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D Attwell

Publications and source records attributed to D Attwell.

At least 37 records · Page 2Linked to original sources
PubMed1998

Patch-clamp, ion-sensing, and glutamate-sensing techniques to study glutamate transport in isolated retinal glial cells.

We have described how a combination of electrical, ion-sensing, and glutamate-sensing techniques has advanced our understanding of glutamate uptake into isolated salamander retinal glial cells. The next steps in understanding glutamate transport will inevitably depend strongly on molecular biological methods, as described elsewhere in this book, but will also require more detailed study of transporters in their normal environment, perhaps by using patch-clamping or imaging techniques to study cells in situ.

ATP-Binding Cassette Transporters↗
PubMed1997

Retinal processing: visionary transgenics.

Two recent reports in which transgene techniques were used to label specific cell classes in the mouse retina have opened the way to new methods of studying retinal signal processing.

Action Potentials↗
PubMed1997

The role of glutamate transporters in glutamate homeostasis in the brain.

Glutamate transporters in neurones and glia, four of which have been cloned from mammals, play a crucial role in controlling the extracellular glutamate concentration in the brain. In normal conditions, they remove glutamate from the extracellular space and thereby help to terminate glutamatergic synaptic transmission and to prevent the extracellular glutamate concentration from rising to neurotoxic values. Glutamate transport on these carriers is thought to be driven by the cotransport of Na+, the counter-transport of K+, and either the cotransport of H+ or the counter-transport of OH-. Activating the transporters also activates an anion conductance in their structure, the anion flux through which is not coupled to glutamate movement and varies widely for the different transporters. During hypoxia or ischaemia, glutamate transporters can run backwards, releasing glutamate into the extracellular space, triggering the death of neurones and thus causing mental and physical handicap. The rate of glutamate release by this process is slowed by the acid pH occurring in hypoxia/ischaemia, which may help protect the brain during transient, but not sustained, ischaemia.

ATP-Binding Cassette Transporters↗
PubMed1996

Postsynaptic glutamate uptake in rat cerebellar Purkinje cells.

1. Whole-cell clamp experiments on Purkinje neurons in rat cerebellar slices were used to test whether glutamate transporters, detected immunocytochemically in the somata and dendrites of the cells, are functional in the cell surface membrane, and to investigate their role in terminating synaptic transmission. 2. A membrane current was detected with the pharmacology, voltage and ion dependence of a glutamate uptake current. Part of the current was generated by an anion conductance activated when uptake occurs. 3. With sodium and glutamate inside the cell, raising the external potassium concentration generated an outward current attributable to reversed operation of glutamate transporters. 4. The magnitude of the uptake current suggested that Purkinje cell transporters could help to terminate transmission at the climbing and parallel fibre to Purkinje cell synapses. Reducing postsynaptic glutamate uptake with intracellular D-aspartate prolonged the climbing fibre EPSC. 5. These data establish the existence of functional postsynaptic glutamate transporters, show that they contribute to terminating synaptic transmission, and suggest that they may play a role in the preferential death of Purkinje cells in ischaemia.

2-Amino-5-phosphonovalerate↗
PubMed1996

Anion conductance behavior of the glutamate uptake carrier in salamander retinal glial cells.

Glutamate uptake is driven by the cotransport of Na+ ions, the countertransport of K+ ions, and either the countertransport of OH- or the cotransport of H+ ions. In addition, activating glutamate uptake carriers has been shown to lead to activation of an anion conductance present in the carrier structure. Here we characterize the ion selectivity and gating of this anion conductance. The conductance is small with Cl- as the permeant anion, but it is large with NO3- or ClO4- present, undermining the earlier use of NO3- and ClO4- to suggest that OH- countertransport rather than H+ cotransport helps drive uptake. Activation of the anion conductance can be evoked by extra- or intracellular glutamate and can occur even when glutamate transport is inhibited. By running the carrier backward and detecting glutamate release with AMPA receptors in neurons placed near the glial cells, we show that anion flux is not coupled thermodynamically to glutamate movement, but OH-/H+ transport is. The possibility that cell excitability is modulated by the anion conductance associated with glutamate uptake suggests a target for therapeutic drugs to reduce glutamate release in conditions like epilepsy.

Amino Acid Transport System X-AG↗
PubMed1996

Modulation of non-vesicular glutamate release by pH.

Glutamate uptake into glial cells helps to keep the brain extracellular glutamate concentration, [glu]o, below levels that kill neurons. Uptake is powered by the transmembrane gradients of Na+, K+ and pH. When the extracellular [K+] rises in brain ischaemia, uptake reverses, releasing glutamate into the extracellular space. Here we show, by monitoring glutamate transport electrically and detecting released glutamate with ion channels in neurons placed outside glial cells, that a raised [H+] inhibits both forward and reversed glutamate uptake. No electroneutral reversed uptake was detected, contradicting the idea that forward and reversed uptake differ fundamentally. Suppression of reversed uptake by the low pH occurring in ischaemia will slow the Ca(2+)-independent release of glutamate with can raise [glu]o to a neurotoxic level, and will thus protect the brain during a transient loss of blood supply.

