Tina Hinton

Novel, Potent, and Selective GABAC Antagonists Inhibit Myopia Development and Facilitate Learning and Memory

This study reports pharmacological and physiological effects of cis- and trans-(3-aminocyclopentanyl)butylphosphinic acid (cis- and trans-3-ACPBPA). These compounds are conformationally restricted analogs of the orally active GABAB/C receptor antagonist (3-aminopropyl)-n-butylphosphinic acid (CGP36742 or SGS742). cis-[IC50(ρ1) = 5.06 μM and IC50(ρ2) = 11.08 μM; n = 4] and trans-3-ACPMPA [IC50(ρ1) = 72.58 μM and IC50(ρ2) = 189.7 μM; n = 4] seem competitive at GABAC receptors expressed in Xenopus laevis oocytes, having no effect as agonists (1 mM) but exerting weak antagonist (1 mM) effects on human GABAA and GABAB receptors. cis-3-ACPBPA was more potent and selective than the trans-compound, being more than 100 times more potent at GABAC than GABAA or GABAB receptors. cis-3-ACPBPA was further evaluated on dissociated rat retinal bipolar cells and dose-dependently inhibited the native GABAC receptor (IC50 = 47 ± 4.5 μM; n = 6). When applied to the eye as intravitreal injections, cis- and trans-3-ACPBPA prevented experimental myopia development and inhibited the associated vitreous chamber elongation, in a dose-dependent manner in the chick model. Doses only 10 times greater than required to inhibit recombinant GABAC receptors caused the antimyopia effects. Using intraperitoneal administration, cis- (30 mg/kg) and trans-3-ACPBPA (100 mg/kg) enhanced learning and memory in male Wistar rats; compared with vehicle there was a significant reduction in time for rats to find the platform in the Morris water maze task (p < 0.05; n = 10). As the physiological effects of cis- and trans-3-ACPBPA are similar to those reported for CGP36742, the memory and refractive effects of CGP36742 may be due in part to its GABAC activity.

Stress and GABAA receptors

J. Neurochem. (2010) 112, 1115–1130.

Stress and GABA receptors.

J. Neurochem. (2010) 112, 1115–1130.

Cigarette smoking during pregnancy regulates the expression of specific nicotinic acetylcholine receptor (nAChR) subunits in the human placenta.

Smoking during pregnancy is associated with low birth weight, premature delivery, and neonatal morbidity and mortality. Nicotine, a major pathogenic compound of cigarette smoke, binds to the nicotinic acetylcholine receptors (nAChRs). A total of 16 nAChR subunits have been identified in mammals (9 α, 4 β, and 1 δ, γ and ε subunits). The effect of cigarette smoking on the expression of these subunits in the placenta has not yet been determined, thus constituting the aim of this study. Using RT-qPCR and western blotting, this study investigated all 16 mammalian nAChR subunits in the normal healthy human placenta, and compared mRNA and protein expressions in the placentas from smokers (n = 8) to controls (n = 8). Our data show that all 16 subunit mRNAs are expressed in the normal, non-diseased human placenta and that the expression of α2, α3, α4, α9, β2 and β4 subunits is greater than the other subunits. For mRNA, cigarette smoke exposure was associated with increased expression of the α9 subunit, and decreased expression of the δ subunit. At the protein level, expression of both α9 and δ was increased. Thus, cigarette smoking in pregnancy is sufficient to regulate nAChR subunits in the placenta, specifically α9 and δ subunits, and could contribute to the adverse effects of vasoconstriction and decreased re-epithelialisation (α9), and increased calcification and apoptosis (δ), seen in the placentas of smoking women.

GABA transporters GAT-1 and GAT-3 in the human dorsolateral prefrontal cortex in schizophrenia.

This study aimed to investigate the binding affinity of [3H]GABA and [3H]β-alanine to GABA transporters GAT-1 and GAT-3 in the human dorsolateral prefrontal cortex (Brodmanns’ area 9) in schizophrenia. Using post mortem tissue from individuals diagnosed with schizophrenia (n = 6) and control subjects (n = 6), the density of GAT-1 was established by displacing [3H]GABA with muscimol, and for GAT-3 [3H]β-alanine was used. Data analysis showed a significant decrease of GAT-1 levels (45%), and a significant increase of GAT-3 density (23%) within the dorsolateral prefrontal cortex of individuals diagnosed with schizophrenia when compared to age- and sex-matched controls. The observed decrease of GAT-1 could be explained as a consequence of the GABA hypo-function or the result of volumetric shrinkage of the cerebral cortex previously reported in this disease. The observed elevation of GAT-3 levels could be due to a compensatory effect for any functional loss of GABA re-uptake by the decreased GAT-1 levels.

