Plasmalemma transport of OH− inChara corallina

Summary The light-mediated, time-dependent rise in the pH value at the center of an alkaline band was analyzed using the methods of numerical analysis. From this analysis an expression of the time-dependent build-up of OH^– efflux was obtained for these bands. This information can now be employed to determine whether the light-activated transport of OH^– and HCO ₃ ^– influences the electrical properties of the plasmalemma. The dark-induced deactivation of OH^– transport was also characterized, revealing a transition from efflux to a transient influx phase during deactivation.Numerical analysis of the steady-state OH^– diffusion pattern, established along the surface of an alkaline band, revealed that the OH^– efflux width was wider than previously envisaged. It was also found that OH^– sink regions exist on either side of the efflux zone. These, and other characteristics revealed by the numerical analysis, enabled us to extend the OH^– transport model proposed by Lucas (J. Exp. Bot. 1975,26:347).     

Models of long-distance transport: how is carrier-dependent auxin transport regulated in the stem?

? This paper presents two models of carrier-dependent long-distance auxin transport in stems that represent the process at different scales. ? A simple compartment model using a single constant auxin transfer rate produced similar data to those observed in biological experiments. The effects of different underlying biological assumptions were tested in a more detailed model representing cellular and intracellular processes that enabled discussion of different patterns of carrier-dependent auxin transport and signalling. ? The output that best fits the biological data is produced by a model where polar auxin transport is not limited by the number of transporters/carriers and hence supports biological data showing that stems have considerable excess capacity to transport auxin. ? All results support the conclusion that auxin depletion following apical decapitation in pea (Pisum sativum) occurs too slowly to be the initial cause of bud outgrowth. Consequently, changes in auxin content in the main stem and changes in polar auxin transport/carrier abundance in the main stem are not correlated with axillary bud outgrowth.     

The energetics of Cl− active transport inChara

Summary The rate of Cl^– influx in intactChara was inhibited whenever the ATP concentration was reduced by application of metabolic inhibitors. In perfused cells, however, a net influx of Cl^– against its electrochemical gradient could be observed in the absence of ATP. Addition of ATP to the perfusion medium slightly stimulated Cl^– influx in one experiment but had no effect in another. Addition of ADP, NADH or metabolic inhibitors did not alter the influx rate. Consideration of the potential energy gradients across theChara plasmalemma in the perfused state leads to the conclusion that Cl^– influx occurs by cotransport with H⁺ or OH^–.     

Mathematical models of α-synuclein transport in axons

To investigate possible effects of diffusion on α-synuclein (α-syn) transport in axons, we developed two models of α-syn transport, one that assumes that α-syn is transported only by active transport, as part of multiprotein complexes, and a second that assumes an interplay between motor-driven and diffusion-driven α-syn transport. By comparing predictions of the two models, we were able to investigate how diffusion could influence axonal transport of α-syn. The predictions obtained could be useful for future experimental work aimed at elucidating the mechanisms of axonal transport of α-syn. We also attempted to simulate possible defects in α-syn transport early in Parkinson's disease (PD). We assumed that in healthy axons α-syn localizes in the axon terminal while in diseased axons α-syn does not localize in the terminal (this was simulated by postulating a zero α-syn flux into the terminal). We found that our model of a diseased axon predicts the build-up of α-syn close to the axon terminal. This build-up could cause α-syn accumulation in Lewy bodies and the subsequent axonal death pattern observed in PD (‘dying back’ of axons).     

A Maxwell's Demon type of membrane transport: possibility for active transport by ABC-type transporters?

ATP-binding-cassette (or ABC)-type transporters constitute one of the largest family of membrane transporters in nature. Many of its members move substrates "actively", i.e. in an ATP-dependent manner against an electrochemical gradient. No consensus is available about the mechanism. Therefore, a novel class of transport mechanisms is proposed based on Maxwell's demon idea. This transport mechanism consists of a gated pore that selectively opens for substrates from one, but not the other side. Thermoenergy (Brownian motion) would suffice for substrate translocation across the membrane; energy for synchronizing gate opening with substrate arrival would come from ATP hydrolysis. Simulations demonstrate that such a mechanism would be thermodynamically and kinetically feasible. It exhibits "active", unidirectional transport, saturation, and other typical features of protein-catalysed reactions. It also shows pore behavior with charged substrates moving under the influence of electrical potentials. Its efficiency depends on a diffusion time constant of the substrate in solution that is slower than the transit time through the membrane, a situation that can realistically be achieved at millimolar or lower substrate concentrations. Features of the novel mechanism that differ significantly from P- or F-type ATPases are: (1) transport cannot be run in "reverse" to synthesize ATP even if sufficient energy is available in the gradient of the transported solute and (2) unidirectional and net substrate fluxes through the transporter diverge with increasing substrate concentration.     

