Inactive Membrane Research Articles

Predictors of stoppage and recurrence of choroidal neovascularization with a Treat- Extend-Stop protocol: 4-year follow-up

To analyze the long-term results of treatment of active age-related macular degeneration (AMD) with anti-vascular endothelial growth factor (VEGF) agents using the treat-extend-stop (TES) approach, defining predictive factors for stoppage of the treatment and recurrences in a real-world setting. Data from 191 eyes treated with intravitreal injections for choroidal neovascularization due to AMD were retrospectively reviewed. Changes in best-corrected visual acuity (BCVA) and membrane activity (optical coherence tomography) were recorded and evaluated over a 48-month follow-up. Logistic regression analysis was used to determine predictors of treatment stoppage and recurrences after stoppage. BCVA improvement was found in 70.5% of eyes at 48 months, and remaining signs of activity in 27.9%. Disease inactivity was achieved in 69 eyes (31.9%), with a relapse of the membrane in 29 of these eyes (42.0%). Significant independent predictors of treatment stoppage were found: no foveal membrane, inactive membrane at 12, 24, 36 and 48 months, extension interval>8 weeks at 12 and 24 months,>15 injections at 24 months, and baseline BCVA>61 letters. Concerning recurrent membranes, only the presence of membrane activity at 36 months and baseline BCVA>61 letters were independent predictors. Anti-VEGF treatment of AMD using the TES protocol allows for successful visual restoration in most patients, with more likely disease inactivity in those eyes with better baseline BCVA, maintaining signs of membrane inactivity during the first two years of follow-up and requiring fewer injections.

Использование электролизных процессов в безреагентной водоочистке: удаление сероводорода, органического железа, синтетических поверхностно-активных веществ

The use of electrotechnical equipment refers to reagent-free methods and makes it possible to simplify the automation schemes for facilities for removing pollutants from aqueous solutions. The article substantiates the approaches and structural diagrams of a reagent-free water purification unit based on the use of electrolysis processes. During the research, the processes of oxidation of pollutants were analyzed when a block of electrochemical action on aqueous solutions was used during processing. A diaphragm cell with an inactive membrane volume of 3.5 liters was used to implement the appropriate reagent-free method for removing pollutants from aqueous solutions. The solution of scientific and practical problems is shown: removal of hydrogen sulfide from water from a well before feeding it to the technological lines of a dairy plant; purification of waste water from an enterprise for the production of gelatinous products from organic iron and related pollutants; reduction of waste water pollutants of a cosmetology enterprise, with a focus on destructive effects on synthetic surfactants. Based on the analysis of the results and treatment modes of three aqueous solutions, the structure of the block of reagent-free electrolysis treatment of aqueous solutions is substantiated. As a result of experimental studies, the prospects of electrical equipment have been confirmed. The reagent-free method will simplify the automation schemes for facilities for removing pollutants from aqueous solutions - accordingly, increase the reliability of such computer-integrated solutions. Further research should be directed to the substantiation, creation and parameterization of mathematical models of combined electrotechnological water treatment complexes. Key words: environmental safety, electrotechnological complex, waste water, reagent-free method, electrolysis processes

Комбинированный электротехнологический комплекс очистки сточных вод бумажно-картонного производства

