Isoguvacine

Manufactured by Merck Group

Sourced in United States, United Kingdom

Isoguvacine is a laboratory reagent used in research applications. It is a GABA(A) receptor agonist that can be utilized in the study of neurological processes and signaling mechanisms.

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Lab products found in correlation

Bumetanide, by Merck Group (6 mentions) Gabazine, by Bio-Techne (3 mentions) β alanine, by Merck Group (2 mentions) Pcdna1, by Thermo Fisher Scientific (2 mentions) 5 aminovaleric acid, by Merck Group (2 mentions) Glycine, by Merck Group (2 mentions) Dl 2 amino 5 phosphonovaleric acid, by Merck Group (2 mentions) Sr 95531 hydrobromide, by Bio-Techne (2 mentions) Gabazine, by Merck Group (2 mentions) Picrotoxin, by Bio-Techne (2 mentions) Clampfit 10, by Molecular Devices (2 mentions) Muscimol, by Merck Group (1 mentions) Mecamylamine hydrochloride, by Bio-Techne (1 mentions) Methyllycaconitine citrate, by Bio-Techne (1 mentions) Dihydro β erythroidine hydrobromide, by Bio-Techne (1 mentions) Digidata 1440a, by Molecular Devices (1 mentions) Gaboxadol, by Merck Group (1 mentions) 4 5 6 7 tetrahydroisoxazolo 5 4 c pyridin 3 ol thip, by Merck Group (1 mentions) Vigabatrin, by Merck Group (1 mentions) Cgp35348, by Merck Group (1 mentions)

13 protocols using isoguvacine

Extracellular Recordings of Hippocampal Slices

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Extracellular field recordings were performed in the CA3 pyramidal layer of hippocampal slices of P16 mice with glass pipettes (Harvard Apparatus, MA, USA) containing ACSF. Spike frequency was recorded for 10 min (control period), then isoguvacine (10 μM; Sigma-Aldrich, MO, USA) was applied in the bath solution for 90 s (isoguvacine period). Following isoguvacine application, spike frequency was recorded for an extra 15 min (wash-out period). Signals were recorded with a low-noise multichannel DAM-80A amplifier (WPI, UK; low-pass filter 1 Hz; high-pass filter 3 kHz; gain ×1000) and digitized online with a Digidata 1400A digitizer (Molecular Devices, CA, USA). Recordings were analyzed using Clampfit 10.4 software (Molecular Devices, CA, USA). The spike detection threshold was defined as three times the standard deviation of the noise recorded in the bath solution. Spike frequency was calculated for control, isoguvacine, and wash-out periods. Slices for which wash-out spike frequency did not come back to control levels (±20% of control) were excluded from this study.

Chiesa M., Nardou R., Lozovaya N., Eftekhari S., Tyzio R., Guimond D., Ferrari D.C, & Ben-Ari Y. (2019). Enhanced Glutamatergic Currents at Birth in Shank3 KO Mice. Neural Plasticity, 2019, 2382639.

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GABAA Agonist Microinjections to Inhibit Kölliker-Fuse Nucleus

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Bilateral microinjections of isoguvacine (a GABA_A agonist, 10 mM, Sigma–Aldrich) containing 5% fluorescent latex microbeads (Lumafluor, New City, NY, USA) were performed into the KF to pharmacologically inhibit this region. Before the isoguvacine microinjections, the KF was functionally identified by microinjections of L-glutamate (10 mM, Sigma–Aldrich), which evoked an increase in cVN post-I activity and PN apnea (Bautista and Dutschmann, 2014 (link)), as demonstrated in Fig. 1. The glass pipettes contained either the drug-microbead mixture or L-glutamate and were positioned in the KF using the following stereotaxic coordinates: 5.5 mm rostral from the calamus scriptorius; 1.75 mm lateral from midline; and 1.5 mm ventral to the dorsal surface. The volume of each microinjection was 70–90 nL. Despite the accuracy of the bilateral injections (as observed with both the histology and the consistent reduction in the cVN post-inspiratory peak), these injections may have not necessarily inhibited all KF neurons.

JENKIN S.E., MILSOM W.K, & ZOCCAL D.B. (2017). THE KÖLLIKER–FUSE NUCLEUS ACTS AS A TIMEKEEPER FOR LATE-EXPIRATORY ABDOMINAL ACTIVITY. Neuroscience, 348, 63-72.

