Biomedical subjects
Publications and source records attributed to C Lüscher.
The biochemical composition of the postsynaptic membrane and the structure of dendritic spines may be rapidly modulated by synaptic activity. Here we review these findings, discuss their implications for long-term potentiation (LTP) and long-term depression (LTD) and propose a model of sequentially occurring expression mechanisms.
Little is known about the mechanisms that regulate the number of ionotropic glutamate receptors present at excitatory synapses. Herein, we show that GluR1-containing alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors (AMPARs) are removed from the postsynaptic plasma membrane of cultured hippocampal neurons by rapid, ligand-induced endocytosis. Although endocytosis of AMPARs can be induced by high concentrations of AMPA without concomitant activation of N-methyl-D-aspartate (NMDA) receptors (NMDARs), NMDAR activation is required for detectable endocytosis induced by synaptically released glutamate. Activated AMPARs colocalize with AP2, a marker of endocytic coated pits, and endocytosis of AMPARs is blocked by biochemical inhibition of clathrin-coated pit function or overexpression of a dominant-negative mutant form of dynamin. These results establish that ionotropic receptors are regulated by dynamin-dependent endocytosis and suggest an important role of endocytic membrane trafficking in the postsynaptic modulation of neurotransmission.
Compounds known to disrupt exocytosis or endocytosis were introduced into CA1 pyramidal cells while monitoring excitatory postsynaptic currents (EPSCs). Disrupting exocytosis or the interaction of GluR2 with NSF caused a gradual reduction in the AMPAR EPSC, while inhibition of endocytosis caused a gradual increase in the AMPAR EPSC. These manipulations had no effect on the NMDAR EPSC but prevented the subsequent induction of LTD. These results suggest that AMPARs, but not NMDARs, cycle into and out of the synaptic membrane at a rapid rate and that certain forms of synaptic plasticity may utilize this dynamic process.
Although much has been learned about the mechanisms underlying NMDA receptor-dependent long-term potentiation (LTP), considerable debate remains as to whether LTP is expressed as an increase in the synaptic release of glutamate or as an increase in the sensitivity of the postsynaptic glutamate receptors. We have directly measured changes in the synaptic release of glutamate by recording synaptically evoked glial glutamate transporter currents with whole-cell recording. Glial cell responses were very sensitive to manipulations known to change the release of glutamate yet remained constant during LTP. These results argue strongly for a postsynaptic expression mechanism for LTP.
To study the role of G protein-coupled, inwardly rectifying K+ (GIRK) channels in mediating neurotransmitter actions in hippocampal neurons, we have examined slices from transgenic mice lacking the GIRK2 gene. The outward currents evoked by agonists for GABA(B) receptors, 5HT1A receptors, and adenosine A1 receptors were essentially absent in mutant mice, while the inward current evoked by muscarinic receptor activation was unaltered. In contrast, the presynaptic inhibitory action of a number of presynaptic receptors on excitatory and inhibitory terminals was unaltered in mutant mice. These included GABA(B), adenosine, muscarinic, metabotropic glutamate, and NPY receptors on excitatory synapses and GABA(B) and opioid receptors on inhibitory synapses. These findings suggest that a number of G protein-coupled receptors activate the same class of postsynaptic K+ channel, which contains GIRK2. In addition, the GIRK2 channels play no role in the inhibition mediated by presynaptic G protein-coupled receptors, suggesting that the same receptor can couple to different effector systems according to its subcellular location in the neuron.
