HPLC-Based Activity Profiling for GABAA Receptor Modulators in Searsia pyroides Using a Larval Zebrafish Locomotor Assay

Planta Medica ◽  
2017 ◽  
Vol 83 (14/15) ◽  
pp. 1169-1175 ◽  
Author(s):  
Fahimeh Moradi-Afrapoli ◽  
Hannes van der Merwe ◽  
Maria De Mieri ◽  
Anke Wilhelm ◽  
Marco Stadler ◽  
...  
Keyword(s):  
Xenopus Oocytes ◽  
High Resolution Mass Spectrometry ◽  
Aminobutyric Acid ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid ◽  
Concentration Dependent Manner ◽  
Hplc Fraction ◽  
Chloride Currents ◽  
Zebrafish Larvae ◽  
Activity Profiling

AbstractA dichloromethane extract from leaves of Searsia pyroides potentiated gamma aminobutyric acid-induced chloride currents by 171.8 ± 54% when tested at 100 µg/mL in Xenopus oocytes transiently expressing gamma aminobutyric acid type A receptors composed of α 1 β 2 γ 2s subunits. In zebrafish larvae, the extract significantly lowered pentylenetetrazol-provoked locomotion when tested at 4 µg/mL. Active compounds of the extract were tracked with the aid of HPLC-based activity profiling utilizing a previously validated zebrafish larval locomotor activity assay. From two active HPLC fractions, compounds 1 – 3 were isolated. Structurally related compounds 4 – 6 were purified from a later eluting inactive HPLC fraction. With the aid of 1H and 13C NMR and high-resolution mass spectrometry, compounds 1 – 6 were identified as analogues of anacardic acid. Compounds 1 – 3 led to a concentration-dependent decrease of pentylenetetrazol-provoked locomotion in the zebrafish larvae model, while 4 – 6 were inactive. Compounds 1 – 3 enhanced gamma aminobutyric acid-induced chloride currents in Xenopus oocytes in a concentration-dependent manner, while 4 – 6 only showed marginal enhancements of gamma aminobutyric acid-induced chloride currents. Compounds 2, 3, and 5 have not been reported previously.

gamma-Aminobutyric acid-induced response in rat dissociated paratracheal ganglion cells

1992 ◽  
Vol 67 (5) ◽  
pp. 1367-1374 ◽  
Author(s):  
S. Itabashi ◽  
K. Aibara ◽  
H. Sasaki ◽  
N. Akaike
Keyword(s):  
Response Curve ◽  
Ganglion Cells ◽  
Aminobutyric Acid ◽  
Equilibrium Potential ◽  
Induced Response ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Concentration Response Curve

1. The pharmacologic properties of gamma-aminobutyric acid (GABA)-induced Cl- current (ICl) were studied in the paratracheal ganglion cells freshly dissociated from 7- to 10-day-old rat trachea in a whole-cell recording mode by the use of a conventional patch-clamp technique. 2. GABA- and muscimol-induced currents increased sigmoidally in a concentration-dependent manner, and both currents reversed at approximately -3 mV, which was close to the Cl- equilibrium potential (ECl). 3. Strychnine (STR) at low concentration and bicuculline (BIC) inhibited GABA response competitively, whereas STR at the higher concentrations, benzylpenicillin (PCG), or picrotoxin (PTX) inhibited noncompetitively. Inhibition of GABA response by PCG but not other antagonists was voltage dependent, indicating that PCG acts as a Cl- channel blocker. 4. The concentration-response curve of pentobarbital sodium (PB)-induced ICl was bell shaped. At concentrations higher than 10(-3) M, both the peak and plateau currents decreased, and a transient "hump" current appeared immediately after washing out PB. In the presence of PB, the concentration-response curve of GABA shifted toward left without changing the maximum response. 5. Although diazepam (DZP) at concentration used did not induce a response, it potentiated the GABA response in a concentration-dependent manner between 10(-8) and 10(-6) M. DZP also caused a parallel shift toward left in the concentration-response curve of GABA. 6. PB or DZP further enhanced the GABA response in the presence of the other agent. 7. It is concluded that the properties of GABAA receptors in the paratracheal ganglion cells are essentially similar to those reported in other preparations.

scholarly journals HPLC-Based Activity Profiling for GABAA Receptor Modulators in Murraya exotica

2019 ◽  
Vol 14 (1) ◽  
pp. 1934578X1901400
Author(s):  
Nova Syafni ◽  
Fahimeh Moradi-Afrapoli ◽  
Ombeline Danton ◽  
Anke Wilhelm ◽  
Marco Stadler ◽  
...  
Keyword(s):  
Gaba Receptors ◽  
Xenopus Oocyte ◽  
Aminobutyric Acid ◽  
Xenopus Laevis Oocytes ◽  
Gamma Aminobutyric Acid ◽  
Activity Assay ◽  
Chloride Currents ◽  
Activity Profiling ◽  
Murraya Exotica ◽  
Receptor Modulators