ATP-Binding Cassette Transporters↗
PubMed1996

The changing pattern of neuroscience PhDs in the UK.

The establishment by the Wellcome Trust of two four-year PhD programmes in neuroscience, in which PhD students will study neuroscience in greater depth and breadth and be able to make a more informed choice of PhD project and supervisor, marks a commitment to improving the quality of graduate training in neuroscience in the UK.

Education, Graduate↗
PubMed1996

Non-synaptic release of ATP by electrical stimulation in slices of rat hippocampus, cerebellum and habenula.

ATP is thought to be a fast neurotransmitter in the medial habenula region of the brain, and may be coreleased with other transmitters, for example with glutamate in the hippocampus. We monitored ATP release in rat brain slices using the bioluminescent indicator system luciferin-luciferase. Electrical stimulation of the hippocampus, cerebellum or habenula led to ATP release, but this release was calcium-independent and was not blocked by tetrodotoxin, or by other agents found to block ATP release from red blood cells. Although calcium-dependent ATP release may occur in response to electrical stimulation, it appears to be overwhelmed by calcium-independent release, which may result from electroporation of cells close to the stimulating electrode. Consistent with this, uptake into cells of the fluorescent dye Lucifer yellow was promoted by electrical stimulation. Our data undermine a previous suggestion, based on use of the luciferin-luciferase technique, that ATP is synaptically released with glutamate in the hippocampus.

Action Potentials↗
PubMed1995

Pre- and postsynaptic determinants of EPSC waveform at cerebellar climbing fiber and parallel fiber to Purkinje cell synapses.

Excitatory postsynaptic currents (EPSCs) at the parallel fiber and climbing fiber to Purkinje cell synapses were studied by whole-cell clamping Purkinje cells in cerebellar slices. Reducing glutamate release with adenosine or GABA decreased the amplitude of the EPSCs, with a larger suppression being produced at the parallel fiber synapse. Reducing glutamate release also speeded the decay of the EPSCs, and this effect was not a series resistance artefact since postsynaptic reduction of the current with CNQX did not speed the EPSC decay. Blocking glutamate uptake slowed the decay of the EPSCs. At the climbing fiber synapse, adenosine had little suppressive effect on the smaller EPSC evoked by the second of a pair of stimuli. Blocking desensitization of postsynaptic AMPA receptors prolonged the EPSC decay, preferentially increased the size of the second EPSC, and resulted in adenosine having a similar suppressive effect on the first and second EPSC. These data suggest that, at these synapses, the fall of glutamate concentration in the synaptic cleft overlaps with the decay of the EPSC, and that the EPSC size and duration are controlled by the amount of glutamate released, the rate of glutamate uptake, and desensitization.

Adenosine↗
PubMed1994

Counter-transport of potassium by the glutamate uptake carrier in glial cells isolated from the tiger salamander retina.

1. To investigate the transport of potassium on the glutamate uptake carrier, the glutamate uptake current in isolated retinal Müller cells was monitored by whole-cell clamping, while measuring changes of potassium concentration outside the cells ([K+]o) with an ion-sensitive microelectrode. 2. Activating glutamate uptake led to an accumulation of potassium outside the cells, consistent with the hypothesis, based on less direct evidence, that the glutamate uptake carrier transports potassium out of the cell. 3. The glutamate-evoked rise of [K+]o showed the pharmacology and sodium dependence of glutamate uptake. 4. The rise in [K+]o was proportional to the uptake current flowing between 0 and -80 mV, implying that the ratio of K+ transported to charge transported by the uptake carrier is constant over this voltage range. The K+ to charge transport ratio was the same for uptake of D-aspartate and L-glutamate. 5. By clamping cells with pipettes containing solutions of different [K+], the dependence of the glutamate and aspartate uptake currents on intracellular [K+] was determined. L- and D-aspartate transport showed a smaller maximum uptake current (Imax), and a smaller apparent Michaelis constant (Km) for activation by intracellular K+, than did L-glutamate transport. The ratio of Imax to Km was the same for these three analogues, a result which can be predicted from simple models of the carrier's operation. 6. Fully activating glutamate uptake in Müller cells in the intact retina would produce a K+ load into the extracellular space of about 0.6 mM s-1. Suppression of glutamate release from photoreceptors by light will reduce K+ efflux from Müller cells in the outer retina; this may contribute to the light-evoked fall of [K+]o observed in the outer retina, and thus contribute to shaping the electroretinogram.