Three decades in transport business: studies of metabolite transport in chloroplasts – a personal perspective

This article gives an historical overview of our group's research on various metabolite translocators of chloroplasts, such as the translocators for phosphorylated intermediates of the Calvin–Benson cycle and of glycolysis, of ADP and ATP, of dicarboxylates, of pyruvate and of hexoses; how it began and where it led to. Wherever appropriate, references will be made to research in other laboratories. This revised version was published online in June 2006 with corrections to the Cover Date.     

Blood-to-brain influx transport of nicotine at the rat blood–brain barrier: Involvement of a pyrilamine-sensitive organic cation transport process

Nicotine is the most potent neural pharmacological alkaloid in tobacco, and the modulation of nicotine concentration in the brain is important for smoking cessation therapy. The purpose of this study was to elucidate the net flux of nicotine transport across the blood–brain barrier (BBB) and the major contributor to nicotine transport in the BBB. The in vivo brain-to-blood clearance was determined by a combination of the rat brain efflux index method and a rat brain slice uptake study, and the blood-to-brain transport of nicotine was evaluated by in vivo vascular injection in rats and a conditionally immortalized rat brain capillary endothelial cell line (TR-BBB13 cells) as an in vitro model of the rat BBB. The blood-to-brain nicotine influx clearance was obtained by integration plot analysis as 272 μL/(min g brain), and this value was twofold greater than the brain-to-blood efflux clearance (137 μL/(min g brain)). Thus, it is suggested that the net flux of nicotine transport across the BBB is dominated by blood-to-brain influx transport. In vivo blood-to-brain nicotine transport was inhibited by pyrilamine. [³H]Nicotine uptake by TR-BBB13 cells exhibited time-, temperature-, and concentration-dependence with a K_m value of 92 μM. Pyrilamine competitively inhibited nicotine uptake by TR-BBB13 cells with a K_i value of 15 μM, whereas substrates and inhibitors of organic cation transporters had little effect. These results suggest that pyrilamine-sensitive organic cation transport process(es) mediate blood-to-brain influx transport of nicotine at the BBB, and this is expected to play an important role in regulating nicotine-induced neural responses.     

Ammonium ion transport—a cause of cell death

Ammonium can be transported into the cell by ion pumps in the cytoplasmic membrane. Ammonia then diffuse out through the cell membrane. A futile cycle is created that results in cytoplasmic acidification and extracellular alkalinisation. Ammonium transport can be quantified by measuring the extracellular pH changes occurring in a cell suspension (in PBS) after addition of ammonium. By using this technique, in combination with specific inhibitors of various ion pumps, it was shown that ammonium ions are transported across the cytoplasmic membrane by the Na⁺K⁺2Cl^--cotransporter in both hybridoma and myeloma cells. Further, the Na⁺/H⁺ exchanger, which regulates intracellular pH by pumping out protons, was shown to be active during ammonium exposure. The viability of hybridoma cells suspended in PBS and exposed to NH _inf4 ^sup+ for only 90 min, was reduced by 11% (50% necrosis and 50% apoptosis). A control cell suspension did not loose viability during this time. Turning off the activity of the Na⁺/H⁺ exchanger (by amiloride) during ammonium exposure decreased viability further, while inhibiting transport itself (by bumetanide) restored viability to the same level as for the control experiment with bumetanide alone. These results show that one effect of ammonia/ammonium on cell physiology is specifically related to the inward transport of ammonium ions by membrane bound ion pumps.Abbreviations q _pH specific rate of pH increase (pH units per min and 10⁶ cells per ml)     

The regulation of transport of glucose and methyl α-glucoside in Pseudomonas aeruginosa

The methyl alpha-glucoside-transport system of Pseudomonas aeruginosa has been characterized with respect to its specificity, energy-dependence, kinetics and regulation. The uptake of glucose involved two components, one of which transported glucose (K(m)=8mum) and methyl alpha-glucoside (K(m)=2.8mm) whereas the other was more complex, involving the extracellular activity of glucose dehydrogenase. Mutants defective in this enzyme have been isolated and characterized. The methyl alpha-glucoside-glucose-transport system was repressed when the organism was grown in the absence of glucose; the induction of this transport system by glucose, and its activity once induced, were inhibited by products of citrate metabolism.     

A time-dependent mathematical expression of the münch hypothesis of phloem transport

A time-dependent mathematical expression of the Münch, osmotically driven mass flow hypothesis of phloem transport is presented. The dependent variables include concentration of solutes, pressure, velocity of phloem sap, osmotic flux of water, and concentration dependent unloading of solutes. The model meets conservation requirements during all iterations, and responds realistically to changes in independent variables. Given the same set of independent variables the time-dependent model converges to the same values as the closed-form steady-state model of Goeschl et al. (1976) regardless of the initial conditions.     