At the moment, complex and unified solutions have not been created, with the intensification of processes and control systems in accordance with integral resource-efficient criteria, which increases the urgency of the task of developing a technological basis for a specialized combined electrotechnological complex for water treatment. The aim of the research is to substantiate the structure and parameters of the combined electrotechnological processes of wastewater treatment in paper and cardboard production. When treating wastewater, electrocoagulation processes were used, characterized by the electrochemical dissolution of metal anodes. Also, during the processing, chemical coagulation was used, which is based on the addition of active coagulants to the wastewater: ammonium, copper, and iron salts. To accelerate the precipitation processes, after coagulation, a biocidal flocculant was used, after which the clarified solution was passed through a diaphragm electrolyzer with an inactive membrane. Also, wastewater was subjected to ozonation for the purpose of additional oxidation of pollutants. To mix the coagulant and effluents, we used an atmospheric air supply. After flotation coagulation, the wastewater was separated by sedimentation and filtered, if necessary, through a polystyrene foam filter. After evaluating the results of experimental wastewater treatment of paper and cardboard production, a combined scheme of an electrotechnological water treatment complex is proposed. Calculation of technical and economic indicators showed that the financial costs of processing glue and dye washes using a combined electrotechnical complex are almost six times less than the services of specialized companies for the disposal of such wastewater. Disposal of untreated aqueous solutions of paper and cardboard production without purification is unacceptable - from the point of view of their possible critical anthropogenic load on the environment in such cases.

Structure of human DPEP3 in complex with the SC-003 antibody Fab fragment reveals basis for lack of dipeptidase activity

Dipeptidase 3 (DPEP3) is one of three glycosylphosphatidylinositol-anchored metallopeptidases potentially involved in the hydrolytic metabolism of dipeptides. While its exact biological function is not clear, DPEP3 expression is normally limited to testis, but can be elevated in ovarian cancer. Antibody drug conjugates targeting DPEP3 have shown efficacy in preclinical models with a pyrrolobenzodiazepine conjugate, SC-003, dosed in a phase I clinical trial (NCT02539719). Here we reveal the novel atomic structure of DPEP3 alone and in complex with the SC-003 Fab fragment at 1.8 and 2.8 Å, respectively. The structure of DPEP3/SC-003 Fab complex reveals an eighteen-residue epitope across the DPEP3 dimerization interface distinct from the enzymatic active site.DPEP1 and DPEP3 extracellular domains share a conserved, dimeric TIM (β/α)8-barrel fold, consistent with 49% sequence identity. However, DPEP3 diverges from DPEP1 and DPEP2 in key positions of its active site: a histidine to tyrosine variation at position 269 reduces affinity for the β zinc and may cause substrate steric hindrance, whereas an aspartate to asparagine change at position 359 abolishes activation of the nucleophilic water/hydroxide, resulting in no in vitro activity against a variety of dipeptides and biological substrates (imipenem, leukotriene D4 and cystinyl-bis-glycine). Hence DPEP3, unlike DPEP1 and DPEP2, may require an activating co-factor in vivo or may remain an inactive, degenerate enzyme.This report sheds light on the structural discriminants between active and inactive membrane dipeptidases and provides a benchmark to characterize current and future DPEP3-targeted therapeutic approaches.

Photografting Graphene Oxide to Inert Membrane Materials to Impart Antibacterial Activity

Surface modification with bactericides is a promising approach to imparting membrane materials with biofouling resistance. However, chemical modification of membranes made from inert materials, such as polyvinylidene fluoride (PVDF) and polysulfone, is challenging because of the absence of reactive functional groups on these materials. In this study, we develop a facile procedure using benzophenone as an anchor to graft biocidal graphene oxide (GO) to chemically inactive membrane materials. GO nanosheets are first functionalized with benzophenone through an amide coupling reaction. Then, benzophenone-functionalized GO nanosheets are irreversibly grafted to the inert membrane surfaces via benzophenone-initiated cross-linking under ultraviolet irradiation. The binding of GO to the membrane surface is confirmed by scanning electron microscopy and Raman spectroscopy. When exposed to a model bacterium (Escherichia coli), GO-functionalized PVDF and polysulfone membranes exhibit strong antibacterial activity, re...

