New research from a collaboration between the Harris and Ehlers laboratories shows that the mobility of cargo along the smooth endoplasmic reticulum (SER) of hippocampal dendrites is reduced where the SER is highly elaborated, particularly along spiny segments and at dendritic branch points. The primary objective of this research proposal is to investigate whether the SER network within developing dendrites is locally regulated in the vicinity of spines and synapses. We will examine changes in dendritic SER organization during a period of robust synaptogenesis from postnatal day (P) 15 to P21. During this one postnatal week, oblique dendrites mature substantially in spine number and synapse size.
Aim 1 will examine how the dendritic SER accommodates new spine formation during normal development. We will test the hypothesis that the local organization of dendritic SER becomes more elaborate as spine density and synapse size increase from P15 to P21. To test this hypothesis, we will use the nanometer resolution of serial section electron microscopy to make three-dimensional (3D) reconstructions of dendrites, synapses, and the dendritic SER network of perfusion-fixed hippocampus in vivo at P15 and P21. To estimate an index of local elaboration, we will compare regions of SER that form complex SER cisterns to regions that mainly consist of simple tubules along spiny and spine-free segments of dendrites. We will also induce long-term potentiation (LTP) in hippocampal slices (as a cellular model of learning) to investigate changes in the organization of dendritic SER during synaptic plasticity. We will use theta-burst stimulation (TBS) to study multiple phases of LTP. The AMPA receptor-dependent, early-phase LTP (E-LTP) lasts up to an hour, while the protein-synthesis dependent, late- phase LTP (L-LTP) endures for more than 3 hrs.
Aim 2 will test whether dendritic SER undergoes plasticity- dependent elaboration during E-LTP.
Aim 3 will test the hypothesis that plasticity-dependent elaboration of dendritic SER is reduced during L-LTP relative to E-LTP. In these experiments, slices that received TBS (1 each for E-LTP and L-LTP) will be compared to a slice that only received control-stimulation. After expression of LTP, we will compare spine density and synapse size between control and TBS dendrites at P15 and P21, and evaluate whether the organization of the dendritic SER near spines is altered relative to the spine-free portions of the dendritic shaft in these slices. LTP-specific effects will be controlled for by administering APV, a competitive NMDA receptor antagonist known to block LTP. The implications of these findings will establish a comprehensive description of the role of SER in the development and plasticity of spines and synapses at a postnatal age where developmental disorders often manifest dendritic abnormalities. The goals put forth in this application serve the mission of a variety of NIH agencies that use basic research to understand the evolving science of brain, behavior, and experience (e.g., NIMH and NICHD) and will help to identify potential therapeutic targets for developmental disorders associated with malformed spines and dysfunctional SER.
The results obtained from this proposal will expand our knowledge of how the smooth endoplasmic reticulum (SER) modifies its organization to support dendritic spine and synapse structure during normal development and synaptic plasticity. We must have a complete understanding of the mechanisms that control how dendritic resources like the SER support synapse development in normal dendrites before we can understand what makes these processes go awry in diseased dendrites.