Amino Acid Transport System X-AG↗
PubMed1994

Arachidonic acid depresses non-NMDA receptor currents.

Arachidonic acid has been proposed as an intercellular messenger in the nervous system. It is released when glutamate acts on postsynaptic receptors, potentiates NMDA receptor currents and depresses glutamate uptake. Here we report the effects of arachidonic acid on non-NMDA receptor currents, studied by whole-cell clamping isolated neurons and neurons in tissue slices. In cultured cerebellar granule cells and in freshly isolated hippocampal pyramidal cells arachidonic acid decreased the current produced by iontophoresed AMPA. This depression was not due to increased desensitization of the AMPA receptor. In cerebellar slices, arachidonic acid depressed the non-NMDA component of the synaptic current at the mossy fibre to granule cell and the parallel fibre to Purkinje cell synapses. However, this depression was not always seen, possibly because the lipophilic arachidonic acid is absorbed by superficial cells in the slice and does not reach the synapse being studied. Depression of non-NMDA receptor currents by arachidonic acid may reflect the presence of an arachidonic acid binding site on the non-NMDA receptor, but non-NMDA receptor subunits show much less sequence homology with fatty acid binding proteins than does the NMDA receptor.

Amino Acid Sequence↗
PubMed1994

Triggering and execution of neuronal death in brain ischaemia: two phases of glutamate release by different mechanisms.

A reduced blood or oxygen supply to the brain leads to neuronal death caused by excessive activation of glutamate receptors. Recent evidence suggests that two distinct phases of glutamate release produce this death. During ischaemia or hypoxia, glutamate is released by reversed operation of glutamate uptake carriers. It activates N-methyl-D-aspartate (NMDA) receptors, increases the intracellular concentration of Ca2+, and triggers a long-lasting potentiation of NMDA-receptor-gated currents. After ischaemia, glutamate released by Ca(2+)-dependent exocytosis activates an excessive influx of Ca2+ largely through potentiated NMDA-receptor-channels, which leads to neuronal death. The therapeutic implications of such a scheme are discussed.

Animals↗
PubMed1994

Neurotransmitter transporters.

In the past year, our knowledge of neurotransmitter transporters has increased significantly. Recently, new members of two families of plasma membrane uptake carriers have been cloned, and the stoichiometries, physiological function and mechanisms of modulation of some of these transporters are now better understood. These developments highlight the possible role of neurotransmitter transporters in disease states, in the development of the nervous system, and as targets for therapeutic drugs.

Animals↗
PubMed1994

Intracellular pH changes produced by glutamate uptake in rat hippocampal slices.

1. The mean intracellular pH in area CA1 of rat hippocampal slices was monitored fluorescently after loading the cells with the dye BCECF-AM. 2. Including L-glutamate in the solution superfusing the slice led to the intracellular pH becoming more acid. This acidification had a roughly Michaelis-Menten dependence on the superfused glutamate concentration with a half-maximal dose around 200 microM: this value must overestimate the glutamate concentration at most of the cells, which will be reduced by uptake. 3. The glutamate-evoked acidification was not significantly reduced by blockers of glutamate-gated ion channels [6-cyano-7-nitroquinoxaline-2,3- dione (CNQX) and D-aminophosphonovalerate (APV)] nor by blockers of gamma-aminobutyric acid (GABA)- and glycine-gated channels (picrotoxin and strychnine), and so was not produced by H+ entry through alpha-amino-3-hydroxy-5-methyl-4- isoxazolepropionic acid (AMPA) or N-methyl-D-aspartate (NMDA) receptor channels nor by HCO3- exit through the chloride channels controlled by GABA or glycine. 4. The glutamate-evoked acidification was not reduced by tetrodotoxin (TTX), ruling out the possibility of it being generated by action potentials. It was also unaffected by saturation of presynaptic L-amino-4-phosphonobutanoate (AP4) receptors with AP4. 5. In the presence of blockers of glutamate-, GABA-, and glycine-gated channels, the acidification showed the pharmacology of glutamate uptake and was reduced by a glutamate uptake blocker. 6. The glutamate-evoked acidification showed an ion dependence similar to that of glutamate uptake. It was abolished by removal of extracellular sodium and was reduced by raising the extracellular potassium concentration. It was unaffected by blockers of Na+/H+ exchange (amiloride) and Na+/HCO3- cotransport [4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS)] and so was not produced by the Na+ influx accompanying glutamate uptake changing the activity of these carriers. 7. These data show that the glutamate uptake carrier acidifies hippocampal cells, possibly because it transports a pH-changing anion out of the cell as in salamander glial cells. Glutamate uptake may thus contribute to activity-induced pH changes in the nervous system.

Acid-Base Equilibrium↗