myo-inositol transport or for myo-inositol and PI biosynthesis were not found in the symbiont. CONCLUSIONS: Altogether, our results indicate that the bacterium depends on the host to obtain PI.
Assuntos
Fosfatidilinositóis/metabolismo , Trypanosomatina/metabolismo , Bactérias/isolamento & purificação , Regulação da Expressão Gênica/fisiologia , Filogenia , Simbiose , Trypanosomatina/genéticaRESUMO
Dopamine is the main catecholamine found in the retina of most species, being synthesized from the L-amino acid tyrosine. Its effects are mediated by G protein coupled receptors subfamilies that are commonly coupled to adenylyl cyclase in opposite manners. There is evidence that this amine works as a developmental signal in the embryonic retina and several distinct roles have been attributed to dopamine in the retina such as proliferation, synaptogenesis, neuroprotection, increased signal transmission in cone, gap junction modulation, neuronal-pigmented epithelium-glial communication, and neuron-glia interaction. Here we describe methods that have been used in the study of the dopaminergic function in the retina in the last 40 years. We emphasize the approaches used in the studies on the development of the avian and rodent retina. The dopaminergic system is one of the first phenotypes to appear in the developing vertebrate retina.
Assuntos
Dopamina/metabolismo , Retina/citologia , Retina/metabolismo , Animais , Benzazepinas/farmacologia , Biomarcadores/metabolismo , Western Blotting , Técnicas de Cultura de Células , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Galinhas , AMP Cíclico/metabolismo , Imuno-Histoquímica , Neuroglia/citologia , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Dopaminérgicos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sinapses/metabolismo , Técnicas de Cultura de TecidosRESUMO
The immunocytochemical staining of L-DOPA decarboxylase (DDC) and tyrosine hydroxylase (TH) in cells of the developing avian retina and in cells of the retina of adult rats and opossum were compared. DDC was identified at embryonic day 8 in the chick, in cells in the inner nuclear layer (INL). At embryonic day 13, two types of DDC positive cells were observed; type 1, with the soma located in the innermost layer of the INL; and type 2, with the soma located two cell rows from the innermost part of the INL. Immunolabeling for DDC in the presumptive outer plexiform layer was more clearly defined at embryonic day 19 and at post-hatched day 7. Processes of DDC labeled cells extended into the inner plexiform layer, supporting the amacrine identity of these cells. Dot-blot analysis revealed that DDC could be detected at embryonic day 4. Confocal microscopy showed that at embryonic day 10, DDC positive cells, but not TH positive cells, were found. After embryonic day 13, cells immunolabeled for DDC and DDC plus TH were detected. The mean density of DDC positive cells quantified in whole-mounted chick retinas showed that in all stages the density of DDC positive cells exceeded that of TH positive cells by 10-13-fold. As for the avian retina, density of DDC positive cells in opossum and rat retinas exceeded the density of TH positive cells. In opossum, Müller fibers were also clearly labeled for DDC but not for TH. We propose the hypothesis that the dopamine synthesis in the developing avian retina as well as in the mature rat and opossum tissue is greater than would be expected from TH staining alone.