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Expressing and Analyzing GABA-ρ Receptors

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The subcloned human GABA-ρ1 cDNA into pcDNA1.1 (Invitrogen, San Diego, CA, USA) was generously donated by Dr. George Uh1 (National Institute for Drug Abuse, Baltimore, MD, USA). The subcloned human GABA-ρ2 cDNA into PKS (Invitrogen) was generously provided by Dr. Garry Cutting (Center for Medical Genetics, Johns Hopkins University School of Medicine, Baltimore, MD, USA).
GABA, muscimol, β-alanine, 5-amino valeric acid, glycine, isoguvacine, and imidazole-4-acetic acid were purchased from Sigma-Aldrich (Sigma-Aldrich Pty Ltd., Castle Hill, NSW 1765, Australia). TACA and CACA were prepared as previously reported [3 (link)].

Naffaa M.M., Hibbs D.E., Chebib M, & Hanrahan J.R. (2022). Pharmacological Effect of GABA Analogues on GABA-ρ2 Receptors and Their Subtype Selectivity. Life, 12(1), 127.

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Pharmacological Modulation of Neuronal Signaling

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For in vitro experiments, SR 95531 hydrobromide (gabazine 5 μM, Tocris Bioscience, UK, Ref. 1262), NMDA (10 μM, Tocris Bioscience, UK, Ref. 0114), 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline-2,3-dione (10 µM, NIH generous gift), GABA (10 μM, Sigma-Aldrich), isoguvacine (10 μM, Sigma-Aldrich, Ref. G002), DL -2-Amino-5-phosphonovaleric acid (40 µM, Sigma-Aldrich, Ref. A5282), and bumetanide (10 μM, Sigma-Aldrich, Ref. B3023) were directly added to the perfusion solutions. For ca experiments, slices were treated with bumetanide for 40 min before and during recordings. Cocktail of nicotinic receptor antagonists included mecamylamine hydrochloride (10 μM, Tocris Bioscience, Ref. 2843/10), methyllycaconitine citrate (0.1 μM, Tocris Bioscience, Ref. 1029/5), and dihydro-β-erythroidine hydrobromide (10 μM, Tocris Bioscience, Ref. 2349/10). For in vivo experiments, bumetanide pretreatment (3 mg kg⁻¹) was given to mice in drinking water during 5 weeks.

Lozovaya N., Eftekhari S., Cloarec R., Gouty-Colomer L.A., Dufour A., Riffault B., Billon-Grand M., Pons-Bennaceur A., Oumar N., Burnashev N., Ben-Ari Y, & Hammond C. (2018). GABAergic inhibition in dual-transmission cholinergic and GABAergic striatal interneurons is abolished in Parkinson disease. Nature Communications, 9, 1422.

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Extracellular Recording of Hippocampal Slices

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Hippocampal slices are individually transferred to a recording chamber maintained at 30–32°C and continuously perfused (2 mL/min) with oxygenated normal or adapted ACSF. Extracellular field recordings are made using tungsten wire electrodes (diameter: 50 μm, California Fine Wire, Grover Beach, CA, United States). Recording electrodes are positioned in a pyramidal cell layer of CA3 subfield, and signals are amplified using custom- DAM-8A amplifiers (WPI, GB; low-pass filter: 0.1 Hz; high-pass filter: 3 kHz; gain: x1000) and then acquired using an A/D converter (Digidata 1440A, Axon Instruments). Clampfit 10.1 (Axon Instruments) software is used for the acquisition and analysis of the network activity. Isoguvacine and bumetanide are purchased from Sigma.

Goubert E., Altvater M., Rovira M.N., Khalilov I., Mazzarino M., Sebastiani A., Schaefer M.K., Rivera C, & Pellegrino C. (2019). Bumetanide Prevents Brain Trauma-Induced Depressive-Like Behavior. Frontiers in Molecular Neuroscience, 12, 12.

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Recombinant Expression of Human GABA Receptor

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Human ρ1 cDNA subcloned into pcDNA1.1 (Invitrogen, San Diego, CA, USA) was kindly provided by Dr. George Uh1 (National Institute for Drug Abuse, Baltimore, MD, USA).
(GABA, 5-aminovaleric acid, β-alanine, glycine, isoguvacine, 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) also known as Gaboxadol, were purchased from Sigma-Aldrich (Sigm-Aldrich Pty Ltd, Castle Hill, NSW 1765 Australia). 1,2,5,6-Tetrahydropyridin-4-yl)methylphosphinic acid (TPMPA) [31 (link)], trans-aminocrotonic acid (TACA) and cis-aminocrotonic acid (CACA) [32 (link)], and 2-aminoethylmethane thiosulfonate (MTSEA) [33 (link)] were prepared as previously reported.