Determining the Composition of Active Cholesterol in the Plasma Membrane of Single Cells by using Electrochemiluminescence

AbstractActive and inactive cholesterol in the plasma membrane of one cell were co‐measured by using luminol electrochemiluminescence, so that the composition of active membrane cholesterol was determined at single cells. The cellular plasma membrane remained intact after the removal of active membrane cholesterol and, thus, inactive cholesterol was activated for reaction with cholesterol oxidase. By utilizing the reactions of active and inactive cholesterol with oxidase in series, both of them generated hydrogen peroxide, inducing the corresponding electrochemiluminescence for the respective quantitation of these two states of membrane cholesterol. The successful determination of active and inactive membrane cholesterol at one cell provides information about the composition of active membrane cholesterol, which is significant for the study of cellular cholesterol trafficking.

An in vivo EGF receptor localization screen in C. elegans Identifies the Ezrin homolog ERM-1 as a temporal regulator of signaling.

The subcellular localization of the epidermal growth factor receptor (EGFR) in polarized epithelial cells profoundly affects the activity of the intracellular signaling pathways activated after EGF ligand binding. Therefore, changes in EGFR localization and signaling are implicated in various human diseases, including different types of cancer. We have performed the first in vivo EGFR localization screen in an animal model by observing the expression of the EGFR ortholog LET-23 in the vulval epithelium of live C. elegans larvae. After systematically testing all genes known to produce an aberrant vulval phenotype, we have identified 81 genes regulating various aspects of EGFR localization and expression. In particular, we have found that ERM-1, the sole C. elegans Ezrin/Radixin/Moesin homolog, regulates EGFR localization and signaling in the vulval cells. ERM-1 interacts with the EGFR at the basolateral plasma membrane in a complex distinct from the previously identified LIN-2/LIN-7/LIN-10 receptor localization complex. We propose that ERM-1 binds to and sequesters basolateral LET-23 EGFR in an actin-rich inactive membrane compartment to restrict receptor mobility and signaling. In this manner, ERM-1 prevents the immediate activation of the entire pool of LET-23 EGFR and permits the generation of a long-lasting inductive signal. The regulation of receptor localization thus serves to fine-tune the temporal activation of intracellular signaling pathways.

Construction of an artificial cell membrane anchor using DARC as a fitting for artificial extracellular functionalities of eukaryotic cells

The need to functionalize cell membranes in a directed way for specific applications as single cell arrays or to force close cell-to-cell contact for artificial intercellular interaction and/or induction concerning stem cell manipulation or in general to have a tool for membrane and cell surface-associated processes, we envisaged a neutral inactive membrane anchor for extracellular entities to facillitate the above mentioned functionalities.The silent Duffy antigen/receptor for chemokines (DARC) is a receptor-like membrane protein of erythrocytes and mediates no cell transduction not at least regarding a missing or truncated G-loop and therefore it seemed to be the candidate for our cell membrane anchor.We isolated the genetic information of DARC from human genomic DNA and cloned it in a mammalian cell line as a fusion protein via a suitable plasmid vector.In this report we demonstrate that the human plasma membrane protein DARC can be used as an artificial anchor molecule in cell surface engineering applications. We constructed the fusion protein SNAP-tag-DARC, consisting of DARC and the self-labeling protein tag SNAP-tag® (Covalys). The SNAP-tag® served as an example for a molecular-technological developed protein that is artificially attached to the extracellular side of the plasma membrane through our DARC-anchor. SnapTag should serve as an example for any extracellular entity and was easy to detect by a commercial detection system. The synthesis of SNAP-tag-DARC, its correct incorporation into the cell membrane and the functionality of the SNAP-tag® were verified by RT-PCR, Western blotting and confocal fluorescence microscopy and showed the desired functionality as an membrane anchor for an extracellular application entity.