A dichloromethane extract from twigs and leaves of Murraya exotica produced allosteric potentiation of gamma aminobutyric acid (GABA) induced chloride currents in a microelectrode assay in Xenopus laevis oocytes expressing GABA receptors of α1, β2, γ2s subunit composition. The activity was tracked by HPLC-based activity profiling utilizing a zebrafish locomotor activity assay. Osthol (9) was identified as the main active compound. In addition, five other coumarins and four flavonols were identified. Osthol (9) and structurally related coumurrayin (10) were tested in the Xenopus oocyte assay. Compound 9 potentiated GABAA-induced chloride currents by 487 ± 42%, with an EC50 of 46 ± 10 μM, while 10 showed negligible effects on chloride currents. In silico evaluation of physicochemical properties showed that 9 and 10 had properties that are favorable for oral bioavailability and BBB permeability.

gamma-Aminobutyric acid stimulates acid secretion from the isolated guinea pig stomach

1987 ◽  
Vol 253 (5) ◽  
pp. G601-G606
Author(s):  
L. H. Tsai ◽  
K. Taniyama ◽  
C. Tanaka
Keyword(s):  
Guinea Pig ◽  
Acid Secretion ◽  
Cholinergic Neurons ◽  
Aminobutyric Acid ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid ◽  
Concentration Dependent Manner ◽  
High Concentrations ◽  
Guinea Pig Stomach ◽  
Spontaneous Secretion

gamma-Aminobutyric acid (GABA) content was measured, and the effect of GABA on acid secretion was studied using the everted preparation of isolated guinea pig stomachs. GABA contents in the mucosa layer and the remaining layer were 20-24 nmol/g tissue and 34-42 nmol/g tissue, respectively. GABA at 10(-6) to 3 X 10(-5) M induced acid secretion, and the maximum secretion was obtained at 3 X 10(-5) M, that is approximately 1.6-fold of the spontaneous secretion and approximately half of the amount secreted by histamine at 3 X 10(-4) M. The GABA-induced acid secretion was inhibited by bicuculline, scopolamine, pirenzepine, proglumide, and tetrodotoxin, but not by cimetidine. Muscimol (3 X 10 to 10(-5) M), but not baclofen, induced acid secretion in a concentration-dependent manner. The responses to GABA and muscimol were antagonized by bicuculline. Scopolamine and tetrodotoxin completely inhibited the acid secretion induced by low concentrations of GABA and muscimol and to some extent the response induced by high concentrations of muscimol. All these results indicate that GABA induces acid secretion via the A type of GABA receptor, probably located mainly on the cholinergic neurons and partially on the nonneuronal cells in the guinea pig stomach.

gamma-Aminobutyric acid-induced response in acutely isolated nucleus solitarii neurons of the rat

1991 ◽  
Vol 260 (4) ◽  
pp. C745-C749 ◽  
Author(s):  
T. Nakagawa ◽  
M. Wakamori ◽  
T. Shirasaki ◽  
T. Nakaye ◽  
N. Akaike
Keyword(s):  
Response Curve ◽  
Aminobutyric Acid ◽  
Equilibrium Potential ◽  
Induced Response ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Voltage Dependent ◽  
Concentration Response Curve

The gamma-aminobutyric acid (GABA)-induced macroscopic Cl- current (ICl) was investigated in acutely isolated nucleus tractus solitarii (NTS) neurons by a conventional patch-clamp technique combined with a rapid drug application method. The GABA- and muscimol-induced ICl increased in a concentration-dependent manner. The reversal potentials were close to the Cl- equilibrium potential. Pentobarbital sodium (PB) itself elicited a current. Bicuculline (BIC), strychnine (STR), picrotoxin, benzylpenicillin (PCG), Cd2+, and Zn2+ suppressed the GABA response in a concentration-dependent manner. Both BIC and STR shifted the concentration-response curve for GABA response to the right, whereas PCG suppressed the maximum response without affecting the threshold, indicating that BIC and STR antagonized competitively and PCG noncompetitively. The inhibitory action of PCG on GABA response was in a highly voltage-dependent manner. PB shifted the concentration-response curve for GABA response to the left. The augmentatory effect of PB was voltage dependent.

scholarly journals The effect of baclofen on spontaneous and evoked behavioural expression of experimental neuropathic chronic pain

1999 ◽  
Vol 57 (3B) ◽  
pp. 753-760 ◽  
Author(s):  
TEREZINHA DE JESUS T. SANTOS ◽  
CARLOS M. DE CASTRO-COSTA ◽  
SÍLVIO D. A. GIFFONI ◽  
FRANKLIN J. C. SANTOS ◽  
RODRIGO S. N. RAMOS ◽  
...  
Keyword(s):  
Chronic Pain ◽  
Nervous System ◽  
Neuropathic Pain ◽  
Aminobutyric Acid ◽  
Thermal Stimulus ◽  
Dependent Manner ◽  
Gamma Aminobutyric Acid ◽  
Analgesic Effects ◽  
Nociceptive Stimulus ◽  
Dose Dependent