The inflammatory response associated with Staphylococcus aureus osteomyelitis results in extensive bone damage characterized by apparent increases in bone resorption and formation. These results suggest an increased local release of agents capable of modulating bone remodelling. Tumor necrosis factor alpha (TNF-alpha) is a proinflammatory cytokine proposed to play an important role both in normal bone remodelling and in bone diseases; however, its potential role in osteomyelitis is unclear. This study evaluated changes in bone TNF levels during infection, using a rat model of acute osteomyelitis due to S. aureus. Following direct tibial infection, bacterial counts in bone were persistently high (approximately 6 log10 CFU/g of bone over 63 days) and bone weights increased. TNF activity was undetectable in uninfected bone (<0.01 ng/g of bone) but dramatically higher in infected bone (up to 5.2 +/- 3.5 ng/g of bone). Although TNF-alpha mRNA was weakly detected in uninfected bone, osteomyelitis was associated with up to 37-fold increases in expression of both the 1.6- and 2.4-kb transcripts. Both TNF activity and mRNA transcript levels remained elevated throughout the course of infection. TNF-alpha mRNA detected by in situ hybridization was present in osteoblasts as well as in populations of marrow cells and/or inflammatory infiltrate cells. Histopathology of infected bone indicated extensive bone resorption and adjacent areas of formation that were associated with cells expressing TNF-alpha mRNA. These data suggest that the elevated TNF levels induced by experimental infection may be directly related to changes in the histology of bone during osteomyelitis.
1. To assess the role of intracellular Ca2+ in action potential (AP) propagation, whole-cell recordings of cultured dorsal root ganglion (DRG) cells were carried out while Ca2+ was simultaneously measured with a laser-scanning confocal microscope. 2. Flash photolytic liberation of a Ca2+ buffer during trains of APs which partly failed to invade the DRG cell body immediately lowered intracellular Ca2+ and restored safe AP propagation. Furthermore, the speed of the propagated AP was reduced considerably when intracellular Ca2+ was increased by flash photolysis of caged Ca2+. 3. Both results suggest that intracellular Ca2+ regulates the safety factor for AP propagation and may thus provide a control mechanism for synaptic integration, which acts pre- as well as postsynaptically.
Previous reports suggest that magnetic cortical stimulation (MCS) of the motor cortex can elicit a sensation of movement (kinaesthesia) in paralysed limbs. To assess this sensory effect of MCS, we examined the accuracy of kinaesthetic perception of stimulus-induced right elbow flexion in six blindfolded, healthy subjects. Matching of movements elicited by MCS was compared with matching of passive elbow movements. Small flexion movements between 1.5 and 5 degrees of angle were regularly overestimated by 50-100% when induced by MCS over the parietal cortex, whereas movements elicited by MCS over the frontal cortex or by passive elbow flexion were accurately estimated. Our results provide data compatible with the hypothesis of the existence of a "central sense of movement". Activation of collateral branches projecting from the motor cortex to the sensory could be the underlying mechanism to this phenomenon.
Here we describe the subcellularly uniform control of the intracellular Ca2+ concentration ([Ca2+]i) by flash photolysis of caged Ca2+ or a caged Ca2+ buffer. A mixture of the two Ca2+ indicators Fluo-3 and Fura-red was used together with a laser-scanning confocal microscope to reveal spatial aspects of intracellular Ca2+ signals. The patch clamp technique in the whole-cell variant was applied to load the cells with the indicator mixture together with either DM-nitrophen or diazo-2 and to measure changes in the membrane current. An in vivo calibration was performed to convert the Fluo-3/Fura-red fluorescence ratios to [Ca2+] values. The resulting calibration curve suggested an apparent KD of 1.6 microM, Rmax of 2.15, Rmin of 0.08 and a Hill-coefficient of 0.75 for the indicator mixture. Controlled rupture of the cell membrane revealed a large fraction of immobile intracellular Fura-red fluorescence that may account for the reduced in vivo Rmax value when compared to the in vitro value of 3.1. In cardiac myocytes, flash photolytic release of Ca2+ from DM-nitrophen generated inwardly directed Na+/Ca2+ exchange currents and Ca2+ signals that were graded with the discharged flash-energy. Rapid line-scans revealed subcellularly homogeneous [Ca2+] jumps regardless of the discharged flash energy. Ca2+ signals evoked by L-type Ca2+ currents (ICa) could be terminated rapidly in a spatially homogeneous manner by UV flash photolysis of diazo-2. No side-effects of the photolytic products of DM-nitrophen or diazo-2 with the mixture of Fluo-3/Fura-red were detectable in our experiments. The combination of UV flash photolysis and laser scanning confocal microscopy enabled us to control [Ca2+]i homogeneously on the subcellular level. This approach may improve our understanding of the subcellular properties of cardiac Ca2+ signalling. The technique can also be applied in other cell types and with other signalling systems for which caged compounds are available.