Inhibition of Brain-Derived Neurotrophic Factor/Tyrosine Kinase B Signaling Suppresses Choriocarcinoma Cell Growth

Brain-derived neurotrophic factor (BDNF) signals through its receptor tyrosine kinase (Trk)B to regulate the development trophoblast cells during peri- and postimplantation periods. Possible involvement of this signaling system in malignant human trophoblastic cell growth has not been investigated. Here, we found the expression of BDNF and neurtropin-4/5 together with TrkB in human trophoblastic choriocarcinoma cells. Treatment of cultured choriocarcinoma cells with a soluble TrkB ectodomain or a Trk receptor inhibitor K252a suppressed cell proliferation and increased apoptosis associated by the disruption of mitochondrial functions, whereas an inactive plasma membrane nonpermeable K252b was ineffective. Studies using these specific inhibitors also indicated the importance of the phosphatidylinositol 3-kinase and ERK pathways in mediating BDNF actions. Based on PCR array analyses to identify changes in expression profiles of cell cycle- and apoptosis-related genes in cultured choriocarcinoma cells, we found that suppression of endogenous TrkB signaling led to decreases in key proproliferation cell cycle genes and increases in two inhibitory cell cycle genes together with the up-regulation of several proapoptotic genes. In vivo studies in athymic nude mice bearing choriocarcinoma cell tumors further demonstrated that treatment with K252a, but not K252b, suppressed tumor growth accompanied by decreased cell proliferation, reduced levels of a tumor marker, human chorionic gonadotropin-beta, and increased levels of apoptosis and caspase-3/7 activities. Thus, autocrine signaling of the BDNF/TrkB system is important for human choriocarcinoma cell growth, and inhibition of BDNF/TrkB signaling in these cells could provide a novel therapy for patients with choriocarcinoma.

Atomic Force Microscopy Study of Different Effects of Natural and Semisynthetic β-Lactam on the Cell Envelope of Escherichia coli

The effects of native and semisynthetic beta-lactam on the surface of E. coli cells were investigated using tapping mode atomic force microscopy (AFM). These studies were done on both single cell and cell colonies. This allowed us to achieve a complete understanding of the effects, since previous related AFM analysis was only performed on single cells. Our results indicated that although amoxycillin and penicillin could both induce nanoporous damage to the envelope of E. coli, the distribution of the pores was different: those induced by penicillin were randomly distributed on the cell surface, while those induced by amoxycillin were far more numerous and mainly on the two ends of the cell. These findings could explain why the effect of amoxycillin is stronger than that of native penicillin. Ofloxacin was used as a control due to its inactive membrane, and no cell wall damages were observed. Our results demonstrate that AFM is a useful tool in discerning and verifying antibiotic mechanisms and can be helpful to explain the relationship between chemical structure and the function of antibiotics.

Keren, a new ligand of the Drosophila epidermal growth factor receptor, undergoes two modes of cleavage.

Spitz (Spi) is the most prominent ligand of the Drosophila EGF receptor (DER). It is produced as an inactive membrane precursor which is retained in the endoplasmic reticulum (ER). To allow cleavage, Star transports Spi to the Golgi, where it undergoes cleavage by Rhomboid (Rho). Since some DER phenotypes are not mimicked by any of its known activating ligands, we identified an additional ligand by database searches, and termed it Keren (Krn). Krn is a functional homolog of Spi since it can rescue the spi mutant phenotype in a Rho- and Star-dependent manner. In contrast to Spi, however, Krn also possesses a Rho/Star-independent ability to undergo low-level cleavage and activate DER, as evident both in cell culture and in flies. The difference in basal activity correlates with the cellular localization of the two ligands. While Spi is retained in the ER, the retention of Krn is only partial. Examining Spi/Krn chimeric and deletion constructs implicates the Spi cytoplasmic domain in inhibiting its basal activity. Low-level activity of Krn calls for tightly regulated expression of the Krn precursor.