Naffaa M.M., Absalom N., Solomon V.R., Chebib M., Hibbs D.E, & Hanrahan J.R. (2016). Investigating the Role of Loop C Hydrophilic Residue ‘T244’ in the Binding Site of ρ1 GABAC Receptors via Site Mutation and Partial Agonism. PLoS ONE, 11(5), e0156618.

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Modulating Chloride Homeostasis and GABA Signaling in Ischemic Injury

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Treatments were applied either during the 2 h exposure to OGD/sham normoxia (T1) or post-exposure, during the 3 h necessary for viability assessment (T2).
We blocked outward Cl⁻ transport with DIOA, a KCC2 blocker (Sigma, 20 µM), and inward Cl⁻ transport with bumetanide (Sigma, 10 µM), a NKCC1 blocker as a means for increasing, respectively decreasing [Cl⁻]_i either during OGD (T1) or post-exposure (T2). We also used the GABA_A receptor antagonist gabazine (Sigma, 30 µM) as well as the GABA_A receptor agonist isoguvacine (Sigma, 40 µM) to modulate the GABA_A channel, both during OGD (T1) and during post-exposure (T2). All treatments were also applied to normoxic cultures. Experimental design and treatment times are illustrated in Figure 1.

Zagrean A.M., Grigoras I.F., Iesanu M.I., Ionescu R.B., Chitimus D.M., Haret R.M., Ianosi B., Ceanga M, & Zagrean L. (2019). Neuronal Transmembrane Chloride Transport Has a Time-Dependent Influence on Survival of Hippocampal Cultures to Oxygen-Glucose Deprivation. Brain Sciences, 9(12), 360.

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Pharmacological Modulation of GABAergic Signaling

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In vitro. Bumetanide (10 μM, Sigma-Aldrich, MO, USA), isoguvacine (10 μM, Sigma-Aldrich, MO, USA), TTX (1 μM, Abcam, Bristol, UK), CNQX (10 μM, Sigma-Aldrich, MO, USA), gabazine (5 μM, Tocris Cookson, Bristol, UK) and DHPG (50 μM, Tocris Bioscience, UK) were directly added to the perfusion solutions.
In vivo. Pregnant mice were randomly assigned for Bumetanide pretreatment (2.5 mg/kg) administered in the drinking water.

Lozovaya N., Nardou R., Tyzio R., Chiesa M., Pons-Bennaceur A., Eftekhari S., Bui T.T., Billon-Grand M., Rasero J., Bonifazi P., Guimond D., Gaiarsa J.L., Ferrari D.C, & Ben-Ari Y. (2019). Early alterations in a mouse model of Rett syndrome: the GABA developmental shift is abolished at birth. Scientific Reports, 9, 9276.

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Neurotransmitter Modulation Assay

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Isoguvacine, baclofen, gabazine, CGP-35348 and vigabatrin were obtained from Sigma Chemical Co. All these chemicals were dissolved in normal saline.

Ding L., Gao R., Xiong X.Q., Gao X.Y., Chen Q., Li Y.H., Kang Y.M, & Zhu G.Q. (2015). GABA in Paraventricular Nucleus Regulates Adipose Afferent Reflex in Rats. PLoS ONE, 10(8), e0136983.

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Drugs Application in Neuronal Bath

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Drugs were applied in the bath via a three-way tap system, by changing the superfusion solution to one differing only in its content of drug(s). Drugs used were: SR 95531 hydrobromide (gabazine), DNQX and picrotoxin purchased from Ascent Scientific (UK), isoguvacine from Sigma-Aldrich (Italy) and TTX from Latoxan (Israel). Stock solutions were made in distilled water and then aliquoted and frozen at −20 °C. DNQX was dissolved in DMSO. The final concentration of DMSO in the bathing solution was 0.1%. At this concentration, DMSO alone did not modify the membrane potential, input resistance or the firing properties of neurons.

Cellot G., Maggi L., Di Castro M.A., Catalano M., Migliore R., Migliore M., Scattoni M.L., Calamandrei G, & Cherubini E. (2016). Premature changes in neuronal excitability account for hippocampal network impairment and autistic-like behavior in neonatal BTBR T+tf/J mice. Scientific Reports, 6, 31696.

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