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A Monte Carlo study was carried out to test the reliability of the Maximum Likelihood Estimator (MLE) approach for quantal analysis. This widely used statistical method was applied to extract a finite mixture of Gaussian distributions from simulated data. The data were generated by convolving a distribution of discrete amplitude steps (multiples of a unitary step Q) with Gaussian noise of various standard deviations (sigma n). Our results offer practical guidance on when to use the MLE, taking into account the determining parameters: signal to noise ratio (Q/sigma n, the most important parameter), number of samples collected and the number of components (k). For a given set of parameters the algorithm always converged to the "true" values, never converged to the "true" values or converged in only a fraction of cases to the "true" values. The behavior of the fitting routine in the parameter space is displayed in contour plots. These contour plots can be used as a guide to test the accuracy of deconvolution results.
1. In this and the companion paper the reliability of action potential (AP) propagation through dorsal root ganglion (DRG) cells was investigated. Experimental data were collected from DRG cells of embryonic rat slice cultures of the spinal cord. A field stimulation electrode was used to elicit an AP in the axon. The propagated AP or, in case of conduction block, its electronic residue (ER), was measured intracellularly in the soma of the DRG cell. 2. The morphological and electrophysiological data combined with published data from voltage-clamp studies were taken to implement a compartmental computer model, which allows a precise description of the propagating AP and the channel kinetics at any point along the axon. 3. The safety factor for conduction was found to be low. Thus failures of AP invasion of the DRG cell soma could occur at sites of impedance mismatch when a hyperpolarizing current was applied, a second stimulus felt into the relative refractory period of the first, or when the axon was repetitively stimulated. 4. The ERs of the failed APs had discrete amplitude levels, suggesting that the failures were always caused at the same site along the axon. These sites of low safety factor were found to be the branch point in the unipolar DRG cell and the entrance of the stem piece into the soma in both cell types, the bipolar as well as the unipolar. 5. A systematic comparison of bipolar and unipolar DRG cells showed that the AP conduction through the latter is more reliable. For large cell bodies, the unipolar configuration is needed for save conduction. 6. Conduction through unipolar DRG cells is faster than through bipolar cells because the electrical load of the soma is masked by the high-resistive stem piece. The length of this stem piece is correlated inversely to the delay caused at the branch point, as the electrical load of the soma is more efficiently masked by a long stem piece.
1. The reliability of the propagation of action potentials (AP) through dorsal root ganglion (DRG) cells in embryonic slice cultures was investigated during repetitive stimulation at 1-20 Hz. Membrane potentials of DRG cells were recorded intracellularly while the axons were stimulated by an extracellular electrode. 2. In analogy to the double-pulse experiments reported previously, either one or two types of propagation failures were recorded during repetitive stimulation, depending on the cell morphology. In contrast to the double-pulse experiments, the failures appeared at longer interpulse intervals and usually only after several tens of stimuli with reliable propagation. 3. In the period with reliable propagation before the failures, a decrease in the conduction velocity and in the amplitude of the afterhyperpolarization (AHP), an increase in the total membrane conductance, and the disappearance of the action potential "shoulder" were observed. 4. The reliability of conduction during repetitive stimulation was improved by lowering the extracellular calcium concentration or by replacing the extracellular calcium by strontium. The reliability of conduction decreased by the application of cadmium, a calcium channel blocker, 4-amino pyridine, a fast potassium channel blocker, or apamin or muscarine, the blockers of calcium-dependent potassium channels. The reliability of conduction was not effected by blocking the sodium potassium pump with ouabain or by replacing extracellular sodium with lithium. 5. In the period with reliable propagation cadmium, apamin, and muscarine reduced the amplitude of the AHP. The shoulder of the action potential was more pronounced and not sensitive to repetitive stimulation when extracellular calcium was replaced by strontium. It disappeared when cadmium was applied. 6. In DRG somata changes of the intracellular Ca2+ concentration were monitored by measuring the fluorescence of the Ca2+ indicator Fluo-3 with a laser-scanning confocal microscope. During repetitive stimulation, an accumulation of intracellular calcium occurred that recovered very slowly (tens of seconds) after the AP trains. 7. Computer model simulations performed in analogy to the experimental protocols produced conduction failures during repetitive stimulation only when the calcium currents during the APs were reduced. 8. From these findings it is concluded that conduction failures during repetitive stimulation are dependent on an accumulation of intracellular calcium leading to an inactivation of calcium currents, combined with small contributions of an accumulation of extracellular potassium and a summation of slow potassium conductances.