Sex-differences and stress: Effects on regional high and low affinity [3H]GABA binding

Sex-differences are observed in the GABAergic neurotransmitter system both at rest and following acute stress, yet the brain regions and functional implications of these differences are unknown. We examined sex-differences in the number of low- and high-affinity [3H]GABA binding sites in various brain regions of male and female mice and the effect of stress on such sex-differences. Male (n=6) and female (n=6) QS mice were exposed to a brief swim stress (3 min at 32+/-1 degrees C) either individually or with cage-mates whilst control males (n=6) and females (n=6) remained undisturbed in the home cage. Using quantitative receptor autoradiography, sections of mouse brain were labelled with either 30 or 1000 nM [3H]GABA to label high or low affinity binding sites, respectively. Results indicated that males had more low affinity [3H]GABA binding sites in various forebrain cortical regions but less high affinity binding sites in many of these regions compared with females. Forced swim stress-induced rapid changes in forebrain GABA binding sites in females and group stressed males, suggesting a mechanism for rapid GABAergic adaptations. However the number of functional binding sites for GABA in certain forebrain regions was altered by stress in opposite directions in males and females, such that baseline sex-differences were removed following stress. These results exemplify sex-differences in brain chemical function and stress responses, and are of potential importance for understanding sex-differences in response to GABAergic compounds and disorders with sex and stress as predisposing factors.

Herbal Products and GABA Receptors

This article profiles ten examples of chemicals derivedfrom herbal preparations that influence ionotropicreceptors for the brain’s major inhibitory neurotrans-mitter g-aminobutyric acid (GABA). Many herbalmedicines are used to influence brain function inordertotreatanxiety,cognitivedisorders, andinsom-nia. Such disorders are considered likely to involveGABA receptors, particularly ionotropic receptorsthat are responsible for fast synaptic transmissionvia ion channels. The evidence of efficacy, or other-wise, in the use of herbal medicines to treat centralnervous system (CNS) disorders remains to be estab-lished unequivocally in many cases. There is nodoubt, however, that these products contain chemi-cals that in their pure form can be shown to influencebrain function in specific ways. The structural diver-sity of these chemicals provides valuable leads for thedevelopment of new therapeutic agents and toolswith which to investigate nervous system function.Herbal medicines are usually complex mixtures ofchemicals. The combined actions of these chemicalsmay contribute to the overall effects of the medicines.Thus, it is important to study interactions among theindividualchemicals.WithrespecttoionotropicGABAreceptors,thereareexamplesofsynergisticinteractionsbetweentwochemicalsandexamplesofwhathasbeentermed second-order modulation, in which a secondmodulator enhances the action of a first-order modu-lator.Thismayrepresentanewformofdrugactionandlead to the identification of new drug targets. Investi-gating the complex ways in which these chemicalsinteract could lead to a better understanding of thefunction of ionotropic GABA receptors.GABA itself is an important plant constituent, andthus it should not be surprising that plants contain arange of other chemicals that can influence GABAfunction. In addition, there are examples of plant-derived GABA mediating some form of communica-tionamonganimals,bacteria,fungi,andplants.Thus,GABA may represent a conserved and ubiquitousbiological signaling molecule.Quality control of herbal medicines is a significantproblem. Identification of key active ingredients inherbal medicines and the interactions among themmay provide the means to better quality control usingfunctional assays in addition to the currently usedchemical assays. Many of the advances in studies ofherbal products and GABA receptors have come fromligand binding studies, for example, from the bindingof benzodiazepines to brain membranes and, morerecently, from functional studies in which GABAreceptorsofknownsubunitcompositionareexpressedincellsthatdonotusuallyexpresssuchreceptors(e.g.,frog oocytes).

Antipsychotic drug administration differentially affects [3H]muscimol and [3H]flunitrazepam GABAA receptor binding sites