Assuntos
Dopa Descarboxilase/metabolismo , Retina/enzimologia , Retina/crescimento & desenvolvimento , Tirosina 3-Mono-Oxigenase/metabolismo , Animais , Embrião de Galinha , Didelphis , Feminino , Immunoblotting , Imuno-Histoquímica , Masculino , Microscopia Confocal , RatosRESUMO
Müller cells represent the main type of glia present in the retina interacting with most, if not all neurons in this tissue. Müller cells have been claimed to function as optic fibers in the retina delivering light to photoreceptors with minimal distortion and low loss [Franze et al (2007) Proc Natl Acad Sci 104:8287-8292]. Most of the mediators found in the brain are also detected in the retinal tissue, and glia cells are active players in the synthesis, release, signaling and uptake of major mediators of synaptic function. Müller glia trophic factors may regulate many different aspects of neuronal circuitry during synaptogenesis, differentiation, neuroprotection and survival of photoreceptors, Retinal Ganglion Cells (RGCs) and other targets in the retina. Here we review the role of several transmitters and trophic factors that participate in the neuron-glia loop in the retina.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neuroglia/metabolismo , Neurotransmissores/metabolismo , Retina/citologia , Animais , Dopamina/metabolismo , Ácido Glutâmico/metabolismo , Neuroglia/citologia , Neurônios/citologia , Neurônios/metabolismo , Purinas/metabolismo , Retina/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
GABA is the main inhibitory aminoacid transmitter present in neurons and glial cells. Its uptake is carried out by specific high-affinity Na(+)/Cl (-) dependent transporters (GATs). It has been reported in the past that, in the avian retina, [(3)H]GABA appears to be exclusively accumulated by horizontal and amacrine cells in the inner nuclear layer, and also by ganglion cells. Purified chick Müller glia cultures were able to take up [(3)H]GABA in a Na(+) and Cl(+) dependent way. Increasing GABA concentration increases GABA uptake by these cells, reaching half-maximal transport efficiency (EC50) around 0.3 mM. [(3)H]GABA uptake by Müller glia neuronal-free cultures was not mediated by neuronal transporters since it was not blocked by NNC-711, but was inhibited by beta-alanine, a specific glial transporter inhibitor. Chick Müller glia in culture express both GAT-1 and GAT-3 GABA transporters. Although mixed neuron-glial dense cultures released GABA upon glutamate, high K[(+) or veratridine stimulation, Müller glial cells did not release [(3)H]GABA upon treatment with these agents, suggesting that different from neurons, transporter mediated GABA release is not a common mechanism operating in these cells. The data also suggest that Müller cells take up GABA unidirectionally, which may constitute an important mechanism of inactivating GABA activity mediated by neurons.
Assuntos
Neuroglia/fisiologia , Retina/citologia , Ácido gama-Aminobutírico/metabolismo , Análise de Variância , Animais , Transporte Biológico/efeitos dos fármacos , Células Cultivadas , Embrião de Galinha , Cloretos/metabolismo , Técnicas de Cocultura/métodos , Relação Dose-Resposta a Droga , Antagonistas GABAérgicos/farmacologia , Neuroglia/efeitos dos fármacos , Neurônios/química , Ácidos Nipecóticos/farmacologia , Oximas/farmacologia , Sódio/metabolismo , Temperatura , Trítio/metabolismoRESUMO
Dopamine is the main catecholamine found in the chick retina whereas norepinephrine is only found in trace amounts. We compared the effectiveness of dopamine and norepinephrine in promoting cyclic AMP accumulation in retinas at embryonic day 13 (E13) and from post-hatched chicken (P15). Dopamine (EC(50)=10microM) and norepinephrine (EC(50)=30microM), but not the beta(1)-adrenergic agonist isoproterenol, stimulated over seven-fold the production of cyclic AMP in E13 retina. The cyclic AMP accumulation induced by both catecholamines in embryonic tissue was entirely blocked by 2microM SCH23390, a D(1) receptor antagonist, but not by alprenolol (beta-adrenoceptor antagonist). In P15 retinas, 100microM isoproterenol stimulated five-fold the accumulation of cAMP. This effect was blocked by propanolol (10microM), but not by 2microM SCH23390. Embryonic and adult retina display beta(1) adrenergic receptor mRNA as detected by RT-PCR, but the beta(1) adrenergic receptor protein was detected only in post-hatched tissue. We conclude that norepinephrine cross-reacts with D(1) dopaminergic receptor with affinity similar to that of dopamine in the embryonic retina. In the mature retina, however, D(1) receptors become restricted to activation by dopamine. Moreover, as opposed to the embryonic tissue, norepinephrine seems to stimulate cAMP accumulation via beta(1)-like adrenergic receptors in the mature tissue.
Assuntos
Agonistas de Dopamina/farmacologia , Norepinefrina/farmacologia , Receptores Adrenérgicos beta 1/metabolismo , Receptores de Dopamina D1/agonistas , Retina/efeitos dos fármacos , Animais , Benzazepinas/metabolismo , Células Cultivadas , Embrião de Galinha , Ensaio Radioligante , Retina/citologia , Retina/embriologiaRESUMO
Some trypanosomatids, such as Crithidia deanei, are endosymbiont-containing species. Aposymbiotic strains are obtained after antibiotic treatment, revealing interesting aspects of this symbiotic association. Ornithine decarboxylase (ODC) promotes polyamine biosynthesis and contributes to cell proliferation. Here, we show that ODC activity is higher in endosymbiont-bearing trypanosomatids than in aposymbiotic cells, but isolated endosymbionts did not display this enzyme activity. Intriguingly, expressed levels of ODC were similar in both strains, suggesting that ODC is positively modulated in endosymbiont-bearing cells. When the aposymbiotic strain was grown in conditioned medium, obtained after cultivation of the endosymbiont-bearing strain, cellular proliferation as well as ODC activity and localization were similar to that observed in the endosymbiont-containing trypanosomatids. Furthermore, dialyzed-heated medium and trypsin treatment reduced ODC activity of the aposymbiont strain. Taken together, these data indicate that the endosymbiont can enhance the protozoan ODC activity by providing factors of protein nature, which increase the host polyamine metabolism.
