Local dendritic Ca2+ signaling induces cerebellar long-term depression.
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The coordinated activity of large numbers of adjacent parallel fiber synapses elevate calcium concentration locally in small regions of Purkinje cell dendrites. Such activity has also been reported to produce long-term depression of parallel fiber synaptic transmission. We have examined the relationship between these two events by combining patch clamp measurements of parallel fiber synaptic transmission with confocal microscopic imaging of the local calcium signals. We find that patterns of parallel fiber activity capable of evoking long-term depression invariably cause increases in Purkinje cell calcium concentration that are very spatially restricted. These results suggest that one function of the local dendritic calcium signals is to induce long-term depression of parallel fiber synapses. (+info)
The role of dendritic action potentials and Ca2+ influx in the induction of homosynaptic long-term depression in hippocampal CA1 pyramidal neurons.
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Long-term depression (LTD) of synaptic efficacy at CA1 synapses is believed to be a Ca(2+)-dependent process. We used high-speed fluorescence imaging and patch-clamp techniques to quantify the spatial distribution of changes in intracellular Ca2+ accompanying the induction of LTD at Schaffer collateral synapses in CA1 pyramidal neurons. Low-frequency stimulation (3 Hz), which was subthreshold for action potentials, produced small changes in [Ca2+]i and failed to elicit LTD. Increasing the stimulus strength so that action potentials were generated produced both robust LTD and increases in [Ca2+]i. Back-propagating action potentials at 3 Hz in the absence of synaptic stimulation also produced increases in [Ca2+]i, but failed to induce LTD. When subthreshold synaptic stimulation was paired with back-propagating action potentials, however, large increases in [Ca2+]i were observed and robust LTD was induced. The LTD was blocked by the N-methyl-D-aspartate receptor (NMDAr) antagonist APV, and stimulus-induced increases in [Ca2+]i were reduced throughout the neuron under these conditions. The LTD was also dependent on Ca2+ influx via voltage-gated Ca2+ channels (VGCCs), because LTD was severely attenuated or blocked by both nimodipine and Ni2+. These findings suggest that back-propagating action potentials can exert a powerful control over the induction of LTD and that both VGCCs and NMDArs are involved in the induction of this form of plasticity. (+info)
Long-term potentiation is associated with new excitatory spine synapses on rat dentate granule cells.
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To investigate possible morphological correlates to long-term potentiation (LTP), three-dimensional reconstruction of serial electron micrographs was employed. LTP was induced in the perforant path/dentate granule cell synapse in two rats. The surgically isolated contralateral side served as control, along with two untreated animals. Longitudinally sectioned and transversally sectioned dendrites were sampled from the middle fifth of the molecular layer and all visibly connected spines were identified. A mixed, unbalanced, nested variance component model was used to make a valid statistical comparison between the LTP and control groups. The spine density was higher in the experimental than in the control groups. The changes were statistically significant in both the longitudinal and transverse sample. In addition, spines with a divided stem and two heads (bifurcating spines) were seen at a higher frequency in the LTP material compared with the contralateral material. From a subset of dendrites all connected spines were reconstructed and detailed measurements of head, neck, and PSD dimensions were made. We failed to find significant differences following LTP on either of the dimensions measured. The results suggest that new spine synapses are formed following LTP, including some of the bifurcating type. (+info)
This study is aimed at testing the hypothesis that sustained phosphorylation underlies long-term desensitization of AMPA receptors, which is thought to be the mechanism of long-term synaptic depression in cerebellar Purkinje cells (PCs). We induced long-term desensitization of AMPA receptors in rat cerebellar slices by (1) a 4-min bath application of quisqualate (0.1 mM) or (2) a 15-min bath application of a protein kinase C (PKC) activator, phorbol-12,13-diacetate (0.5 microM) or -dibutyrate (0.6 microM), followed by a 4-min AMPA (0.1 mM) application. In slices so treated, labeling with an antibody (12P3) against a peptide corresponding to part of AMPA receptor subunit GluR2 including serine 696 and phosphorylated at this serine site revealed phosphorylation of the AMPA receptors in PC dendrites that was sustained for at least 1 hr. At an early phase, within 20 min after the chemical stimulation, the phosphorylation was resistant to an Ca2+ chelator (BAPTA-AM), a metabotropic glutamate receptor antagonist (MCPG), and a PKC inhibitor (calphostin C), whereas at a late phase, 30 min or more after the chemical stimulation, it was blocked by these reagents similarly to long-term desensitization of AMPA receptors. Taken together with data obtained previously using different protocols of chemical stimulation, the present results strongly support the above-mentioned hypothesis. (+info)
Modulation of mammalian dendritic GABA(A) receptor function by the kinetics of Cl- and HCO3- transport.
