A novel class of amino-alkylcyclohexanes as uncompetitive,fast, voltage-dependent,N-methyl-D-aspartate (NMDA) receptor antagonists – <Emphasis Type="Italic">in vitro</Emphasis> characterization

The fact that potent NMDA receptor channel blockers produce phencyclidine-like psychotropic symptoms in man and rodents implies that uncompetitive antagonism of NMDA receptors may not be a promising therapeutic approach. However, recent data indicate that agents with moderate affinity such as memantine and neramexane (MRZ 2/579) are useful therapeutics due to their strong voltage-dependency and rapid unblocking kinetics. Merz has developed a series of novel uncompetitive NMDA receptor antagonists based on an amino-alkylcyclohexane structure. These compounds displaced [(3)H]-MK-801 binding to rat cortical membranes with K(i) values between 1 and 100 microM and inward current responses of cultured hippocampal neurons to NMDA were antagonized in a strongly voltage-dependent manner with rapid blocking/unblocking kinetics. Three of these compounds, with similar biophysical properties to memantine, were chosen for development. MRZ 2/759 (1-ethenyl-3,3,5,5-tetramethyl-cyclohexylamine), 2/1010 (1,3,3,5-tetramethyl-6-azabicyclo[3.2.1]octane) and 2/1013 (8,8,10,10-tetramethyl-1-azaspiro[5.5] undecane) displaced [(3)H]-MK-801 binding with K(i) values of 1.18, 2.59 and 3.64 microM, respectively. They were similarly potent against NMDA-induced currents in hippocampal neurons - IC(50) values of 1.51, 3.06 and 2.20 microM, respectively. In line with their moderate affinity, all were voltage-dependent (delta = 0.86, 0.96 and 0.89, respectively) and fast, open-channel blockers (k(on) 7.90, 1.70 and 2.60 x 10(4) M(-1) sec(-1), k(off) 0.13, 0.12 and 0.24 sec(-1), respectively). These compounds are also NMDA receptor antagonists in the CNS following systemic administration and have good therapeutic indices in a variety of in vivo behavioural models where glutamate is known to play a pivotal role. In view of their relatively low affinity and associated rapid kinetics, they should prove to be useful therapeutics in a wide range of CNS disorders.     

Variations in activity and inhibition with pH: the protonated amine is the substrate for monoamine oxidase,but uncharged inhibitors bind better

Summary It has been accepted that, as required mechanistically, the neutral form of the amine is the substrate for monoamine oxidase, despite the amine pK _a of above 9.5. The pH dependence of the kinetic parameters for kynuramine oxidation by purified human MAO-A and for phenylethylamine oxidation by MAO-B in granulocytes at pH values from 5 to 10 was consistent with the protonated amine being used. Deprotonation of a group of pK _a = 7.1 in MAO-B and pK _a = 7.5 ± 0.1 (n = 4) in MAO-A was important for efficient catalysis. The K_i values for two oxazolidinone inhibitors of MAO-A gave opposite pH-dependence indicating that the uncharged form of each inhibitor bound better than the charged form. Decreased pH induced a blue shift in the spectral maximum of MAO-A indicative of a more hydrophobic environment around the flavin, and also influenced the redox properties of the flavin.     

GABAergic currents in RT and VB thalamic nuclei follow kinetic pattern of alpha3- and alpha1-subunit-containing GABAA receptors

Inhibitory postsynaptic currents (IPSCs) of the thalamic reticular (RT) nucleus are dramatically slower than in the neighboring ventrobasal (VB) neurons. It has been suggested that alpha3-subunit-containing receptors underlie slow IPSCs in RT neurons, while rapid synaptic currents in the VB nucleus are due to gamma-aminobutyric acid A receptors (GABAARs), including the alpha1-subunit. In our recent study [Barberis et al. (2007) Eur. J. Neurosci., 25, 2726-2740] we have found that profound differences in kinetics of currents mediated by alpha3beta2gamma2 and alpha1beta2gamma3 receptors resulted from distinct binding and desensitization properties. However, a direct comparison between kinetics of neuronal GABAARs from RT and VB neurons and alpha3- and alpha1-subunit-containing receptors has not been made. For this purpose, current responses to ultrafast GABA applications were recorded from patches excised from neurons in VB and RT areas. Deactivation kinetics determined for RT and VB neurons closely resembled that in currents mediated by alpha3beta2gamma2 and alpha1beta2gamma2 receptors. In RT neurons, currents elicited by non-saturating [GABA] had a remarkably slow onset, a hallmark of alpha3-subunit-containing receptors. In VB and RT neurons, single-channel currents elicited by brief GABA pulses had similar characteristics to those of alpha1beta2gamma2 and alpha3beta2gamma2 receptors. However, in stationary conditions, similarity between single-channel currents in neurons and respective recombinant receptors was less apparent. We propose that the non-stationary kinetics of GABAergic currents in VB and RT nuclei mimic that of currents mediated by alpha1- and alpha3-subunit-containing receptors. The dissimilarity between stationary kinetics of neuronal and recombinant receptors probably reflects differences between GABAARs mediating phasic and tonic currents in these neurons.     