Intracellular transport of mouse kidney β-glucuronidase induced by gonadotrophin

1. The response of renal beta-glucuronidase with time to the injection of gonadotrophin was investigated in each submicrosomal fraction of rough and smooth microsomal fractions of mouse kidney homogenate. 2. The increase in beta-glucuronidase activity appeared initially in membranes of the rough microsomal fraction, 24h after injection. 3. Afterwards the newly synthesized enzyme appeared in the contents of the rough microsomal fraction and was subsequently found in the smooth microsomal fraction, reaching a maximum concentration in this fraction at 72h. 4. At this juncture, a decrease in the enzyme activity was observed in rough microsomal contents whereas the lysosomal fraction had reached its maximum value. 5. The time-course of the appearance of beta-glucuronidase in the submicrosomal fractions after the gonadotrophin stimulation suggests that the newly synthesized enzyme at the site of membrane-bound ribosomes is transferred across the membrane into cisternae of the rough endoplasmic reticulum, and then is transported into lysosomes via the smooth endoplasmic reticulum. 6. The properties of microsomal and lysosomal beta-glucuronidases were compared.     

Characterization of β-hydroxybutyrate transport in rat erythrocytes and thymocytes

A method was developed for study of β-hydroxybutyrate transport in erythrocytes and thymocytes. Critical to the method was a centrifugal separation of cells from medium which took advantage of β-hydroxybutyrate transport's temperature dependence and inhibition by phloretin and methylisobutylxanthine, all of which are demonstrated in this work. These properties suggested mediated transport, as did saturation kinetics and inhibition by several agents including pyruvate and α-cyanocinnamate. Most conclusive in this regard was a 2-fold preference for d- over l-β-hydroxybutyrate. Entry was not Na⁺ dependent. It was stimulated by substitution of SO₄^2? for most of the Cl^?. The equilibrium β-hydroxybutyrate space was much higher than the Cl^? space of thymocytes, suggesting that β-hydroxybutyrate entry is not associated with net inward negative current and is not coupled to outward Cl^? or inward K⁺ movement (assuming that K⁺ is at electrochemical equilibrium). Coupling to H⁺ entry or OH^? exit is compatible with the result. These findings are consistent with β-hydroxybutyrate entry by the carboxylate transport site which has been studied extensively with pyruvate and lactate as permeants. The Cl^?/HCO₃^? exchange carrier did not appear to contribute significantly to β-hydroxybutyrate transport.     

Revisiting the Münch pressure-flow hypothesis for long-distance transport of carbohydrates: modelling the dynamics of solute transport inside a semipermeable tube

A mathematical model of the Münch pressure-flow hypothesis for long-distance transport of carbohydrates via sieve tubes is constructed using the Navier-Stokes equation for the motion of a viscous fluid and the van't Hoff equation for osmotic pressure. Assuming spatial dimensions that are appropriate for a sieve tube and ensuring suitable initial profiles of the solute concentration and solution velocity lets the model become mathematically tractable and concise. In the steady-state case, it is shown via an analytical expression that the solute flux is diffusion-like with the apparent diffusivity coefficient being proportional to the local solute concentration and around seven orders of magnitude greater than a diffusivity coefficient for sucrose in water. It is also shown that, in the steady-state case, the hydraulic conductivity over one metre can be calculated explicitly from the tube radius and physical constants and so can be compared with experimentally determined values. In the time-dependent case, it is shown via numerical simulations that the solute (or water) can simultaneously travel in opposite directions at different locations along the tube and, similarly, change direction of travel over time at a particular location along the tube.     

The reverse transport of DA, what physiological significance?

It is well established that midbrain dopamine neurons innervating the striatum, release their neurotransmitter through an exocytotic process triggered by the neural firing and involving a transient calcium entry in the terminals. Long ago, it had been proposed, however, that another mechanism of release could co-exist with classical exocytosis, involving the reverse-transport of the cytosolic amine by the carrier, ordinarily responsible for uptake function. This atypical mode of release could be evoked directly at the preterminal level by multiple environmental endogenous factors involving transient alterations of the sodium gradient. It cannot be excluded that this mode of release participates in the firing-induced release. In contrast with the classical exocytosis of a preformed DA pool, the reverse-transport of DA requires simultaneous alterations of intraterminal amine metabolism including synthesis and displacement from storage compartment. The concept of a reverse-transport of dopamine is coming from the observations that releasing substances, such as amphetamine-related molecules, actually induce this type of transport. A large set of arguments advocates that reverse-transport plays a role in the maintenance of basal extracellular DA concentration in striatum. It was also often evoked in physiopathological situations including ischemia, neurodegenerative processes, etc. The most recent studies suggest that this release could occur mainly outside the synapses, and thus could constitute a major feature in the paracrine transmission, sometimes evoked for DA.     

Is defective electron transport at the hub of aging?

The bulwark of the mitochondrial theory of aging is that a defective respiratory chain initiates the death cascade. The increased production of superoxide is suggested to result in progressive oxidant damage to cellular components and particularly to mtDNA that encodes subunits assembled in respiratory complexes. Earlier studies of respiration in muscle mitochondria obtained from large cohorts of patients supported this notion by showing that either singly or in combinations, the respiratory complexes exhibited decreased activity in the elderly. The following critique of the most cited publications over the past decade points out the systematic errors that put earlier work at odds with recent findings. These later investigations indicate that aging has no overt effect on either the electron transport system or oxidative phosphorylation.