Kavain, the Major Constituent of the Anxiolytic Kava Extract, Potentiates GABAA Receptors: Functional Characteristics and Molecular Mechanism.

Extracts of the pepper plant kava (Piper methysticum) are effective in alleviating anxiety in clinical trials. Despite the long-standing therapeutic interest in kava, the molecular target(s) of the pharmacologically active constituents, kavalactones have not been established. γ-Aminobutyric acid type A receptors (GABAARs) are assumed to be the in vivo molecular target of kavalactones based on data from binding assays, but evidence in support of a direct interaction between kavalactones and GABAARs is scarce and equivocal. In this study, we characterised the functional properties of the major anxiolytic kavalactone, kavain at human recombinant α1ß2, ß2γ2L, αxß2γ2L (x = 1, 2, 3 and 5), α1ßxγ2L (x = 1, 2 and 3) and α4ß2δ GABAARs expressed in Xenopus oocytes using the two-electrode voltage clamp technique. We found that kavain positively modulated all receptors regardless of the subunit composition, but the degree of enhancement was greater at α4ß2δ than at α1ß2γ2L GABAARs. The modulatory effect of kavain was unaffected by flumazenil, indicating that kavain did not enhance GABAARs via the classical benzodiazepine binding site. The ß3N265M point mutation which has been previously shown to profoundly decrease anaesthetic sensitivity, also diminished kavain-mediated potentiation. To our knowledge, this study is the first report of the functional characteristics of a single kavalactone at distinct GABAAR subtypes, and presents the first experimental evidence in support of a direct interaction between a kavalactone and GABAARs.

Probing α4ßδ GABAA receptor heterogeneity: differential regional effects of a functionally selective α4ß1δ/α4ß3δ receptor agonist on tonic and phasic inhibition in rat brain.

In the present study, the orthosteric GABAA receptor (GABAAR) ligand 4,5,6,7-tetrahydroisothiazolo[5,4-c]pyridin-3-ol (Thio-THIP) was found to possess a highly interesting functional profile at recombinant human GABAARs and native rat GABAARs. Whereas Thio-THIP displayed weak antagonist activity at α1,2,5ß2,3γ2S and ρ1 GABAARs and partial agonism at α6ß2,3δ GABAARs expressed in Xenopus oocytes, the pronounced agonism exhibited by the compound at α4ß1δ and α4ß3δ GABAARs was contrasted by its negligible activity at the α4ß2δ subtype. To elucidate to which extent this in vitro profile translated into functionality at native GABAARs, we assessed the effects of 100 µm Thio-THIP at synaptic and extrasynaptic receptors in principal cells of four different brain regions by slice electrophysiology. In concordance with its α6ß2,3δ agonism, Thio-THIP evoked robust currents through extrasynaptic GABAARs in cerebellar granule cells. In contrast, the compound did not elicit significant currents in dentate gyrus granule cells or in striatal medium spiny neurons (MSNs), indicating predominant expression of extrasynaptic α4ß2δ receptors in these cells. Interestingly, Thio-THIP evoked differential degrees of currents in ventrobasal thalamus neurons, a diversity that could arise from differential expression of extrasynaptic α4ßδ subtypes in the cells. Finally, whereas 100 µm Thio-THIP did not affect the synaptic currents in ventrobasal thalamus neurons or striatal MSNs, it reduced the current amplitudes recorded from dentate gyrus granule cells, most likely by targeting perisynaptic α4ßδ receptors expressed at distal dendrites of these cells. Being the first published ligand capable of discriminating between ß2- and ß3-containing receptor subtypes, Thio-THIP could be a valuable tool in explorations of native α4ßδ GABAARs.

GABA in the nervous system of the cestodes Diphyllobothrium dendriticum (Diphyllobothriidea) and Caryophyllaeus laticeps (Caryophyllidea), with comparative analysis of muscle innervation.

The nervous system (NS) of the cestodes Diphyllobothrium dendriticum (Diphyllobothriidea) and Caryophyllaeus laticeps (Caryophyllidea) was investigated using immunocytochemistry. The GABA neurotransmitter was identified in the NS of both species; GABAergic neurons were detected in the main nerve cords (MC). GABA-like immunoreactive neurons were predominantly unipolar and exhibited more intensive immunoreactivity in the neurite than in the perikaryon. In C. laticeps , GABA-like immunoreactive somas are located in both the MCs and peripheral NS near the longitudinal muscles. The innervation of the body musculature was studied using a combination of antibodies against GABA, serotonin (5-HT), and FMRFamide and with complementary staining of F-actin. In both species, the location of GABAergic neurites is associated with all muscle layers including the subtegumental, longitudinal, transverse, and dorsoventral muscles. The cytomorphology of 5-HTergic motoneurons in the MCs of both species is described and differences in muscle innervation between D. dendriticum and C. laticeps are demonstrated. No evidence for co-localization of GABA with 5-HT or FMRFamide neurotransmitters at any particular neuron was found. Neuropiles in MCs and peripheral NS had separate sets of immunoreactive neurites. A functional role for GABA in muscle innervation is discussed.

Physical and sensory characteristics of pork sausages from enzymatically modified blends of lard and rapeseed oil during storage.

Physical and sensory characteristic of pork sausages produced from enzymatic interesterified blends of lard and rapeseed oil during storage were evaluated. All three enzymatic interesterified blends (IE90, IE70 and IE50) had ratios of unsaturated to saturated fatty acids within the range of 1.47-2.84 which is favourable for cardiovascular disease risk reduction. Blends of IE90 and IE70 were found to have suitable solid fat content, melting and crystallization profile suitable for sausages production. Sausages were produced from blends of IE90 and IE70 with different muscle types (musculus longissimus dorsi and musculus sternomandibularis) and processing conditions such as cooling rates and final processing temperature. Cooling rate was found to have no significant (P>0.05) effect on hardness of the sausages throughout storage. Both musculus longissimus dorsi and high final processing temperature of 20 degrees C increased the hardness of the sausages during storage. In terms of fat particle size distribution, it was found that sausages IE70 had significantly (P<0.05) lower amount of small fat particles (<4 microm) and higher amount of big fat particles (4-500 microm). This is in agreement with the findings on softer texture of sausages IE70. All the sausages produced from interesterified blends of lard and rapeseed oil had no apparent fat excretion and were rated as having acceptable sensory attributes as compared to reference sausage which was produced from pure lard.