人眼微光下夜近视机制的研究

本文利用激光散斑屈光测试系统详细探讨了夜近视(night myopia)的产生机制。夜近视在环境亮度降低到产生暗视觉时出现,并随亮度继续降低而增大,平均值为1.35D。实验证明眼睛球差和色差不是造成夜近视的主要因素。夜近视数值在暗适应过程中呈增大的趋势。在暗视觉时,由于视野中缺少适宜的刺激,眼睛处于暗焦(dark focus)休止状态,这是造成夜近视的主要原因。     

Biochemical and genetic characterization of three hamster cell mutants resistant to diphtheria toxin

We describe here three different hamster cell mutants which are resistant to diphtheria toxin and which provide models for investigating some of the functions required by the toxin inactivates elongation factor 2 (EF-2). Cell-free extracts from mutants Dtx(r)-3 was codominant. The evidence suggests that the codominant phenotype is the result of a mutation in a gene coding for EF-2. The recessive phenotype might arise by alteration of an enzyme which modifies the structure of EF-2 so that it becomes a substrate for reaction with the toxin. Another mutant, Dtx(r)-2, contained EF-2 that was sensitive to the toxin and this phenotype was recessive. Pseudomonas aeruginosa exotoxin is known to inactivate EF-2 as does diphtheria toxin and we tested the mutants for cross-resistance to pseudomonas exotoxin. Dtx(r)-1 and Dtx(r)-3 were cross-resistant while Dtx(r)-2 was not. It is known that diphtheria toxin does not penetrate to the cytoplasm of mouse cells and that these cell have a naturally occurring phenotype of diphtheria toxin resistance. We fused each of the mutants with mouse 3T3 cells and measured the resistance. We fused each of the mutants with mouse 3T3 cells and measured the resistance of the hybrid cells to diphtheria toxin. Intraspecies hybrids containing the genome of mutants Dtx(r)-1 and Dtx(r)-3 had some resistance while those formed with Dtx(r)-2 were as sensitive as hybrids derived from fusions between wild-type hamster cells and mouse 3T3 cells.     

Interaction of three-finger proteins from snake venoms and from mammalian brain with the cys-loop receptors and their models

With the use of surface plasmon resonance (SPR) it was shown that ws-Lynx1, a water-soluble analog of the three-finger membrane-bound protein Lynx1, that modulates the activity of brain nicotinic acetylcholine receptors (nAChRs), interacts with the acetylcholine-binding protein (AChBP) with high affinity, K_D = 62 nM. This result agrees with the earlier demonstrated competition of ws-Lynx1 with radioiodinated α-bungarotoxin for binding to AChBP. For the first time it was shown that ws-Lynx1 binds to GLIC, prokaryotic Cys-loop receptor (K_D = 1.3 μM). On the contrary, SPR revealed that α-cobratoxin, a three-finger protein from cobra venom, does not bind to GLIC. Obtained results indicate that SPR is a promising method for analysis of topography of ws-Lynx1 binding sites using its mutants and those of AChBP and GLIC.     

alpha-Neurotoxins and alpha-conotoxins--nicotinic cholinoreceptor blockers

The review is devoted to the competitive blockers of different nicotinic acetylcholine receptors, alpha-neurotoxins from snake venoms, and alpha-conotoxins from marine snails of the Conidae family. The relationship between the structure and function of these toxins is discussed. Recent data on the mechanism of alpha-neurotoxin and alpha-conotoxin interaction with the nicotinic acetylcholine receptor are presented.     