交联β-环糊精-可溶性淀粉聚合物微球的合成及载药性质

目的用β-环糊精和可溶性淀粉作为原料合成不溶于水的微球,并对其吸附载药性质进行研究。方法使用反相悬浮交联法制备聚合物微球,对其乳化及交联过程的各个因素进行考察,并且以水杨酸为模型药物,考察了交联反应不同因素对水杨酸载药量的影响。探讨了药物的吸附动力学和吸附热力学。用傅立叶红外色谱仪、粉末X射线衍射、激光衍射粒度分析仪等手段对微球的性质进行研究。结果与结论β-环糊精与可溶性淀粉和环氧氯丙烷的用量比例为9 g∶1 g∶12 mL、氢氧化钠质量分数为40%、水相油相体积比为15∶120、乳化剂用量为7.0 g、搅拌速度为400 r.min-1、60℃下交联8 h可以获得粒径为100μm左右、粒径分布较窄、形状圆整的微球。红外光谱证明β-环糊精和可溶性淀粉在交联后结构和性质发生变化,交联后生成的三维网络结构使其具有较大的机械强度和吸附能力,可以作为药物载体。实验结果表明交联微球对水杨酸的吸附很快,1 h可达到平衡,平衡时包封率达到58.35%,载药质量分数为4.856%,而且主要为物理吸附过程。     

In vitro evaluation of UV opacity potential of <Emphasis Type="Italic">Aloe vera</Emphasis> L. gel from different germplasms

In this study, lyophilized crude and methanolic extracts of aloe gel from different germplasms (S24, RM, TN, OR, and RJN) of Aloe vera L. were tested for their ultraviolet (UV) opacity potential. UV absorption profiles, sun protection factor (SPF), and percentage blocking of UVA and UVB were considered to test UV opacity potential. Both the extracts showed UV absorption and followed the same path in the wavelength range of 250–400 nm in all the germplasms. Methanolic extract showed a stronger absorptivity than the crude lyophilized extract. Among the tested germplasms, maximum UV opacity property with a SPF of 9.97% and 79.12% UVB blocking was obtained with RJN, whereas a poor response was evident in TN with a SPF of 1.37% and 28.5% UVB blocking at 4 mg/ml methanolic extract. To our knowledge the present work for the first time documents UV opacity properties of A. vera L. gel and opens up new vistas in Aloe gel characterization.     

盐酸氯米帕明大鼠在体肠吸收动力学研究

目的研究盐酸氯米帕明在大鼠肠道的吸收特性。方法采用大鼠在体肠灌流实验方法,利用高效液相色谱法同时测定肠回流液中药物及酚红的浓度,通过酚红的浓度校正相应时刻供试液的体积。结果盐酸氯米帕明浓度为5、10、25μg/ml时小肠的吸收速率常数(ka)为(0.74±0.04)、(0.78±0.03)、(0.77±0.05)h-1;在十二指肠、空肠、回肠和结肠的ka分别为(0.590±0.026)、(0.670±0.032)、(0.680±0.030)和(0.560±0.031)h-1。结论盐酸氯米帕明在大鼠肠道均有良好的吸收。盐酸氯米帕明在小肠的吸收呈表观一级动力学过程,吸收机制为被动扩散。     

The phospholipase A2 inhibitor methyl indoxam suppresses diet-induced obesity and glucose intolerance in mice

Background and purpose:

Previous results have shown that mice lacking in the group 1B phospholipase A₂ (Pla2g1b) are resistant to obesity and diabetes induced by feeding a diabetogenic high-fat/high-carbohydrate diet. This study examined the potential of using the Pla2g1b inhibitor methyl indoxam as therapy to suppress diet-induced obesity and diabetes.

Experimental approach:

Male C57BL/6 mice were fed the diabetogenic diet with or without methyl indoxam supplementation. Body weight gain, fasting plasma glucose levels, glucose tolerance and postprandial lysophospholipid absorption were compared.

Key results:

Wild-type C57BL/6 mice fed the diabetogenic diet without Pla2g1b inhibitor showed 31 and 69% body weight gain after 4 and 10 weeks respectively. These animals also showed elevated plasma glucose levels and were glucose intolerant. In contrast, C57BL/6 mice fed the diabetogenic diet with 90 mg·kg⁻¹ of methyl indoxam gained only 5% body weight after 10 weeks. These animals were also euglycaemic and displayed normal glucose excursion rates in glucose tolerance test. Methyl indoxam suppression of diet-induced body weight gain and glucose intolerance was correlated with the inhibition of Pla2g1b-mediated postprandial lysophospholipid absorption.