Assuntos
Fenômenos Fisiológicos Bacterianos , Proteínas de Bactérias/metabolismo , Crithidia/enzimologia , Crithidia/microbiologia , Ornitina Descarboxilase/metabolismo , Simbiose/fisiologia , Animais , Crithidia/classificação , Ativação Enzimática , Especificidade da EspécieRESUMO
In the chick retina, dopaminergic cells are generated between embryonic days 3 and 7 (E3/E7). However, the expression of tyrosine hydroxylase (TH), the first enzyme in the catecholamine synthetic pathway, is only detected after E11/E12. During the interval comprising E7 to E12, signals conveyed by cAMP are important to determine the TH phenotype. The present study shows that pituitary adenylyl cyclase-activating polypeptide (PACAP), via cAMP, is a major endogenous component in defining the TH phenotype of retina dopaminergic cells during development. PACAP type 1 receptor and its mRNA were detected in retinas since E6. PACAP was also immunodetected in cells localized in the inner nuclear layer of retinas since E8. This peptide promoted greater than 10-fold increase in cAMP accumulation of retinas obtained from embryos since E8, an effect that was blocked by PACAP6-38 (PAC1 receptor antagonist). In cultured retina cells from E8 and E9, maintained for 6 days in vitro with 10 nM PACAP (for 5 days), the number of dopaminergic cells expressing tyrosine hydroxylase increased 2.4-fold. The cAMP analog, 8-Br-cAMP and 3-isobutyl-1-methylxanthine (IBMX, a phosphodiesterase inhibitor) also increased the number of tyrosine hydroxylase-positive cells by 4- to 6-fold. IBMX plus PACAP treatment resulted in 17-fold increase in the number of cells positive for tyrosine hydroxylase. Under this condition the amount of tyrosine hydroxylase expression, as detected by western blot analysis, was also increased. The protein kinase-A inhibitor, rp-cAMPS, significantly reduced the effect of PACAP. Our data show that this peptide is an important factor influencing the definition of the tyrosine hydroxylase phenotype of retina dopaminergic cells within a narrow window of development.
Assuntos
Dopamina/metabolismo , Fatores de Crescimento Neural/fisiologia , Neurônios/metabolismo , Neuropeptídeos/fisiologia , Neurotransmissores/fisiologia , Retina , Tirosina 3-Mono-Oxigenase/metabolismo , 1-Metil-3-Isobutilxantina/farmacologia , 8-Bromo Monofosfato de Adenosina Cíclica/farmacologia , Fatores Etários , Animais , Animais Recém-Nascidos , Western Blotting/métodos , Contagem de Células/métodos , Técnicas de Cultura de Células , Embrião de Galinha , Colforsina/farmacologia , AMP Cíclico/metabolismo , Relação Dose-Resposta a Droga , Interações Medicamentosas , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Imuno-Histoquímica/métodos , Microscopia Confocal/métodos , Fatores de Crescimento Neural/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Neuropeptídeos/antagonistas & inibidores , Neurotransmissores/antagonistas & inibidores , Fenótipo , Inibidores de Fosfodiesterase/farmacologia , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , RNA Mensageiro/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores de Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Receptores de Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Retina/citologia , Retina/embriologia , Retina/enzimologia , Retina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Fatores de Tempo , Tirosina 3-Mono-Oxigenase/genéticaRESUMO
Glia represents the most numerous group of nervous system cells and CNS development and function depend on glial cells. We developed a purified Muller glia culture to investigate the expression of several neurotransmitter markers on these cells, such as dopaminergic, cholinergic, GABAergic and peptidergic receptors or enzymes, based on functional assays measuring second messenger levels or Western blot for specific proteins. Purified Muller cell culture was obtained from 8-day-old (E8) embryonic chick. Glial cells cultured for 15 days (E8C15) expressed D1A and D1B receptors mRNAs, but not D1D, as detected by RT-PCR. The binding of [3H]-SCH 23390 revealed an amount of expressed receptors around 40 fmol/mg protein. Dopamine (100 microM), PACAP (50 nM) and forskolin (10 microM) induced a 50-, 30- and 40-fold cAMP accumulation on glial cells, respectively, but not ip3 production. The dopamine-promoted cAMP accumulation was blocked by 2 microM SCH 23390. Carbachol stimulated a 3-fold ip3 accumulation. Western blot analysis also revealed the expression of tyrosine hydroxylase, L-dopa decarboxylase, PAC1 receptor, GAD67 and beta2-nicotinic receptor subunit by these cells. These results indicate that several components of neurotransmitter signaling and metabolism are found in cultured Muller cells.
