D1R/GluN1 complexes in the striatum integrate dopamine and glutamate signalling to control synaptic plasticity and cocaine-induced responses.

Convergent dopamine and glutamate signalling onto the extracellular signal-regulated kinase (ERK) pathway in medium spiny neurons (MSNs) of the striatum controls psychostimulant-initiated adaptive processes underlying long-lasting behavioural changes. We hypothesised that the physical proximity of dopamine D1 (D1R) and glutamate NMDA (NMDAR) receptors, achieved through the formation of D1R/NMDAR complexes, may act as a molecular bridge that controls the synergistic action of dopamine and glutamate on striatal plasticity and behavioural responses to drugs of abuse. We found that concomitant stimulation of D1R and NMDAR drove complex formation between endogenous D1R and the GluN1 subunit of NMDAR. Conversely, preventing D1R/GluN1 association with a cell-permeable peptide (TAT-GluN1C1) left individual D1R and NMDAR-dependent signalling intact, but prevented D1R-mediated facilitation of NMDAR-calcium influx and subsequent ERK activation. Electrophysiological recordings in striatal slices from mice revealed that D1R/GluN1 complexes control the D1R-dependent enhancement of NMDAR currents and long-term potentiation in D1R-MSN. Finally, intra-striatal delivery of TAT-GluN1C1 did not affect acute responses to cocaine but reduced behavioural sensitization. Our findings uncover D1R/GluN1 complexes as a major substrate for the dopamine-glutamate interaction in MSN that is usurped by addictive drugs to elicit persistent behavioural alterations. They also identify D1R/GluN1 complexes as molecular targets with a therapeutic potential for the vast spectrum of psychiatric diseases associated with an imbalance between dopamine and glutamate transmission.

Mechanisms of synaptic depression triggered by metabotropic glutamate receptors.

Glutamate, by activation of metabotropic receptors (mGluRs), can lead to a reduction of synaptic efficacy at many synapses. These forms of synaptic plasticity are referred to as long-term depression (mGluR-LTD). We will distinguish between mGluR-LTD induced by pre- or postsynaptic receptors and mGluR-LTD induced by the locus of the expression mechanism of the synaptic depression. We will also review recent evidence that mGluR-mediated responses themselves are subject to depression, which may constitute a form of metaplasticity.

[Impulse control disorders and Parkinson's disease]. / Troubles du contrôte des impulsions et maladie de Parkinson.

A variety of behavioral disorders occurring abruptly in patients with Parkinson's disease (PD) has been recently published and attracted considerable attention in the press. Taking the form of pathological gambling, compulsive shopping, addiction to Internet and to other recreational activities, hypersexuality or bulimia, impulse control disorders (ICD) related to PD are probably more frequent than previously appreciated and may have consequences as spectacular as disastrous for the involved patients. ICD are currently viewed as particular adverse reactions to antiparkinsonian medications, notably to dopamine agonists, and, accordingly, tend to improve or disappear when PD therapy is appropriately adjusted.

Effect of high-pressure-induced ice I-to-ice III phase transitions on inactivation of Listeria innocua in frozen suspension.

The inactivation of Listeria innocua BGA 3532 at subzero temperatures and pressures up to 400 MPa in buffer solution was studied to examine the impact of high-pressure treatments on bacteria in frozen matrices. The state of aggregation of water was taken into account. The inactivation was progressing rapidly during pressure holding under liquid conditions, whereas in the ice phases, extended pressure holding times had comparatively little effect. The transient phase change of ice I to other ice polymorphs (ice II or ice III) during pressure cycles above 200 MPa resulted in an inactivation of about 3 log cycles, probably due to the mechanical stress associated with the phase transition. This effect was independent of the applied pressure holding time. Flow cytometric analyses supported the assumption of different mechanisms of inactivation of L. innocua in the liquid phase and ice I (large fraction of sublethally damaged cells due to pressure inactivation) in contrast to cells subjected to ice I-to-ice III phase transitions (complete inactivation due to cell rupture). Possible applications of high-pressure-induced phase transitions include cell disintegration for the recovery of intracellular components and inactivation of microorganisms in frozen food.

Restless AMPA receptors: implications for synaptic transmission and plasticity.

A central assumption in neurobiology holds that changes in the strength of individual synapses underlie changes in behavior. This concept is widely accepted in the case of learning and memory where LTP and LTD are the most compelling cellular models. It is therefore of great interest to understand, on a molecular level, how the brain regulates the strength of neuronal connections. We review a large body of evidence in support of the very straightforward regulation of synaptic strength by changing the number of postsynaptic receptors, and discuss the molecular machinery required for insertion and removal of AMPA receptors.

Epilepsy, hyperalgesia, impaired memory, and loss of pre- and postsynaptic GABA(B) responses in mice lacking GABA(B(1)).

GABA(B) (gamma-aminobutyric acid type B) receptors are important for keeping neuronal excitability under control. Cloned GABA(B) receptors do not show the expected pharmacological diversity of native receptors and it is unknown whether they contribute to pre- as well as postsynaptic functions. Here, we demonstrate that Balb/c mice lacking the GABA(B(1)) subunit are viable, exhibit spontaneous seizures, hyperalgesia, hyperlocomotor activity, and memory impairment. Upon GABA(B) agonist application, null mutant mice show neither the typical muscle relaxation, hypothermia, or delta EEG waves. These behavioral findings are paralleled by a loss of all biochemical and electrophysiological GABA(B) responses in null mutant mice. This demonstrates that GABA(B(1)) is an essential component of pre- and postsynaptic GABA(B) receptors and casts doubt on the existence of proposed receptor subtypes.

Synaptic plasticity and dynamic modulation of the postsynaptic membrane.

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.

Role of AMPA receptor cycling in synaptic transmission and plasticity.

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.

Dynamin-dependent endocytosis of ionotropic glutamate receptors.

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.

Monitoring glutamate release during LTP with glial transporter currents.

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.

G protein-coupled inwardly rectifying K+ channels (GIRKs) mediate postsynaptic but not presynaptic transmitter actions in hippocampal neurons.

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.

Local expression of tumor necrosis factor alpha in an experimental model of acute osteomyelitis in rats.

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.

The perception of movements elicited by magnetic cortex stimulation depends on the site of stimulation.

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.

Photolysis of caged compounds characterized by ratiometric confocal microscopy: a new approach to homogeneously control and measure the calcium concentration in cardiac myocytes.

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.

Control of action potential propagation by intracellular Ca2+ in cultured rat dorsal root ganglion cells.

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.

Practical guidance for testing the accuracy of deconvolution results from quantal analysis.

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.

Action potential propagation through embryonic dorsal root ganglion cells in culture. I. Influence of the cell morphology on propagation properties.

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.