光遗传学技术对抑郁症的研究应用

光遗传学(optogenetics)技术是近年来快速发展的一项生物工程技术,可以对神经元进行精确控制。本文就现有抑郁症治疗的不足做了分析与总结,介绍了光遗传学技术在抑郁症研究应用方面的新进展,提出抑郁症相关行为表现与VTA多巴胺神经元的放电模式、投射通路的特异性、压力类型和压力严重程度之间存在相关联系。     

氚标舒必利放射配基结合分析法的建立

研究了氚标舒必利([~3H]-Sulpiride)与大鼠纹状体膜蛋白的结合特性,结果表明两者结合具有特异性、饱和性、可逆性和配体选择性,B_(max)=917.0±133.2pmol/g蛋白,K_d=5.17±1.13nmol/l;并对根据这一结合特性建立的[~3H]-Sulpiride放射配基结合分析法作为测定血中舒必利浓度的方法进行分析,该方法的线性范围是1—100nmol/l,灵敏度是49.25ng/l,批内、批间变异系数分别为3.35％和10.6％。     

多巴胺D2受体的同源模建研究

多巴胺是大脑中含量最丰富的儿茶酚胺类神经递质,主要通过多巴胺受体调控中枢神经系统的多种生理功能,其中多巴胺D2受体与药物成瘾、精神分裂症、帕金森病等多种疾病的发生相关。然而多巴胺D2受体的晶体结构至今尚未解析出来,给相关疾病的药物设计与开发带来困难。本文采用同源模建的方法,用目前与多巴胺D2受体同源性最高的多巴胺D3受体(3PBL)作为模板,构建多巴胺D2受体的三维结构。经过优化和分子动力学模拟,用Profile-3D和Ramachandran plot对模型进行评估,然后用多巴胺D2受体拮抗剂千金藤啶碱(stepholidine,SPD)进行对接验证,证明构建的多巴胺D2受体模型合理、可靠。     

Noble metal dispersed multiwalled carbon nanotubes immobilized ss-DNA for selective detection of dopamine

A hybrid nanocomposite consisting of Pt nanoparticles decorated functionalized multiwalled carbon nanotubes (f-MWNT) immobilized with single strand-DNA (ss-DNA) has been devised for the selective detection of dopamine (DA). AFM and TEM analyses show that wrapping of ss-DNA over Pt/f-MWNT reduces the aggregation of the nanotubes arising from van der Waals interaction. In addition to serving as a noncovalent dispersion agent, ss-DNA facilitated electron transfer towards dopamine, as analyzed by cyclic voltammetric studies (CV) and amperometry. The sequence dependency of ss-DNA for DA detection has been analyzed using AC and GT ss-DNA. The hybrid nanocomposite biosensor consisting of AC/ss-DNA exhibits linearity of detection upto ∼315 μM, with a detection limit 0.8 μM towards dopamine. The best sensing performance with linearity of ∼800 μM and detection limit ∼0.45 μM has been obtained with GT/ss-DNA immobilized Pt/f-MWNT sensor. Further, the nafion coated ss-DNA wrapped Pt/f-MWNT immobilized biosensor exhibits good stability, fast response time (<3 s) and selective detection of DA in the presence of ascorbic acid and uric acid.     

Evaluation of the transdermal permeation behavior of Proterguride from drug in adhesive matrix patches through hairless mouse skin

The transdermal in vitro permeation behavior of the highly potent dopamine agonist Proterguride was investigated using hairless mouse skin as a model membrane. Drug in adhesive matrix formulations based on different types of pressure-sensitive adhesives (Eudragit® E 100 and Gelva®7883 as acrylates, Oppanol® B 15 SFN as polyisobutylene, and BioPSA® 7-4202 as silicone) with a drug load of 3% by weight were manufactured. All patches were examined for drug crystallization by polarized microscopy immediately after the manufacturing process and after storage for 30 days in sealed aluminium laminate bags at ambient temperature and at 40°C, respectively. Furthermore, the influence of the drug load in acrylate-based formulations onto the steady-state flux of Proterguride was examined. The Eudragit® E 100 system delivered a significantly higher steady-state flux than the systems based on Oppanol® B 15 SFN and also a somewhat higher steady-state flux than the Gelva®-based patch. An addition of 10% by weight of the crystallization inhibitor povidone 25 did not significantly influence the steady-state flux of Proterguride from acrylate matrices. The lipophilic silicone and polyisobutylene adhesives facilitated drug crystallization within the short storage periods at both conditions, probably due to the absence of povidone 25, which was incompatible with these polymers. Varying the drug load in acrylate-based formulations led to a linear increase of the steady-state flux until the steady-state flux of Proterguride leveled off and the patches tended to drug crystallization. It was found that Gelva®-based patches show good physical stability, good skin adhesion, and moderate flux values and, thus, can be evaluated as a basis for a suitable formulation for the transdermal administration of Proterguride.     

Capability of a carbon-polyvinylchloride composite electrode for the detection of dopamine, ascorbic acid and uric acid

In this work, the composite carbon-polyvinylchloride (C-PVC) was used as an electrode for the detection of dopamine, ascorbic acid, uric acid and their mixtures by differential pulse voltammetry (DPV). The results showed that the untreated C-PVC electrode was selective and stable for the oxidation of dopamine in a mixture containing uric acid and an excess of ascorbic acid in acidic medium. The pre-treated C-PVC electrode in a neutral medium exhibited good resolution of the mixture components in the micro molar concentration range of DA. The ageing of the C-PVC electrode during longer time periods did not affect the peak potential and the detection of dopamine, uric acid and ascorbic acid in 0.1 M H₂SO₄. The practical analytical utility of the C-PVC electrode was demonstrated by the measurement of uric acid in human urine and serum samples without any preliminary pre-treatment.     