Post-mortem studies of the human brain indicate that certain GABA(A) receptor subtypes may be differentially altered in schizophrenia. Increased binding to the total population of GABA(A) receptors using [3H]muscimol is observed in the post-mortem schizophrenic brain, yet a proportion of these receptors which bind benzodiazepines and are labelled with [3H]flunitrazepam, show decreased or unaltered expression. Data from animal studies suggest that antipsychotic drugs alter GABA(A) receptor expression in a subtype selective manner, but in the opposite direction to that observed in schizophrenia. To broaden our understanding of the effects of antipsychotic drugs on GABA(A) receptors, we examined the saturation binding maximum (B(max)) and binding affinity (K(D)) of [3H]muscimol and [3H]flunitrazepam in the prefrontal cortex (PFC), hippocampus and thalamus of male SD rats that received a sucrose solution containing either haloperidol (1.5 mg/kg), olanzapine (6.5 mg/kg) or no drug daily for up to 28 days using quantitative receptor autoradiography. [3H]Muscimol binding density was increased most prominently in the PFC after 7 days, with larger and more prolonged effects being induced by the atypical antipsychotic drug olanzapine in subcortical regions. While no changes were observed in [3H]muscimol binding in any region after 28 days of drug administration, [3H]flunitrazepam binding density (B(max)) was increased for both antipsychotic treatments in the PFC only. These findings confirm that the subset of GABA(A) receptors sensitive to benzodiazepines are regulated differently from other GABA(A) receptor subtypes following antipsychotic drug administration, in a time- and region-dependent manner.

Nicotinic acetylcholine receptors (nAChR) are increased in the pre-eclamptic placenta.

Objective: The role of the nicotinic acetylcholine receptors (nAChR) in pre-eclampsia is unknown. Given that ACh levels are affected in pre-eclampsia, it has been suggested that compensatory changes in nAChR expression may ensue. This study aimed to determine the effects of pre-eclampsia on the mRNA and protein expression of 12 mammalian nAChR subunits. Methods: Placentas were collected from healthy term pregnancies (n = 8) and pregnancies complicated by pre-eclampsia (n = 7), both being non-cigarette smoke exposed to rule out any role of nicotine. Using real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR), 12 subunits (α2, α3, α4, α5, α6, α7, α9, β1, β2, β4, δ, and γ) were able to be studied at the mRNA level, while at the protein level using Western blotting, nine subunits (α2, α3, α4, α5, α7, α9, β1, β2, and γ) were studied. Results: At the mRNA level, pre-eclamptic placentas showed an increase in α2 (p = 0.003), α9 (p < 0.001), β1 (p = 0.03) and β2 (p = 0.02) subunit expression, while at the protein level, α7 (p = 0.004), α9 (p = 0.02), and δ (p = 0.003) subunits were increased compared to controls. Conclusion: Certain nAChR subunits are increased in the pre-eclamptic placenta. Given the absence of cigarette smoking, the changes in expression are hypothesised to be due to the hypoxic environment resulting from the pathophysiology of pre-eclampsia, which subsequently affects endogenous ACh levels, yielding compensatory increases in α2, α7, α9, β1, β2, and δ nAChR subunits.

An evaluation of pharmacology curricula in Australian science and health-related degree programs

BackgroundPharmacology is a biomedical discipline taught in basic science and professional degree programs. In order to provide information that would facilitate pharmacology curricula to be refined and developed, and approaches to teaching to be updated, a national survey was undertaken in Australia that investigated pharmacology course content, teaching and summative assessment methods.MethodsTwenty-two institutions participated in a purpose-built online questionnaire, which enabled an evaluation of 147 courses taught in 10 different degrees. To enable comparison, degrees were grouped into four major degree programs, namely science, pharmacy, medicine and nursing. The pharmacology content was then classified into 16 lecture themes, with 2-21 lecture topics identified per theme. The resultant data were analysed for similarities and differences in pharmacology curricula across the degree programs.ResultsWhile all lecture themes were taught across degree programs, curriculum content differed with respect to the breadth and hours of coverage. Overall, lecture themes were taught most broadly in medicine and with greatest coverage in pharmacy. Reflecting a more traditional approach, lectures were a dominant teaching method (at least 90% of courses). Sixty-three percent of science courses provided practical classes but such sessions occurred much less frequently in other degree programs, while tutorials were much more common in pharmacy degree programs (70%). Notably, problem-based learning was common across medical programs. Considerable diversity was found in the types of summative assessment tasks employed. In science courses the most common form of in-semester assessment was practical reports, whereas in other programs pen-and-paper quizzes predominated. End-of-semester assessment contributed 50-80% to overall assessment across degree programs.ConclusionThe similarity in lecture themes taught across the four different degree programs shows that common knowledge- and competency-based learning outcomes can be defined for pharmacology. The authors contend that it is the differences in breadth and coverage of material for each lecture theme, and the differing teaching modes and assessment that characterise particular degree programs. Adoption of pharmacology knowledge-based learning outcomes that could be tailored to suit individual degree programs would better facilitate the sharing of expertise and teaching practice than the current model where pharmacology curricula are degree-specific.