Baclofen (beta-p-chlorophenyl-GABA) has been used in humans to treat spasticity, as well as trigeminal neuralgia. Since GABA (gamma-aminobutyric acid) has been implicated in inhibitory and analgesic effects in the nervous system, it was of interest to study the effect of baclofen in experimental neuropathic pain. With this purpose, experiments were carried out in 17 neuropathic rats with constrictive sciatic injury, as described by Bennet and Xie (1988), taking as pain parameters scratching behaviour and the latency to the thermal nociceptive stimulus. The results showed that baclofen induces, in a dose-dependent manner, significant decrease (p < 0.05) of scratching behaviour and significant increase (p < 0.05) of the latency to the nociceptive thermal stimulus. The absence of antagonism of naloxone suggested a non-participation of an opioid-mediated mechanism in this analgesic effect of baclofen on experimental neuropathic pain.

Evidence for a glutamatergic neural pathway in the myenteric plexus

1991 ◽  
Vol 261 (4) ◽  
pp. G693-G700 ◽  
Author(s):  
J. W. Wiley ◽  
Y. X. Lu ◽  
C. Owyang
Keyword(s):  
Myenteric Plexus ◽  
Longitudinal Muscle ◽  
Muscle Tension ◽  
Nmda Receptor Antagonist ◽  
Aminobutyric Acid ◽  
Dependent Manner ◽  
Neural Pathway ◽  
Concentration Dependent Manner ◽  
Excitatory Neurotransmitter ◽  
Somatostatin 14

The objective of this study was to determine whether L-glutamate (L-Glu) may serve as a neurotransmitter candidate in the guinea pig myenteric plexus. We observed that [3H]Glu and gamma-[3H]aminobutyric acid were synthesized from [3H]glutamine and released from neurons of the myenteric plexus during K+ and 1,1-dimethyl-4-phenylpiperazinium-evoked depolarization in a concentration-dependent manner. Muscle tension studies performed on ileal longitudinal muscle-myenteric plexus (LM-MP) preparations revealed that L-Glu [mean effective dose (ED50) 2.5 x 10(-5) M] produced concentration-dependent contractions, which were unaffected by hexamethonium but abolished by tetrodotoxin, atropine, and magnesium, suggesting that L-Glu acts via N-methyl-D-aspartate (NMDA)-type receptors that stimulate a cholinergic neural pathway unaffected by ganglionic blockade. In addition, L-Glu (ED50 4 x 10(-5) M) and NMDA (ED50 2 x 10(-4) M) stimulated concentration-dependent release of [3H]acetylcholine (ACh) from LM-MP sections, which was inhibited by tetrodotoxin, magnesium, and the NMDA receptor antagonist D-2-amino-5-phosphonovaleric acid (AP-5). L-Glu-mediated release of [3H]ACh was enhanced by theophylline (10-6 M) and 3-isobutyl-1-methylxanthine (1 mM) and was significantly reduced by the adenylate cyclase inhibitor, 2',5'-dideoxyadenosine (10(-4) M) and somatostatin-14 (10(-6) M), which inhibits adenosine 3',5'-cyclic monophosphate (cAMP)-dependent cholinergic transmission in the myenteric plexus. These studies suggest that L-Glu may serve as an excitatory neurotransmitter in the myenteric plexus via its action on NMDA-type receptors, which are coupled to cAMP-dependent release of ACh.

4-Aminopyridine block of the noninactivating cloned K+ channel Kv1.5 expressed in Xenopus oocytes

1995 ◽  
Vol 269 (2) ◽  
pp. H556-H564 ◽  
Author(s):  
T. Yamane ◽  
T. Furukawa ◽  
M. Hiraoka
Keyword(s):  
Xenopus Oocytes ◽  
Inhibitory Concentration ◽  
External Solution ◽  
K Channel ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
External Application ◽  
Charged Form ◽  
Cytoplasmic Side ◽  
Voltage Clamp Method

The blocking action of 4-aminopyridine (4-AP) on the cloned K+ channel Kv1.5 expressed in Xenopus oocytes was studied using the two-microelectrode voltage-clamp method. Application of 4-AP to the bath solution reversibly suppressed the expressed current in a voltage- and concentration-dependent manner decreasing with membrane depolarization and with a half-maximal inhibitory concentration of 0.14 mM (at +40 mV). Both block and unblock occurred mainly during a depolarization when channels were activated. With successive depolarizations, 4-AP decreased not only the peak amplitudes of the current in successive pulses, but also the current during a depolarization. Upon washout of 4-AP, the current recovered with successive depolarizations, whereas no recovery of the current was noted in the absence of depolarizations. The extent of block markedly increased with alkalization of the external solution and decreased with acidification. External application of 4-amino-pyridine methiodide, a charged form of a quaternary 4-AP derivative, did not affect the current, but internal application markedly suppressed the current, indicating the drug gained access to the channel from the cytoplasmic side. These data suggest that 4-AP crosses the membrane in its uncharged form and acts from inside of the cell in its charged form, resulting in block of the channels with higher affinity to the open state.

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