Polyaniline–polyelectrolyte–gold(0) ternary nanocomposites: Synthesis and electrochemical properties

Ternary composites based on polyaniline (PAni), a polyelectrolyte-namely poly(diallyldimethylammoniumchloride) (PDDMAC) and gold (Au(0)) nanoparticles have been formulated and synthesized where the high concentration of PDDMAC acted as medium of reaction. The nanocomposites are characterized by FT-IR, UV–vis, XRD, XPS, SEM, AFM and TEM techniques. XRD showed the presence of all three viz., polyaniline, PDDMAC and Au(0) components in the ternary system. The composites exhibited higher conductivities in the range 26 × 10⁻⁶ to 217 × 10⁻⁶ S/cm compared with the binary composite of PAni–PDDMAC. The ternary composites were adsorbed on a GC electrode and used for sensing dopamine. The composites are useful in sensing as low as 0.05 mM concentration of dopamine at lower potential values compared to some binary PAni–Au nanocomposites.     

Fabrication of a PANI/Au nanocomposite modified nanoelectrode for sensitive dopamine nanosensor design

Polyaniline/Au nanocomposite modified nanoelectrodes based dopamine nanosensors have been developed. The polyaniline/Au nanocomposite film was deposited at the exposed end of the nanoelectrode tip by a surface-graft polymerization method to fabricate a desired modified nanoelectrode. With this modified method, the nanocomposites firmly adhered on the electrode surface and the modified nanoelectrode still had a sharp tip, which was proved by the scanning electron microscope. The electrochemical measurement shows the polyaniline/Au nanocomposite modified film has a good and stable redox activity in neutral solution. The modified nanoelectrode exhibits the excellent electrocatalytic activity towards the oxidations of ascorbic acid and dopamine in phosphate buffer solution. The separation of anodic peak potential of dopamine and ascorbic acid reaches 250 mV. Differential pulse voltammograms results illustrate that dopamine can be selectively determined in the presence of thousands times higher concentration of ascorbic acid with a wide linear range from 200 to 0.3 μM and the detection limit is 0.1 μM. This study provides a simple method for the construction of dopamine nanosensors that have a good sensitivity, wide linear range and stable response. The nanosensors are hopeful to be applied to the detection of dopamine in vivo.     

Evaluation of the Transdermal Permeation Behavior of Proterguride from Drug in Adhesive Matrix Patches Through Hairless Mouse Skin

The transdermal in vitro permeation behavior of the highly potent dopamine agonist Proterguride was investigated using hairless mouse skin as a model membrane. Drug in adhesive matrix formulations based on different types of pressure-sensitive adhesives (Eudragit® E 100 and Gelva®7883 as acrylates, Oppanol® B 15 SFN as polyisobutylene, and BioPSA® 7-4202 as silicone) with a drug load of 3% by weight were manufactured. All patches were examined for drug crystallization by polarized microscopy immediately after the manufacturing process and after storage for 30 days in sealed aluminium laminate bags at ambient temperature and at 40°C, respectively. Furthermore, the influence of the drug load in acrylate-based formulations onto the steady-state flux of Proterguride was examined. The Eudragit® E 100 system delivered a significantly higher steady-state flux than the systems based on Oppanol® B 15 SFN and also a somewhat higher steady-state flux than the Gelva®-based patch. An addition of 10% by weight of the crystallization inhibitor povidone 25 did not significantly influence the steady-state flux of Proterguride from acrylate matrices. The lipophilic silicone and polyisobutylene adhesives facilitated drug crystallization within the short storage periods at both conditions, probably due to the absence of povidone 25, which was incompatible with these polymers. Varying the drug load in acrylate-based formulations led to a linear increase of the steady-state flux until the steady-state flux of Proterguride leveled off and the patches tended to drug crystallization. It was found that Gelva®-based patches show good physical stability, good skin adhesion, and moderate flux values and, thus, can be evaluated as a basis for a suitable formulation for the transdermal administration of Proterguride.     

Comparison of glassy carbon and boron doped diamond electrodes: Resistance to biofouling

Carbon based electrodes are widely used for in vivo and in vitro electrochemical studies. In particular, monoamine neurochemistry has been investigated using carbon microfibre electrodes. Similarly, glassy carbon (GC) is the preferred material for many biochemical applications, such as electrochemical detection in chromatography. More recently, boron doped diamond (BDD) has been utilized for biosensing, as its carbon sp³ structure is expected to provide better resistance to analyte fouling. However, the main factor limiting the use of electrochemical sensors for biological studies is the effect of the biological matrix. Indeed, in vivo or in situ measurements expose the sensor to a complex matrix of proteins, which adsorb on the sensing surface and interfere with the electrochemical measurements.Here, we compare the performance of three carbon based electrodes: GC, GC with low surface oxides and BDD. The redox species ruthenium(III) hexaammine (outer-sphere), ferrocyanide (surface sensitive) and the biologically significant dopamine have been investigated in protein and blood-mimicking matrices. Cyclic voltammetry and electrochemical impedance spectroscopy have been used to examine the effect of spectator molecules and reaction products on electrode mechanisms.Our results show that BDD generally exhibits the best performance for most conditions and reactions and should therefore be preferred for measurements in biologically fouling environments. Furthermore, surface oxides seem also to improve resistance of the GC electrode to biofouling.