Biomedical subjects

O Prange

Publications and source records attributed to O Prange.

6 recordsLinked to original sources

PubMed2001

Modular transport of postsynaptic density-95 clusters and association with stable spine precursors during early development of cortical neurons.

The properties of filopodia and spines and their association with the postsynaptic density (PSD) protein PSD-95 were studied during early development of cultured cortical neurons using time-lapse confocal microscopy. Neurons were transfected with recombinant PSD-95 constructs fused to green fluorescent protein (GFP) for, on average, either 8 d in vitro (DIV) or 14 DIV. We find that, during 1 hr of imaging, filopodia and spines bearing PSD-95/GFP clusters are significantly more stable (i.e., do not turnover) than those lacking clusters. When present within a spine precursor, a PSD-95/GFP cluster appeared to nucleate a relatively stable structure around which filopodium-spine membranes can move. Although processes bearing clusters were generally stable, in 8 DIV neurons, we observed that a subset ( approximately 10%) of PSD-95/GFP clusters underwent rapid modular translocation between filopodia-spines and dendritic shafts. We conclude that, during early synaptic maturation, prefabricated PSD-95 clusters are trafficked in a developmentally regulated process that is associated with filopodial stabilization and synapse formation.

Animals↗

PubMed2000

Vesicle number does not predict postsynaptic measures of miniature synaptic activity frequency in cultured cortical neurons.

We tested the hypothesis that heterogeneity in the frequency of miniature synaptic activity reflects differences in the number of vesicles present in presynaptic terminals. Using imaging techniques, we measured dendritic miniature synaptic calcium transients attributed to the spontaneous release of single transmitter quanta. Following imaging, the identified neurons were processed for serial transmission electron microscopy. At sites of quantal Ca(2+) transients mediated by N-methyl-D-aspartate receptors, we confirmed the presence of excitatory synapses and measured the total number of vesicles and the number of docked vesicles. We observed no correlation between the frequency of spontaneous miniature activity and either the total vesicle number or the number of docked vesicles. We conclude that the presynaptic vesicle complement as measured by ultrastructural analysis does not necessarily determine the frequency of spontaneous activity at synapses mediated by N-methyl-D-aspartate receptors.

Animals↗

PubMed1999

Correlation of miniature synaptic activity and evoked release probability in cultures of cortical neurons.

Spontaneous miniature synaptic activity is caused by action potential (AP)-independent release of transmitter vesicles and is regulated at the level of single synapses. In cultured cortical neurons we have used this spontaneous vesicle turnover to load the styryl dye FM1-43 into synapses with high rates of miniature synaptic activity. Automated selection procedures restricted analysis to synapses with sufficient levels of miniature activity-mediated FM1-43 uptake. After FM1-43 loading, vesicular FM1-43 release in response to AP stimulation was recorded at single synapses as a measure of release probability. We find that synapses with high rates of miniature activity possess significantly enhanced evoked release rates compared with a control population. Because the difference in release rates between the two populations is [Ca(2+)](o)-dependent, it is most likely caused by a difference in release probability. Within the subpopulation of synapses with high miniature activity, we find that the probabilities for miniature and AP-evoked release are correlated at single synaptic sites. Furthermore, the degree of miniature synaptic activity is correlated with the vesicle pool size. These findings suggest that both evoked and miniature vesicular release are regulated in parallel and that the frequency of miniature synaptic activity can be used as an indicator for evoked release efficacy.

Action Potentials↗

PubMed1999

Ultrastructural correlates of quantal synaptic function at single CNS synapses.

We have tested the hypothesis that functional differences between synapses are associated with ultrastructure in cultured cortical neurons. Using Ca(2+) imaging, we measured NMDA receptor-mediated miniature synaptic calcium transients attributed to the spontaneous release of single transmitter quanta. After imaging, the identified neurons were processed for serial transmission electron microscopy. At sites of quantal NMDA receptor-dependent Ca(2+) transients, we confirmed the presence of excitatory synapses and measured spine size and synaptic contact area. Our results demonstrate that synapse size correlates positively with the amplitude of the NMDA receptor-mediated postsynaptic response, suggesting that larger synapses express a greater number of NMDA receptors. Therefore, regulation of quantal amplitude may involve processes that alter synapse size.

Animals↗

PubMed1999

Analysis of multiquantal transmitter release from single cultured cortical neuron terminals.

Application of single synapse recording methods indicates that the amplitude of postsynaptic responses of single CNS synapses can vary greatly among repeated stimuli. To determine whether this observation could be attributed to synapses releasing a variable number of transmitter quanta, we assessed the prevalence of multiquantal transmitter release in primary cultures of cortical neurons with the action potential (AP)-dependent presynaptic turnover of the styryl dye FM1-43 (,; ). It was assumed that if a high proportion of vesicles within a terminal were loaded with FM1-43 the amount of dye released per stimulus would be proportional to the number of quanta released and/or the probability of release at a terminal. To rule out differences in the amount of release (between terminals) caused by release probability or incomplete loading of terminals, conditions were chosen to maximize both release probability and terminal loading. Three-dimensional reconstruction of terminals was employed to ensure that bouton fluorescence was accurately measured. Analysis of the relationship between the loading of terminals and release indicated that presumed larger terminals (>FM1-43 uptake) release a greater amount of dye per stimulus than smaller terminals, suggesting multiquantal release. The distribution of release amounts across terminals was significantly skewed toward higher values, with 13-17% of synaptic terminals apparently releasing multiple quanta per AP. In conclusion, our data suggest that most synaptic terminals release a relatively constant amount of transmitter per stimulus; however, a subset of terminals releases amounts of FM1-43 that are greater than that expected from a unimodal release process.

2-Amino-5-phosphonovalerate↗

PubMed1999

Amplification of calcium signals at dendritic spines provides a method for CNS quantal analysis.

It has been proposed that the small volume of a dendritic spine can amplify Ca2+ signals during synaptic transmission. Accordingly, we have performed calculations to determine whether the activation of N-methyl-D-aspartate (NMDA) type glutamate receptors during synaptic transmission results in significant elevation in intracellular Ca2+ levels, permitting optical detection of synaptic signals within a single spine. Simple calculations suggest that the opening of even a single NMDA receptor would result in the influx of approximately 310 000 Ca2+ ions into the small volume of a spine, producing changes in Ca2+ levels that are readily detectable using high affinity Ca2+ indicators such as fura-2 or fluo-3. Using fluorescent Ca2+ indicators, we have imaged local Ca2+ transients mediated by NMDA receptors in spines and dendritic shafts attributed to spontaneous miniature synaptic activity. Detailed analysis of these quantal events suggests that the current triggering these transients is attributed to the activation of <10 NMDA receptors. The frequency of these miniature synaptic Ca2+ transients is not randomly distributed across synapses, as some synapses can display a >10-fold higher frequency of transients than others. As expected for events mediated by NMDA receptors, miniature synaptic Ca2+ transients were suppressed by extracellular Mg2+ at negative membrane potentials; however, the Mg2+ block could be removed by depolarization.

Animals↗