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Glutamine and its Neuroactive Derivatives in the Retina
Published in Elling Kvamme, Glutamine and Glutamate in Mammals, 1988
The initial rate of GABA accumulation, following inhibition of its catabolic enzyme GABA-T, with, e.g., γ-vinyl GABA or gabaculine, potentially provides an index of GABA utilization.91 So far, in studies on the retina, this method has been applied only to the chick92 and rat,87,93 and results with the rat suggest that the total turnover of GABA decreases when the tissue is exposed to low levels of illumination.93 Rates may rise again at higher light levels,87,93 but there are anomalies93 and more work is needed. Label and chase techniques, using 3H-glutamine as precursor, add some support to these findings as the relative specific activity of GABA in the isolated, dark-adapted rat retina decreases on exposure to dim light (Section V) and may rise again in bleached tissue.94
The Genetically Epilepsy-Prone Rat: Neuronal Networks And Actions Of Amino Acid Neurotransmitters
Published in Carl L. Faingold, Gerhard H. Fromm, Drugs for Control of Epilepsy:, 2019
Carl L. Faingold, Dean K. Naritoku
Microinjection of a variety of agents that enhance the action of GABA into the IC and other brain regions of AGS-susceptible rats blocks AGS susceptibility. Thus, focal microinjection into the IC of GABA agonists, including GABA (in high doses), muscimol, a predominantly GABAA agonist, the GABAA agonist, THIP (4,5,6,7-tetrahydroisoxa-zolo[5,4-c]pyridin-3-ol), and baclofen, a GABAB agonist, blocked seizure susceptibility in the GEPR-9 or the GEPR-antecedent,26,30,36,82 (Table 3). Bilateral microinjection of gaba-culine, an irreversible inhibitor of GABA transaminase, into the IC of the GEPR-9 completely blocks seizure susceptibility.33 The delayed onset and prolonged duration of the effect of gabaculine as compared to GABA agonists is consistent with the time course for elevation of GABA levels produced by this agent. The metabolic effect of gabaculine was confirmed by the significant increase in GABA levels produced in the IC by gabaculine microinfusion.83 The GABA uptake inhibitor, NO-328 (tiagabine), also is effective in blocking AGS in the GEPR when injected bilaterally into the IC84 (see Table 3). Taurine microinjection into IC also produced a long-lasting blockade of seizure susceptibility in GEPR-antecedents.23 AGS in rats undergoing ethanol withdrawal were blocked by microinfusion into the IC of the GABAA agonists, muscimol and THIP, the GABAB agonist, baclofen, and the benzodiazepine, chlordiazepoxide.26,39 These data, coupled with the ability of bilateral microinjection of the GABAA antagonist, bicuculline, into the IC to produce susceptibility to AGS in normal rats,25 strongly support a vital role for GABA in initiation mechanisms for AGS in the IC.
High-yield synthesis and purification of recombinant human GABA transaminase for high-throughput screening assays
Published in Journal of Enzyme Inhibition and Medicinal Chemistry, 2021
Mingu Gordon Park, Ah-reum Han, Su Yeon Kim, Tai Young Kim, Ho Min Kim, C. Justin Lee
GABase (Merck, G7509) is a convenient and inexpensive source of SSADH, as it is a commercially available mixture composed of bacterial GABA-T and SSADH obtained from Pseudomonas fluorescens. However, GABase was not able to be used directly in our human GABA-T assay due to the presence of bacterial GABA-T. So we used the bacterial SSADH isolation method as previously described39, which allowed large amounts of bacterial SSADH to be isolated quickly and conveniently from GABase by irreversibly inhibiting the bacterial GABA-T activity. Five milligrams of gabaculine (Santa Cruz Biotechnology, sc-200473), a naturally occurring GABA-T irreversible inhibitor, was added to 25 ml of 1 U/mL GABase solution (reconstituted with 50 mM K4P2O7 buffer at pH 8.6), followed by maintaining the reaction at 4 °C for 30 min. The residual gabaculine, which also can inactivate recombinant human GABA-T, was eliminated by dialysis at 4 °C with 2 L of K4P2O7 buffer, changed 3 times per 9 h followed by the final dialysis overnight. The isolated bacterial SSADH was stored at −80 °C for long-term storage.