人工膜と生体膜の接点を探る―膜構造と機能発現 構造形成のメカニズム「人工膜」

I intended to correlate the structure of a biological membrane and that of the artificial membranes which had been employed widely in industrial fields. Two types of the membranes such as an ion-exchange membrane and an inactive membrane against ions were manufactured by the application of the common phenomena named as the micro-phase separation. This gave rise to the similar supermolecular structure for the resultant membranes : The former and latter ones were composed of the ion-cluster particles with ca. 5 nm in size and the primary particles with 10-15 nm, respectively. Although the affinity or chemical potential governed the formation of the structure of the membrane after the phase separation, the interfacial force working between the particles and the medium affected on the formation of the orientation of molecular chians at the surface. When we pay an attention to the similarity in the structure between both membranes of artificial and biological ones, the common driving forces for the formation of the membrane structure may be realized.

Inactive membrane protein kinase Cs: a possible target for receptor signalling.

The activation of the multifunctional cell signalling enzymes, the protein kinase Cs (PKCs), is generally thought to result from the translocation of inactive cytosolic enzymes to activation sites in cell membranes. However, recent studies suggest that PKCs may also be stimulated in cells by processes independent of translocation. One possible mechanism is the modulation of the activity of PKCs already resident in membranes. A PKC assay that measures enzyme activity directly in isolated native membranes has revealed the presence of an activatable pool of PKCs resident in native membranes of various cells and tissues. In 3T3-L1 cells, some or all of this pool of membrane PKCs was stimulated within 10 min of exposing the cells to 10 ng/ml epidermal growth factor or 100 ng/ml fibroblast growth factor. Similar increases in PKC activity were observed in native membranes isolated from CTLL-2, WEHI-231 and S49 lymphoma cells that had been exposed to interleukin-2. These growth factors all stimulated membrane PKC activity without detectably translocating cytosolic enzymes to the membranes. In intact WEHI cells, low concentrations (5-10 microM) of a diacylglycerol, 1-oleoyl-2-acetyl-sn-glycerol (OAG), or low concentrations (2-10 nM) of phorbol 12-myristate 13-acetate sufficed to activate PKCs already resident in membranes, but much higher concentrations (50-100 microM and 50-100 nM respectively) were needed to detectably stimulate the translocation of cytosolic PKCs. A phosphatidylcholine-specific phospholipase C also selectively stimulated membrane PKCs in WEHI cells at concentrations that were much less than those needed to induce the translocation of cytosolic enzymes. Furthermore, interleukin-2 and low concentrations of OAG both stimulated the phosphorylation of the 85 kDa PKC-selective substrate protein in intact WEHI cells in which translocation of PKCs was not evident. These results suggest that the membranes of some cells maintain a pool of activatable PKCs that respond to lower levels of extracellular stimuli than cytosolic PKCs, and that can be stimulated by signals which produce diacylglycerols through the hydrolysis of phospholipids other than polyphosphoinositides.

Evidence that the activation of an inactive pool of membrane-associated protein kinase C is linked to the IL-2-dependent survival of T lymphocytes.

The activation of protein kinase C (PKC) is believed to result from the translocation of inactive cytosolic enzyme to the lipid environment of membranes. However, by using a novel method for measuring PKC activity directly in isolated membranes, we have previously shown that a significant proportion of the PKC present in a variety of cells associates with membranes in an inactive state, and that this pool of inactive PKC can be stimulated specifically in cells in the absence of translocation. IL-2 did not stimulate the translocation of PKC to membranes in the IL-2-dependent mouse T cell line, CTLL-2. Nevertheless, a transient, two approximately threefold increase in membrane PKC activity was observed within 10 min of IL-2 addition to these cells. This increase was entirely caused by the activation of a pool of inactive membrane PKC previously associated with the membrane. The inhibition of PKC activity by the specific inhibitors bisindolylmaleimide (BIS) and 1-O-hexadecyl-2-O-methyl-rac-glycerol (AMG) blocked the ability of IL-2 to suppress the onset of apoptosis in IL-2 and serum-deprived CTLL-2 cells. The inhibition of this important function of IL-2 was most pronounced when the PKC inhibitors were added to the medium within 2 h of stimulating the cells with IL-2. The results suggest that transient activation of inactive membrane PKC is linked to the IL-2 receptor signaling, and may be an important step in the mechanism(s) by which the cytokine suppresses cell death in T lymphocytes.

