Several technical aspects of our experiments were essential selleck chemical to drawing our conclusions. One is that we were able to compare synaptic density and ultrastructural features of connections onto stable and extending dendritic branches within the same dendritic arbors. Consequently, it is clear that differences in synapse density and maturation on stable and dynamic branches do not arise from

heterogeneity of the postsynaptic neurons. This analysis also allows us to conclude that mature synapses are found preferentially on stable dendritic branches. Second, we were able to compare connectivity of presynaptic boutons as they relate to the dynamics of axon branches. This demonstrated that the reduced divergence from MSBs and the decreased convergence onto stable dendrites seen in this study are not necessarily accompanied by large-scale changes in axonal or dendritic arbor structure and would not have been detected without the combined use of in vivo time-lapse

imaging to distinguish stable and dynamic branches and the spatial resolution of EM. High-density clusters of immature synapses on newly extended dendrites would be difficult Neratinib research buy to distinguish from fewer more mature synapses on stable dendrites based on fluorescent light microscopy of synaptic markers. Similarly, because the distances between individual synaptic contacts within a MSB are less than 1 μm, the gain or loss of contacts from MSBs occurs at a suboptical resolution and

may have been underestimated in previous light microscope based studies (Alsina et al., 2001, Meyer and Smith, 2006 and Ruthazer et al., 2006). Third, we have been able to make Megestrol Acetate direct comparisons between the synaptic rearrangements that occur over a 24 hr time interval and a 4 hr interval, which indicate that synapse formation, maturation, and elimination occur over a time scale of hours during activity-dependent microcircuit development in vivo. Consequently, our experiments provide direct evidence for a previously unrecognized role for synaptic dynamics and synapse elimination in fine-scale circuit development. The potential role of synaptic connections in regulating the elaboration of neuronal structure has been proposed by Vaughn (1989) in the synaptotrophic model of neuronal development (Vaughn, 1989), which states that formation of synaptic connections stabilize pre- and postsynaptic neuronal branches and promote further growth of the neuronal arbor. Studies in which synaptic activity was shown to regulate neuronal arbor development provide support for the synaptotrophic hypothesis (Cline and Haas, 2008); however, other studies suggested that neuronal development can occur without synaptic transmission (Verhage et al., 2000).