Study of solid-phase catalytic isotopic exchange of hydrogen in alpha-conotoxin G1 under the effect of spillover-tritium

Tritium-labeled alpha-conotoxin G1 with a molar radioactivity of 35 Ci/mmol and full biological activity (according to the binding to nicotinic acetylcholine receptor) was obtained by the high-temperature solid-state catalytic isotope exchange (HSCIE). The tritium distribution in the molecule of alpha-conotoxin G1 was revealed by 3H NMR spectroscopy. Tritium was found in all amino acid residues except for the Asn4-Pro5-Ala6 fragment. The data on the comparative reactivity of C-H bonds, the ab initio quantum-chemical calculation of the hydrogen exchange reaction, and the information on the spatial structures of alpha-conotoxin G1 in solution and in crystal state allowed us to establish that the reactivity of H atoms may be increased by their interaction with the electron donor O and N atoms at the transition state of the HSCIE reaction. A decrease in the rate of the HSCIE reaction could be caused by both a poor spatial accessibility of C-H bonds and a limited mobility of the peptide fragment containing these bonds.     

Azemiopsin from Azemiops feae viper venom, a novel polypeptide ligand of nicotinic acetylcholine receptor

Azemiopsin, a novel polypeptide, was isolated from the Azemiops feae viper venom by combination of gel filtration and reverse-phase HPLC. Its amino acid sequence (DNWWPKPPHQGPRPPRPRPKP) was determined by means of Edman degradation and mass spectrometry. It consists of 21 residues and, unlike similar venom isolates, does not contain cysteine residues. According to circular dichroism measurements, this peptide adopts a β-structure. Peptide synthesis was used to verify the determined sequence and to prepare peptide in sufficient amounts to study its biological activity. Azemiopsin efficiently competed with α-bungarotoxin for binding to Torpedo nicotinic acetylcholine receptor (nAChR) (IC(50) 0.18 ± 0.03 μm) and with lower efficiency to human α7 nAChR (IC(50) 22 ± 2 μm). It dose-dependently blocked acetylcholine-induced currents in Xenopus oocytes heterologously expressing human muscle-type nAChR and was more potent against the adult form (α1β1εδ) than the fetal form (α1β1γδ), EC(50) being 0.44 ± 0.1 μm and 1.56 ± 0.37 μm, respectively. The peptide had no effect on GABA(A) (α1β3γ2 or α2β3γ2) receptors at a concentration up to 100 μm or on 5-HT(3) receptors at a concentration up to 10 μm. Ala scanning showed that amino acid residues at positions 3-6, 8-11, and 13-14 are essential for binding to Torpedo nAChR. In biological activity azemiopsin resembles waglerin, a disulfide-containing peptide from the Tropidechis wagleri venom, shares with it a homologous C-terminal hexapeptide, but is the first natural toxin that blocks nAChRs and does not possess disulfide bridges.     

Photoactivatable alpha-conotoxins reveal contacts with all subunits as well as antagonist-induced rearrangements in the Torpedo californica acetylcholine receptor.

Azidobenzoyl (AzBz) and benzoylbenzoyl (BzBz) derivatives of alpha-conotoxin MI and L-benzoylphenylalanine (Bpa) analogs of alpha-conotoxin GI were synthesized. All these compounds, similarly to native alpha-conotoxins, completely displaced the radioiodinated MI or GI from the membrane-bound nicotinic acetylcholine receptor (AChR) of Torpedo californica. However, the GI(Bpa11) analog was considerably less potent than GI in competing with radioiodinated alpha-bungarotoxin (alphaBgt). Irradiation of iodinated AzBz derivatives bound to AChR resulted in labeling of all AChR subunits. The BzBz and Bpa derivatives gave lower levels of specific cross-linking but considerable labeling at additional sites that was enhanced, rather than suppressed, by an excess of native alpha-conotoxins or alphaBgt. Both equilibrium binding of benzophenone-derivatized alpha-conotoxins and their cross-linking could be totally abolished by physostigmine. The results obtained demonstrate that (a) specific binding sites for alpha-conotoxins and alphaBgt are overlapping but not identical, (b) each of the AChR subunits can be labeled with photoactivatable alpha-conotoxins and (c) enhancement of benzophenone-derivatized alpha-conotoxins cross-linking at additional (physostigmine-related) sites by alphaBgt or GI indicates that these antagonists induce structural alterations in the AChR outside their binding sites.     