A note of caution in determining glucose in molasses-based alcoholic fermentation broths by an enzymatic electrode

A technique for the determination of glucose in molasses-based alcoholic fermentation broths using an enzyme electrode was assessed. Electrochemical interference produced during the alcoholic fermentation was measured as glucose. An enzymatically inactive membrane discriminates between actual glucose and electrochemical interference.

The activation of inactive membrane-associated protein kinase C is associated with DMSO-induced erythroleukemia cell differentiation

The rapid redistribution of cytosolic protein kinase C (PKC) to membranes and its subsequent proteolytic activation to PKM have been implicated in the DMSO/HMBA-induced differentiation of murine erythroleukemia (MEL) cells. However, DMSO was found not to induce detectable changes in PKC distribution in a MEL cell subline (MEL1) which differentiated normally in response to the agent. Nevertheless, the differentiation of MEL cells appeared dependent on an early PKC-related event because hemoglobinization was partially blocked by the PKC inhibitor H-7 added to cells within the first 2 h after DMSO induction. Indeed, a rapid (15–60 min) increase in membrane PKC activity was detected in DMSO-treated MEL1 cells using a novel method which quantitates the amount of ‘active’ PKC in intact membranes. This transient PKC increase resulted from the activation of ‘inactive’ enzyme already associated with membranes, and not from the translocation of cytosolic PKC. Conventional PKC assays cannot distinguish between active and inactive membrane PKC pools. DMSO also activated inactive membrane PKC in HL-60 cells, but not in S49T-lymphoma and WEHI-231 B-lymphoma cells which do not differentiate in response to DMSO. The results suggest that a rapid and transient increase in membrane PKC activity may be an important early step in DMSO-induced differentiation of erythroleukemia cells.

Changes in the number, size, and shape of synaptic vesicles in an experimental, projected cortical epileptic focus in the rat.

A projected epileptic focus was induced in the brains of experimental rats by applying penicillin G sodium to the contralateral hemisphere. Synapses of type I (after Gray) in the second cortical layer of the mirror area were analyzed morphometrically in the electron microscope 30 min after application of the epileptogenic drug. The number of agranular synaptic vesicles was counted per 0.01 micron2 over the active zone of the presynaptic membrane in an area 0.1 micron wide adjacent to the active zone and in an area of the same width 0.1 micron distant from the active zone. A statistically significant increase in the number of synaptic vesicles was found in the experimental animals as compared with the controls. The number of synaptic vesicles per 0.01 micron2 area was evaluated along the inactive presynaptic membrane, both in the area adjacent to the active zone of the presynaptic membrane and in other areas at successively greater distances from the active zone. The counts in some of these latter areas were statistically significantly higher in the experimental animals. The size and shape of synaptic vesicles were evaluated in three zones, each 0.1 micron wide, parallel to the membrane of the synaptic cleft. Zone I was directly adjacent to the membrane of the synaptic cleft; zone II was 0.1 micron distant; and zone III 0.1 micron distant from the membrane. The results show that the synaptic vesicles of both the experimental and control animals are significantly smaller in zone I than in the more distant zones. This trend is more marked in the controls than in the experimental animals. Histograms of vesical areas of zones II and III in the controls point to two populations of vesicles. The volumes of vesicles are approximately 10% greater in zone I of experimental animals than in the controls. The elongation of the vesicles in the three zones does not differ statistically either in controls or in experimental animals. Experimental animals, however, have statistically more rounded vesicles than the controls. Assuming that the synpatic vesicles contain the transmitters, our results might contribute to a better understanding of the increased excitability of the epileptic focus area.

