Activity-dependent upregulation of presynaptic kainate receptors at immature CA3-CA1 synapses.

Presynaptic kainate-type glutamate receptors (KARs) regulate glutamate release probability and short-term plasticity in various areas of the brain. Here we show that long-term depression (LTD) in the area CA1 of neonatal rodent hippocampus is associated with an upregulation of tonic inhibitory KAR activity, which contributes to synaptic depression and causes a pronounced increase in short-term facilitation of transmission. This increased KAR function was mediated by high-affinity receptors and required activation of NMDA receptors, nitric oxide (NO) synthetase, and postsynaptic calcium signaling. In contrast, KAR activity was irreversibly downregulated in response to induction of long-term potentiation in a manner that depended on activation of the TrkB-receptor of BDNF. Both tonic KAR activity and its plasticity were restricted to early stages of synapse development and were lost in parallel with maturation of the network due to ongoing BDNF-TrkB signaling. These data show that presynaptic KARs are targets for activity-dependent modulation via diffusible messengers NO and BDNF, which enhance and depress tonic KAR activity at immature synapses, respectively. The plasticity of presynaptic KARs in the developing network allows nascent synapses to shape their response to incoming activity. In particular, upregulation of KAR function after LTD allows the synapse to preferentially pass high-frequency afferent activity. This can provide a potential rescue from synapse elimination by uncorrelated activity and also increase the computational dynamics of the developing CA3-CA1 circuitry.

Transsynaptic expression of a presynaptic glutamate receptor during hippocampal long-term potentiation.

Repetitive activation of excitatory synapses in the hippocampus produces a persistent enhancement of synaptic efficiency known as long-term potentiation (LTP). In anesthetized and in freely moving rats, the induction of LTP in the perforant path led to a transient increase in the amount of messenger RNA (mRNA) coding for a presynaptic glutamate receptor (GR33) in dentate granule cells. The amount of GR33 mRNA was increased for at least 5 hours after the induction of LTP but was indistinguishable from control values 1 day after induction. The N-methyl-D-aspartate receptor antagonist 2-aminophosphonovalerate prevented the induction of both LTP and the increase in GR33 mRNA. The amount of GR33 protein was increased in the mossy fiber terminal zone of dentate granule cells 5 hours after the induction of LTP. These results suggest that the induction of LTP in synapses at one stage in a neural network may lead to modification in synaptic function at the next stage in the network.

A Presynaptic Regulatory System Acts Transsynaptically via Mon1 to Regulate Glutamate Receptor Levels in Drosophila.

Mon1 is an evolutionarily conserved protein involved in the conversion of Rab5 positive early endosomes to late endosomes through the recruitment of Rab7. We have identified a role for Drosophila Mon1 in regulating glutamate receptor levels at the larval neuromuscular junction. We generated mutants in Dmon1 through P-element excision. These mutants are short-lived with strong motor defects. At the synapse, the mutants show altered bouton morphology with several small supernumerary or satellite boutons surrounding a mature bouton; a significant increase in expression of GluRIIA and reduced expression of Bruchpilot. Neuronal knockdown of Dmon1 is sufficient to increase GluRIIA levels, suggesting its involvement in a presynaptic mechanism that regulates postsynaptic receptor levels. Ultrastructural analysis of mutant synapses reveals significantly smaller synaptic vesicles. Overexpression of vglut suppresses the defects in synaptic morphology and also downregulates GluRIIA levels in Dmon1 mutants, suggesting that homeostatic mechanisms are not affected in these mutants. We propose that DMon1 is part of a presynaptically regulated transsynaptic mechanism that regulates GluRIIA levels at the larval neuromuscular junction.

Lipid bilayer membrane-triggered presynaptic vesicle assembly.

The formation of functional synapses on artificial substrates is a very important step in the development of engineered in vitro neural networks. Spherical supported bilayer lipid membranes (SS-BLMs) are used here as a novel substrate to demonstrate presynaptic vesicle accumulation at an in vitro synaptic junction. Confocal fluorescence microscopy, cryo-transmission electron microscopy (cryo-TEM), and fluorescence recovery after photobleaching (FRAP) experiments have been used to characterize the SS-BLMs. Conventional immunocytochemistry combined with confocal fluorescence microscopy was used to observe the formation of presynaptic vesicles at the neuron-SS-BLM contacts. These results indicate that lipid phases may play a role in the observed phenomenon, in addition to the chemical and electrostatic interactions between the neurons and SS-BLMs. The biocompatibility of lipid bilayers along with their membrane tunability makes the suggested approach a useful "toolkit" for many neuroengineering applications including artificial synapse formation and synaptogenesis in vivo.

Development of P2X receptor clusters on smooth muscle cells in relation to nerve varicosities in the rat urinary bladder.

Postnatal development of the distribution of different isoforms of purinergic (P2X) receptors on smooth muscle cells in relation to the development of the innervation of the cells by nerve varicosities in the rat urinary bladder has been determined with immunofluorescence and confocal microscopy. Antibodies against the extracellular domains of the P2X(1) to P2X(6) receptors were used to detect the receptors in the bladder. Several other antibodies were used to identify sympathetic varicosities and Schwann cells. At one day postnatal (D1) there were few strings of varicosities denoting isolated axons, with most axons confined to large nerve trunks. Small size clusters of P2X(1) to P2X(6) receptor subtypes (about 0.4 microm diameter) were observed in the muscle which were independent of each other, and sometimes juxtaposed to the rare isolated varicosity strings. At D4 large numbers of strings of varicosities could be discerned throughout the detrusor. Most of these clouds of small P2X(1) to P2X(6) receptor clusters in their immediate vicinity. Some of these were colocalised with the varicosities, which were of parasympathetic origin as they failed to counter-stain with antibodies to tyrosine hydroxylase. Up to D14 there was a gradual coalescence of many of the isolated P2X(1-6) small receptor clusters so that they became colocalized, often at varicosities. Most of the varicosities in isolated strings possessed receptor clusters at this time. By D21 it was rare to find varicosity strings in the detrusor that were not either in close juxtaposition with P2X small receptor clusters or possessing such clusters in colocalization. However, large numbers of small P2X receptor clusters, many of which consisted of a mixture of isoforms, could be found spatially unrelated to nerve varicosities throughout the detrusor muscle. In the adult, single axons were either coextensive with one or more isoforms of P2X receptor clusters or these were immediately juxtaposed to the axons so that is was rare to find a varicosity that did not possess a receptor cluster. However, different combinations of colocalized P2X receptor isoforms could still be discerned in small clusters unrelated to varicosities. These observations are discussed in relation to the mechanism of formation of the receptor clusters and their migration beneath parasympathetic varicosities during development.