Binding effects of Alpinia galanga oil and its nanoemulsion 1 to GABAA receptors in rat cortical membranes

Anesthetic activity of Alpinia galanga oil (AGO) has been reported however the mechanism of action in mammals has not been clear. In the present study, the binding effects of AGO and its three active components to gamma-aminobutyric acid type A (GABAA) receptor in cortical membranes of Sprague-Dawley rats were firstly investigated using a [3H]muscimol binding assay. Dimethyl sulfoxide (DMSO) was used to deliver these test samples. The results showed that only AGO and methyl eugenol displayed positive modulation at the highest concentration whereas 1,8-cineole and 4-allylphenyl acetate were inactive. An oil-in-water nanoemulsion containing 20%w/w AGO (NE-AGO) was formulated to deliver AGO instead of DMSO. This NE-AGO significantly enhanced a specific [3H]muscimol binding to 179% of the control with EC50 of 391 µg/mL. The result correlates well to the amount of methyl eugenol in AGO. This result confirms that the anesthetic activity of AGO and methyl eugenol is associated with GABAA receptor modulation, while that of 1,8-cineole and 4-allylphenyl acetate is not and may instead be related to other mechanisms. AGO showed well-tolerated by human cells. Therefore, the formulated NE-AGO might be a promising alternative anesthetic product for humans.

Progesterone's Effects on Cognitive Performance of Male Mice Are Independent of Progestin Receptors but Relate to Increases in GABAA Activity in the Hippocampus and Cortex

Progestogens' (e.g., progesterone and its neuroactive metabolite, allopregnanolone), cognitive effects and mechanisms among males are not well-understood. We hypothesized if progestogen's effects on cognitive performance are through its metabolite allopregnanolone, and not actions via binding to traditional progestin receptors (PRs), then progesterone administration would enhance performance in tasks mediated by the hippocampus and cortex, coincident with increasing allopregnanolone concentrations, brain derived neurotrophic factor (BDNF) and/or muscimol binding of PR knock out (PRKO) and wild-type PR replete mice. Experiment 1: Progesterone (4 mg/kg, subcutaneously (SC; n = 12/grp), or oil vehicle control, was administered to gonadally-intact adult male mice PRKO mice and their wild-type counterparts and cognitive behaviors in object recognition, T-maze and water maze was examined. Progesterone, compared to vehicle, when administered post-training increased time investigating novel objects by the PRKO and wild-type mice in the object recognition task. In the T-maze task, progesterone administration to wild-type and PRKO mice had significantly greater number of spontaneous alternations compared to their vehicle-administered counterparts. In the water maze task, PRKO mice administered vehicle spent significantly fewer seconds in the quadrant associated with the escape platform on testing compared to all other groups. Experiment 2: Progesterone administered to wild-type and PRKO mice increased plasma progesterone and allopregnanolone levels (n = 5/group). PRKO mice had higher allopregnanolone levels in plasma and hippocampus, but not cortex, when administered progesterone and compared to wild-type mice. Experiment 3: Assessment of PR binding revealed progesterone administered wild-type mice had significantly greater levels of PRs in the hippocampus and cortex, compared to all other groups (n = 5/group). Wild-type mice administered progesterone, but not vehicle, had increased BDNF levels in the hippocampus, but not the cortex, compared to PRKOs. Wild-type as well as PRKO mice administered progesterone experienced significant increases in maximal GABAA agonist, muscimol, binding in hippocampus and cortex, compared to their vehicle-administered counterparts. Thus, adult male mice can be responsive to progesterone for cognitive performance, and such effects may be independent of PRs trophic actions of BDNF levels in the hippocampus and/or increases in GABAA activity in the hippocampus and cortex.

Increased GABAA receptor binding in amygdala after prenatal administration of valproic acid to rats

ObjectivePrenatal exposure to valproic acid (VPA) enhances the risk for later development of autism spectrum disorders (ASD). An altered gamma-aminobutyric acid (GABA) system may be a key factor in ASD. Here we investigated possible changes in the GABA system in rats exposed to a low dose of prenatal VPA.MethodWe performed autoradiography with [3H]muscimol, (a GABAA receptor agonist), and [11C]Ro15-4513 (a partial agonist of the GABAA α1+5 receptor subtypes), in brain sections containing amygdala, thalamus and hippocampus of rats treated prenatally with 20 mg/kg VPA or saline from the 12th day of gestation.ResultPrenatal VPA significantly increased [11C]Ro15-4513 binding in the left amygdala compared with controls (p<0.05). This difference was not observed in the hippocampus, thalamus or right amygdala. No differences were observed in [3H]muscimol binding.ConclusionWe observed an asymmetric increase in GABAA receptor binding. Disturbances in the GABAA receptor system have also been detected in human autism with [11C]Ro15-4513.

Acute neurosteroid modulation and subunit isolation of the gamma-aminobutyric acidA receptor in the bullfrog, Rana catesbeiana

The inhibitory neurotransmitter gamma-aminobutyric acid (GABA) has multiple receptors. In mammals, the GABA(A) receptor subtype is modulated by neurosteroids. However, whether steroid interaction with the GABA(A) receptor is unique to mammals or a conserved feature in vertebrates is unknown. Thus, neurosteroid modulation of the GABA(A) receptor was investigated in the brain of the bullfrog (Rana catesbeiana) using the mammalian GABA(A) receptor agonist [(3)H]muscimol. Two neurosteroids, allopregnanolone and pregnenolone sulfate, affected [(3)H]muscimol specific binding in bullfrog brain membrane preparations. Allopregnanolone significantly increased [(3)H]muscimol specific binding in a dose- and time-dependent manner. The pattern of allopregnanolone modulation supports the hypothesis that the bullfrog brain possesses both high-affinity and low-affinity [(3)H]muscimol binding sites. Unlike allopregnanolone, pregnenolone sulfate showed biphasic modulation with increased [(3)H]muscimol specific binding at low nanomolar concentrations and decreased specific binding at micromolar concentrations. Additionally, three cDNA fragments with significant homology to mammalian GABA(A) receptor subunits were isolated from the bullfrog brain. These fragments belong to the alpha1, beta1, and gamma2 subunit families. In mammals, GABA(A) receptors composed of these specific subunit isoforms are effectively modulated by neurosteroids, including allopregnanolone. Neurosteroid modulation of the amphibian brain GABA(A) receptor is therefore supported by both [(3)H]muscimol binding studies and subunit sequences. Allopregnanolone and pregnenolone sulfate modulation of this receptor may thus represent a significant mechanism for steroid influence on amphibian brain and behavior.