Navegando por Assunto "Neurotransmissores"

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Acesso aberto (Open Access)
GABA and glutamate transporters: new events and function in the vertebrate retina
(2013) NASCIMENTO, José Luiz Martins do; SAWADA, Luis Armando; OLIVEIRA, Karen Renata Matos; CRESPO LÓPEZ, Maria Elena; SILVA, Anderson Manoel Herculano Oliveira da; HAMOY, Moisés; SILVA, Consuelo Yumiko Yoshioka e; BASTOS, Gilmara de Nazareth Tavares; PANTOJA, Wendell Mauro Soeiro
The neural retina is a highly complex tissue composed of excitatory and inhibitory neurons and glial cells. Glutamate, the main excitatory neurotransmitter, mediates information transfer from photoreceptors, bipolar cells, and ganglion cells, whereas interneurons, mainly amacrine and horizontal cells, use γ-aminobutyric acid (GABA), the main inhibitory neurotransmitter. In this review we place an emphasis on glutamate and GABA transporters as highly regulated molecules that play fundamental roles in neurotransmitter clearance, neurotransmitter release, and oxidative stress. We pharmacologically characterized glutamate transporters in chicken retina cells and identified two glutamate transporters: one Na+-dependent transporter and one Na+-independent transporter. The Na+-dependent uptake system presented characteristics related to the high-affinity xAG- system (EAAT1), and the Na+-independent uptake system presented characteristics related to the xCG- system, which highly contributes to glutamate transport in the retina. Glutamate shares the xCG- system with another amino acid, L-cysteine, suggesting the possible involvement of glutathione. Both transporter proteins are present mainly in Müller glial cells. GABA transporters (GATs) mediate high-affinity GABA uptake from the extracellular space and terminate the synaptic action of GABA in the central nervous system. GABA transporters can be modulated by molecules that act on specific sites to promote transporter phosphorylation and dephosphorylation. In addition to a role in the clearance of GABA, GATs may also release GABA through a reverse transport mechanism. In the chicken retina, a GAT-1 blocker, but not GAT2/3 blocker, was shown to inhibit GABA uptake, suggesting that GABA release from retina cells is mainly mediated by a GAT-1-like transporter.
Acesso aberto (Open Access)
Método baseado me cromatografia líquida de alta eficiência para analisar a atividade dos transportadores de GABA no sistema nervoso central
(Universidade Federal do Pará, 2022-08) MORAES, Edinaldo Rogério da Silva; SILVA, Anderson Manoel Herculano Oliveira da; http://lattes.cnpq.br/8407177208423247
The GATs are the membrane proteins responsible for the uptake of GABA in the central nervous system. Alterations in GAT activity are implicated in several neurological diseases, including retinopathies. The present study describes an alternative method to determine GAT activity in tissue preparations of the central nervous system, using high performance liquid chromatography (HPLC) with fluorescence detection. The GABA concentration in the medium was determined using the o-phthaldehyde (OPA)-derivation protocol validated by the Brazilian Health Regulatory Agency (ANVISA) and the United States Food and Drug Administration (US-FDA). The GAT activity in the retinal preparations was determined through the evaluation of the GABA uptake, which was measured by assessing the difference between the initial and final concentrations of GABA in the incubation médium (Δ𝐺𝑎𝑏𝑎 = [𝐺𝑎𝑏𝑎] 𝑡_0 − [𝐺𝑎𝑏𝑎] 𝑡_𝑒𝑛𝑑). The evaluation of the GAT kinetics returned values of Km = 382.5 ± 32.2 μM and Vmax = 34 nmol/mg of protein. The data also demonstrated that the GABA uptake was predominantly Na+ and temperature-dependent, and was also inhibited by incubation with nipecotic acid, a substrate of GABA transporters. Taken together, these findings confirm that our approach provided a specific measure of GAT activity in retinal tissue. The data presented here thus validate, for the first time, an alternative, simple and sensitive method for the evaluation of GAT activity using high performance chromatography on preparations of the central nervous system.