Conclusions and implications:

These results show that oral supplementation of a diabetogenic diet with the Pla2g1b inhibitor methyl indoxam effectively suppresses diet-induced obesity and diabetes in mice. This suggests that Pla2g1b inhibition may be a potentially effective oral therapeutic option for treatment of obesity and diabetes.     

Evaluation of percutaneous absorption of the repellent diethyltoluamide and the sunscreen ethylhexyl p‐methoxycinnamate‐loaded solid lipid nanoparticles: an in‐vitro study

Objectives Diethyltoluamide and ethylhexyl p‐methoxycinnamate (OMC) are two active ingredients in insect repellent and sunscreen products, respectively. The concurrent application of these two substances often increases their systemic absorption, compromising the safety and efficiency of the cosmetic product. In this study, diethyltoluamide and OMC were incorporated into solid lipid nanoparticles, a colloidal drug delivery system, to reduce percutaneous absorption and avoid toxic effects and also maintain the efficacy of the two active compounds on the skin surface for a long duration. Methods Solid lipid nanoparticles were prepared based on an ultrasonication technique and characterized by differential scanning calorimetry (DSC) analyses. In‐vitro studies determined the percutaneous absorption of diethyltoluamide and OMC. Key findings DSC data carried out on unloaded and diethyltoluamide‐ and/or OMC‐loaded solid lipid nanoparticles highlighted that diethyltoluamide and OMC modified the temperature and the enthalpy change associated to the calorimetric peak of solid lipid nanoparticles. The concurrent presence of the two compounds in the solid lipid nanoparticles caused a synergic effect, indicating that the lipid matrix of nanoparticles guaranteed a high encapsulation of both diethyltoluamide and OMC. Results from the in‐vitro study demonstrated that the particles were able to reduce the skin permeation of the two cosmetic ingredients in comparison with an oil‐in‐water emulsion. Conclusions This study has provided supplementary evidence as to the potential of lipid nanoparticles as carriers for topical administration of cosmetic active compounds.     

Whole‐body physiologically based pharmacokinetic population modelling of oral drug administration: inter‐individual variability of cimetidine absorption

Objectives Inter‐individual variability of gastrointestinal physiology and transit properties can greatly influence the pharmacokinetics of an orally administered drug in vivo. To predict the expected range of pharmacokinetic plasma concentrations after oral drug administration, a physiologically based pharmacokinetic population model for gastrointestinal transit and absorption was developed and evaluated. Methods Mean values and variability measures of model parameters affecting the rate and extent of cimetidine absorption, such as gastric emptying, intestinal transit times and effective surface area of the small intestine, were obtained from the literature. Various scenarios incorporating different extents of inter‐individual physiological variability were simulated and the simulation results were compared with experimental human study data obtained after oral cimetidine administration of four different tablets with varying release kinetics. Key findings The inter‐individual variability in effective surface area was the largest contributor to absorption variability. Based on in‐vitro dissolution profiles, the mean plasma cimetidine concentration–time profiles as well as the inter‐individual variability could be well described for three cimetidine formulations. In the case of the formulation with the slowest dissolution kinetic, model predictions on the basis of the in‐vitro dissolution profile underestimated the plasma exposure. Conclusions The model facilitates predictions of the inter‐individual pharmacokinetic variability after oral drug administration for immediate and extended‐release formulations of cimetidine, given reasonable in‐vitro dissolution kinetics.     

Cell division modulates prion accumulation in cultured cells

The phenotypic effect of prions on host cells is influenced by the physical properties of the prion strain and its level of accumulation. In mammalian cell cultures, prion accumulation is determined by the interplay between de novo prion formation, catabolism, cell division, and horizontal cell-to-cell transmission. Understanding this dynamic enables the analytical modeling of protein-based heritability and infectivity. Here, we quantitatively measured these competing effects in a subline of neuroblastoma (N2a) cells and propose a concordant reaction mechanism to explain the kinetics of prion propagation. Our results show that cell division leads to a predictable reduction in steady-state prion levels but not to complete clearance. Scrapie-infected N2a cells were capable of accumulating different steady-state levels of prions, dictated partly by the rate of cell division. We also show that prions in this subline of N2a cells are transmitted primarily from mother to daughter cells, rather than horizontal cell-to-cell transmission. We quantitatively modeled our kinetic results based on a mechanism that assumes a subpopulation of prions is capable of self-catalysis, and the levels of this subpopulation reach saturation in fully infected cells. Our results suggest that the apparent effectiveness of antiprion compounds in culture may be strongly influenced by the growth phase of the target cells.