ANTIPYRETIC ACTION OF DEXAMETHASONE ON EGTAZIC ACIDINDUCED FEVER IN RABBITS

本文用脑室灌注和Ｆｕｒａ２测定细胞内游离钙技术观察了地塞米松（ｄｅｘａｍｅｔｈａｓｏｎｅ,ＤＥＸ）对家兔乙二醇双（２氨基乙醚）四乙酸性发热效应和下丘脑细胞内游离钙浓度（［Ｃａ２＋］ｉ）的影响,借此深入探讨地塞米松解热作用的中枢机制。结果发现：脑室灌注乙二醇双（２氨基乙醚）四乙酸（０６ｎｍｏｌ）引起家兔结肠温度明显升高,静脉注射地塞米松（５ｍｇ／ｋｇ）显著抑制家兔乙二醇双（２氨基乙醚）四乙酸性发热,地塞米松（６０～１２０μｍｏｌ／Ｌ）并不影响下丘脑细胞内［Ｃａ２＋］ｉ,而事先脑室灌注抑制基因转录的放线菌素Ｄ（３ｎｍｏｌ）则完全取消了地塞米松对乙二醇双（２氨基乙醚）四乙酸性发热的解热作用。这些结果提示：地塞米松显著抑制家兔乙二醇双（２氨基乙醚）四乙酸性发热,其机制与地塞米松激活脑内某些基因的表达有关,而与下丘脑神经细胞跨膜钙离子流无关。     

Instant success for Instant Pharmacology

Instant Pharmacology (1999). Saeb-Parsy K, Assomull RG, Kahn FZ, Saeb-Parsy K, and Kelly E. John Wiley and Sons 349 pp. £19.99 pbk; ISBN 0471976393 © 1999 John Wiley & Sons, Inc.     

Interaction of α-conotoxin ImII and its analogs with nicotinic receptors and acetylcholine-binding proteins: additional binding sites on Torpedo receptor

α-Conotoxins interact with nicotinic acetylcholine receptors (nAChRs) and acetylcholine-binding proteins (AChBPs) at the sites for agonists/competitive antagonists. α-Conotoxins blocking muscle-type or α7 nAChRs compete with α-bungarotoxin. However, α-conotoxin ImII, a close homolog of the α7 nAChR-targeting α-conotoxin ImI, blocked α7 and muscle nAChRs without displacing α-bungarotoxin ( Ellison et al. 2003, 2004 ), suggesting binding at a different site. We synthesized α-conotoxin ImII, its ribbon isomer (ImII iso ), 'mutant' ImII(W10Y) and found similar potencies in blocking human α7 and muscle nAChRs in Xenopus oocytes. Both isomers displaced [¹²⁵I]-α-bungarotoxin from human α7 nAChRs in the cell line GH₄C₁ (IC₅₀ 17 and 23 μM, respectively) and from Lymnaea stagnalis and Aplysia californica AChBPs (IC₅₀ 2.0–9.0 μM). According to SPR measurements, both isomers bound to immobilized AChBPs and competed with AChBP for immobilized α-bungarotoxin ( K _d and IC₅₀ 2.5–8.2 μM). On Torpedo nAChR, α-conotoxin [¹²⁵I]-ImII(W10Y) revealed specific binding ( K _d 1.5–6.1 μM) and could be displaced by α-conotoxin ImII, ImII iso and ImII(W10Y) with IC₅₀ 2.7, 2.2 and 3.1 μM, respectively. As α-cobratoxin and α-conotoxin ImI displaced [¹²⁵I]-ImII(W10Y) only at higher concentrations (IC₅₀≥ 90 μM), our results indicate that α-conotoxin ImII and its congeners have an additional binding site on Torpedo nAChR distinct from the site for agonists/competitive antagonists.