Assuntos
Neuroglia/metabolismo , Neurotransmissores/metabolismo , Receptores de Neurotransmissores/metabolismo , Animais , Benzazepinas/metabolismo , Biomarcadores , Western Blotting , Células Cultivadas , Embrião de Galinha , Colforsina/farmacologia , AMP Cíclico/metabolismo , Dopamina/metabolismo , Dopamina/farmacologia , Antagonistas de Dopamina/metabolismo , Inositol 1,4,5-Trifosfato/metabolismo , Fatores de Crescimento Neural/farmacologia , Neuroglia/enzimologia , Neurônios/metabolismo , Neuropeptídeos/farmacologia , Neurotransmissores/farmacologia , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Racloprida/metabolismo , Receptores de Dopamina D1/genética , Receptores de Dopamina D1/metabolismo , Retina/citologia , Retina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Cytotoxic-activated macrophages control Toxoplasma gondii growth by producing nitric oxide (NO). However, the parasite can partially inhibit NO production. NO is generated from arginine within the polyamine biosynthetic pathway. Two enzymes of this pathway are ornithine, decarboxylase (ODC) and arginine decarboxylase (ADC). The aim of the present work was to investigate whether T. gondii is able to modulate polyamine metabolism in macrophages. Toxoplasma gondii infection did not affect basal ODC or ADC activity. However, lipopolysaccharide induced an increase in ODC activity. Polyamine-treated macrophages exhibited a T. gondii-infection index similar to controls but a higher adhesion index; the parasite did not grow in methyl-ornithine (ODC inhibitor)-treated macrophages. The parasites were able to take up putrescine with a Km of 0.92 microM, indicating the presence of a high-affinity putrescine-transporter system. Putrescine-treated T. gondii actively penetrated macrophages and Vero cells. However, NO production and lysosomal parasitophorous vacuole fusion were not inhibited. Considered together, these results demonstrate that T. gondii requires polyamines for multiplication. However, as opposed to Trypanosoma cruzi and because of a relatively high-affinity putrescine-transporter system in the parasite, constitutive macrophage levels of putrescine seem sufficient to support T. gondii survival and multiplication.
Assuntos
Macrófagos Peritoneais/parasitologia , Poliaminas/metabolismo , Toxoplasma/crescimento & desenvolvimento , Animais , Carboxiliases/metabolismo , Células Cultivadas , Chlorocebus aethiops , Interações Hospedeiro-Parasita , Macrófagos Peritoneais/enzimologia , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos , Óxido Nítrico/biossíntese , Ornitina/metabolismo , Ornitina Descarboxilase/metabolismo , Putrescina/metabolismo , Espermidina/metabolismo , Células VeroRESUMO
Glutamate and GABA are the major excitatory and inhibitory neurotransmitters in the CNS. In the retina, it has been shown that glutamate and aspartate and their agonists kainate and NMDA promote the release of GABA. In the chick retina, at embryonic day 14 (E14), glutamate and kainate were able to induce the release of GABA from amacrine and horizontal cells as detected by GABA-immunoreactivity. NMDA also induced GABA release restricted to amacrine cell population and its projections to the inner plexiform layer (E14 and E18). Although aspartate reduced GABA immunoreactivity, specifically in amacrine cells of E18 retinas, it was not efficient to promote GABA release from retinas at E14. As observed in differentiated retinas, dopamine inhibited the GABA release promoted by NMDA and aspartate but not by kainate. Our data show that different retinal sites respond to distinct EAAs via different receptor systems.