Electrical Characteristics of the Membrane of An Identified Insect Motor Neurone

ABSTRACT The electrical properties of the membrane of an identified locust motor neurone, the fast extensor tibiae in the metathoracic ganglion, have been investigated to determine : the distribution of excitable and inexcitable membrane; the impulse initiation zone; and the conduction velocity of the spike in the ganglion and in the axon. The waveform of extracellularly recorded spikes indicates that the transition from inactive to active membrane occurs along the region of the neurite which bears many arborizations within the neuropile. Measurements of the delay between orthodromically or antidromically evoked spikes, recorded at the soma and other points along the neurite, place the impulse initiating zone close to the transition between active and inactive membrane. Within the ganglion, the spike is conducted at different velocities over different parts of the neurite. The average velocity within the ganglion is, however, only about a seventh of that in the axon (0· 54 m.s-1 against 4· 1 m.s-1).

A comparison of current‐voltage relations for full and partial agonists.

1. Local conductance changes produced by various bath-applied agonists at frog end-plate membrane were measured using focal recording of extracellular potential in voltage-clamped muscle fibres. The potential difference between a focal micropipette placed on the nerve terminal and another micro-pipette placed on or near inactive membrane was taken as proportional to the agonist-induced current through a small patch of an end-plate membrane. 2. The current-voltage (I--V) relation of active membrane was obtained directly by increasing the membrane potential in a ramp fashion. The change in membrane potential was slow enough for post-synaptic gating processes to reach equilibrium during the ramp. 3. During application of sufficiently low concentrations of full agonists (carbachol, (ACh) and partial agonists (choline and decamethonium) the I--V relation of end-plate membrane showed strong curvature in the range of -60 to -130 mV. The slope of I--V relations increased exponentially with membrane hyperpolarization, an e-fold change in conductance occurring for about 50 mV potential shift. 4. The curvature of the I--V relation of end-plate-membrane activated by the partial agonists choline and decamethonium became less as the agonist concentration was increased, and with high concentrations (choline 15 mM; decamethonium 250 micrometer) the I--V relation became almost straight. 5. When end-plate currents produced by high concentrations of partial agonists were matched by application of equi-active concentrations of carbachol, the carbachol-activated membrane still showed as much curvature in its I--V relation as when low concentrations of carbachol were used. 6. Choline and decamethonium concentrations for which the I--V relation was straight produced much greater depression of miniature end-plate currents than did carbachol concentrations which produced the same membrane current at the holding potential. 7. I--V relations for full agonists at high concentrations were obtained after alpha-bungarotoxin pre-treatment. During application of carbachol (400--500 micrometer) and ACh (30--40 micrometer; after complete inhibition of acetylcholinesterase activity) the I--V relation of end-plate membrane is much less curved than during application of low concentrations. 8. It is concluded that either the voltage sensitivity of agonist-induced end-plate conductance reflects voltage sensitivity of agonist binding, or the partial agonists used can exert a voltage-dependent 'local anaesthetic' action in addition to their agonist activity.

Sodium-dependent amino acid transport by cultured hamster cells: membrane vesicles retain transport changes due to glucose starvation and cycloheximide.

Enhanced alpha-aminoisobutyric acid transport by hamster cells cultured in the absence of D-glucose has been demonstrated in isolated membrane vesicles. The observed enhancement was seen in the presence but not in the absence of Na+. Kinetic analysis of transport using both the intact cells and the membrane vesicles showed that the overall enhancement was associated with an increase in Vmax. Decreases in transport activity by intact cells resulting from extended exposure of the cells to inhibitors of protein synthesis, such as cycloheximide, were also evident in membrane vesicles. The use of metabolically inactive membrane vesicles demonstrated that amino acid uptake by intact cells is a transport property of the plasma membrane. In addition, this study shows that membrane vesicle preparations can be exploited for the purpose of studying the regulation of amino acid transport. Taken together, the data suggest that carrier turnover is involved in the regulation of amino acid transport in animal cells.