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1. During prolonged activation of dendritic GABAA receptors, the postsynaptic membrane response changes from hyperpolarization to depolarization. One explanation for the change in direction of the response is that opposing HCO3- and Cl- fluxes through the GABAA ionophore diminish the electrochemical gradient driving the hyperpolarizing Cl- flux, so that the depolarizing HCO3- flux dominates. Here we demonstrate that the necessary conditions for this mechanism are present in rat hippocampal CA1 pyramidal cell dendrites. 2. Prolonged GABAA receptor activation in low-HCO3- media decreased the driving force for dendritic but not somatic Cl- currents. Prolonged GABAA receptor activation in low-Cl- media containing physiological HCO3- concentrations did not degrade the driving force for dendritic or somatic HCO3- gradients. 3. Dendritic Cl- transport was measured in three ways: from the rate of recovery of GABAA receptor-mediated currents between paired dendritic GABA applications, from the rate of recovery between paired synaptic GABAA receptor-mediated currents, and from the predicted vs. actual increase in synaptic GABAA receptor-mediated currents at progressively more positive test potentials. These experiments yielded estimates of the maximum transport rate (vmax) for Cl- transport of 5 to 7 mmol l-1 s-1, and indicated that vmax could be exceeded by GABAA receptor-mediated Cl- influx. 4. The affinity of the Cl- transporter was calculated in experiments in which the reversal potential for Cl- (ECl) was measured from the GABAA reversal potential in low-HCO3- media during Cl- loading from the recording electrode solution. The calculated KD was 15 mM. 5. Using a standard model of membrane potential, these conditions are demonstrated to be sufficient to produce the experimentally observed, activity-dependent GABA(A) depolarizing response in pyramidal cell dendrites. (+info)
Ultrastructural localization of the serotonin transporter in limbic and motor compartments of the nucleus accumbens.
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Extracellular levels of serotonin [5-hydroxytryptamine (5-HT)] in the nucleus accumbens (NAc) can influence both cognitive and motor functions involving extensive connections with the frontal cortex. The 5-HT levels reflect vesicular release and plasmalemmal reuptake through the serotonin transporter (SERT). We used electron microscopic immunocytochemistry to determine the sites for SERT activation in the limbic shell and motor-associated core of the rat NAc. Of the SERT-immunoreactive profiles in each region, >90% were serotonergic axons and axon terminals; the remainder were nonserotonergic dendrites and glia. Axonal SERT immunogold labeling was seen mainly at nonsynaptic sites on plasma membranes and often near 5-HT-containing large dense core vesicles (DCVs). SERT-labeled axonal profiles were larger and had a higher numerical density in the shell versus the core but showed no regional differences in their content of SERT immunogold particles. In contrast, immunoreactive dendrites had a lower numerical density in the shell than in the core. SERT labeling in dendrites was localized to segments of plasma membrane near synaptic contacts from unlabeled terminals and/or dendritic appositions. Our results suggest that in the NAc (1) reuptake into serotonergic axons is most efficient after exocytotic release from DCVs, and (2) increased 5-HT release without concomitant increase in SERT expression in individual axons may contribute to higher extracellular levels of serotonin in the shell versus the core. These findings also indicate that SERT may play a minor substrate-dependent role in serotonin uptake or channel activity in selective nonserotonergic neurons and glia in the NAc. (+info)
Volatile anesthetics block actin-based motility in dendritic spines.
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Dendritic spines form the postsynaptic contact sites for most excitatory synapses in the brain. Spines occur in a wide range of different shapes that can vary depending on an animal's experience or behavioral status. Recently we showed that spines on living neurons can change shape within seconds in a process that depends on actin polymerization. We have now found that this morphological plasticity is blocked by inhalational anesthetics at concentrations at which they are clinically effective. These volatile compounds also block actin-based motility in fibroblasts, indicating that their action is independent of neuron-specific components and thus identifying the actin cytoskeleton as a general cellular target of anesthetic action. These observations imply that inhibition of actin dynamics at brain synapses occurs during general anesthesia and that inhalational anesthetics are capable of influencing the morphological plasticity of excitatory synapses in the brain. (+info)
Microtubule-dependent recruitment of Staufen-green fluorescent protein into large RNA-containing granules and subsequent dendritic transport in living hippocampal neurons.
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Dendritic mRNA transport and local translation at individual potentiated synapses may represent an elegant way to form synaptic memory. Recently, we characterized Staufen, a double-stranded RNA-binding protein, in rat hippocampal neurons and showed its presence in large RNA-containing granules, which colocalize with microtubules in dendrites. In this paper, we transiently transfect hippocampal neurons with human Staufen-green fluorescent protein (GFP) and find fluorescent granules in the somatodendritic domain of these cells. Human Stau-GFP granules show the same cellular distribution and size and also contain RNA, as already shown for the endogenous Stau particles. In time-lapse videomicroscopy, we show the bidirectional movement of these Staufen-GFP-labeled granules from the cell body into dendrites and vice versa. The average speed of these particles was 6.4 microm/min with a maximum velocity of 24. 3 microm/min. Moreover, we demonstrate that the observed assembly into granules and their subsequent dendritic movement is microtubule dependent. Taken together, we have characterized a novel, nonvesicular, microtubule-dependent transport pathway involving RNA-containing granules with Staufen as a core component. This is the first demonstration in living neurons of movement of an essential protein constituent of the mRNA transport machinery. (+info)