A modification of a commercially available digital-audio tape (DAT) for DC recordings such as those used in patch clamp and other electrophysiological experiments is introduced. The modified DAT can record data from 2 channels plus a separate trigger signal for up to 2 h. The bandwidth ranges from direct current to 21 kHz; the signal-to-noise ratio is above 90 dB and the A-D resolution is 16 bit. A 120-min cassette yields a storage capacity of 1.4 Gbytes. The trigger is 'hidden' on 1 channel as a burst of 1-ms duration, thus minimizing interference with the recorded data. The modified DAT offers a low-cost yet high quality solution to most analog data storage needs.
1. In embryonic cocultures of spinal cord, dorsal root ganglia, and muscle, excitatory postsynaptic potentials (EPSPs) were recorded in motoneurons during focal electrical stimulation of the dorsal root ganglia or the spinal cord. 2. EPSPs were depressed in amplitude at high-frequency stimulation relative to a control frequency of 0.5 Hz by 47 and 75% at 5 and 10 Hz, respectively. This was true for composite EPSPs and unitary EPSPs. 3. The depression showed a wide range of variability between individual experiments. The degree of depression at 5 Hz was negatively correlated to the rate of spontaneous excitatory input the motoneurons received. There was no correlation to the soma size, the average amplitude of the EPSPs, the rheobase, or the input resistance of the motoneurons. 4. An increase in latency of EPSPs was observed concomitant with or preceding the synaptic depression in most experiments. Total transmission failures, which were absent at low-frequency stimulation, appeared during depression. 5. Large incremental steps in amplitude could be seen during depression, suggesting that several release sites were switched off and on together. 6. Decreasing the extracellular calcium concentration from 5 to 1 mM led to a decrease in the frequency sensitivity of the synaptic efficacy and to a decrease of the EPSP amplitude and latency. 7. Measurements of the antidromic conduction of action potentials evoked in the axons and recorded in the somata of dorsal root ganglion cells revealed an increase in latency and the appearance of conduction failures at stimulation frequencies of 1-10 Hz. The frequency modulation of conduction was decreased in 1 mM compared with 5 mM external calcium. 8. Together these findings suggest that conduction failures in the presynaptic axons contribute to the synaptic depression of EPSPs in embryonic motoneurons.
An HIV-positive patient presented the classical syndrome of pure alexia. Neuroradiologic investigation by computed tomography showed a ring-like lesion in the left posterior white matter. The clinical manifestations as well as the radiologic findings resolved after antiprotozoal treatment.
The study of all the patients sent to the PPU by the disablement insurance in 1975 and 1976 (389 cases). The comparison of the propositions made in medical reports and the actual allowances provided by the disablement insurance reveals the following data: in 326 cases (84%) out of 389, medical doctors only conclude to the presence or absence of a certain rate of disability (from 0 to 100%), to the exclusion of any rehabilitational measure. In 63 cases (16%) measures were proposed such as a professional rehabilitation or a placing in a specialized institution for insertion to a new profession. For this second group, only in 27 cases was the disablement insurance able to follow the medical propositions. Only in 3 cases the measures taken can be considered fully successful, and in 1 case partially successful. All the other measures taken failed. A first examination of the group of 326 cases (propositions only for disablement) which concerned 297 cases also shows a certain number of discordances between the medical propositions and the allowances provided by the disablement insurance. The majority of the discordances has to do with an undervaluation of the rate of disablement by the medical experts. Moreover, the examination of the diagnosis shows that readaptation measures are more easily proposed to patients with psychosomatic complaints, to young men, and allowances rather to women and to patients with obvious psychiatric disorders. The authors discuss some assumptions regarding these observations.