Contacts among non-sister dendritic branches at bifurcations shape neighboring dendrites and pattern their synaptic inputs.

The size and shape of neuronal dendritic arbors affect the number and pattern of synaptic inputs, as well as the complexity and function of brain circuits. However, the means by which different dendritic arbors take their final shape and how these shapes are associated with distinct synaptic patterns is still largely unknown. Dendritic ramification is influenced by dendrite-dendrite interactions that stabilize specific branching directions and ensure appropriate synaptic contacts. Yet, it is not clear by which mechanism these contacts are allocated. We found that stable dendro-dendritic contacts occur preferentially between non-sister dendritic branches at sites of bifurcations, and that this process is promoted by synaptic activity. Moreover, these contacts are associated with synaptic connections of higher density, higher level of synaptophysin, NR1, GluR2 subunits of glutamate receptors and elevated secretion capability than synaptic connections found on contacts made by non-bifurcating branches or along non-contacting parts of the dendrites. Thus, in cultured neurons, stabilization of hetero-neuronal dendro-dendritic contacts at bifurcations is a new mean to pattern and associate morphogenesis and synaptic input distribution in neighboring dendritic trees.

Convergence among non-sister dendritic branches: an activity-controlled mean to strengthen network connectivity.

The manner by which axons distribute synaptic connections along dendrites remains a fundamental unresolved issue in neuronal development and physiology. We found in vitro and in vivo indications that dendrites determine the density, location and strength of their synaptic inputs by controlling the distance of their branches from those of their neighbors. Such control occurs through collective branch convergence, a behavior promoted by AMPA and NMDA glutamate receptor activity. At hubs of convergence sites, the incidence of axo-dendritic contacts as well as clustering levels, pre- and post-synaptic protein content and secretion capacity of synaptic connections are higher than found elsewhere. This coupling between synaptic distribution and the pattern of dendritic overlapping results in 'Economical Small World Network', a network configuration that enables single axons to innervate multiple and remote dendrites using short wiring lengths. Thus, activity-mediated regulation of the proximity among dendritic branches serves to pattern and strengthen neuronal connectivity.

SGP-1 increases dendritic and synaptic development dependent on synaptic activity.

Neurotrophic factors are a group of secreted proteins which generally regulate neurite outgrowth and synaptic development. SGP-1 has been reported as a neurotrophic factor, though little is known of its effect on neurite outgrowth, and it is unknown whether SGP-1 affects synaptic development. We report here that SGP-1 is distributed in vesicle-like puncta in somas and dendrites of primary neurons in culture, and that SGP-1 is secreted in culture and is taken up by endocytosis in dendrites. Endogenous extracellular activity of SGP-1 promotes dendritic, but not axonal outgrowth. Furthermore, endogenous activity of SGP-1 increases synaptogenesis in hippocampal neurons as determined by measuring the density and size of synaptophysin puncta and by determining the density of dendritic spines, their surface expression of GluR2 and their immunoreactivity for GluR1. The effect of SGP-1 on the amount of postsynaptic receptors in dendritic spines depends on synaptic activity and apparently on activation of MAPK, as inhibition of either of these abolished the affect. Hence, SGP-1 has neurotrophic effects, increasing dendritic growth and promoting synaptic development in an activity-dependent fashion.

Silencing of ZnT-1 expression enhances heavy metal influx and toxicity.

ZnT-1 reduces intracellular zinc accumulation and confers resistance against cadmium toxicity by a mechanism which is still unresolved. A functional link between the L-type calcium channels (LTCC) and ZnT-1 has been suggested, indicating that ZnT-1 may regulate ion permeation through this pathway. In the present study, immunohistochemical analysis revealed a striking overlap of the expression pattern of LTCC and ZnT-1 in cardiac tissue and brain. Using siRNA to silence ZnT-1 expression, we then assessed the role of ZnT-1 in regulating cation permeation through the L-type Ca(2+) channels in cells that are vulnerable to heavy metal permeation. Transfection of cortical neurons with ZnT-1 siRNA resulted in about 70% reduction of ZnT-1 expression and increased Ca(2+) influx via LTCC by approximately fourfold. Moreover, ZnT-1 siRNA transfected neurons showed approximately 30% increase in synaptic release, monitored using the FM1-43 dye. An increased cation influx rate, through the LTCC, was also recorded for Zn(2+) and Cd(2+) in cells treated with the ZnT-1 siRNA. Furthermore, Cd(2+)-induced neuronal death increased by approximately twofold after transfection with ZnT-1 siRNA. In addition, ZnT-1 siRNA transfection of the ovarian granulosa cell line, POGRS1, resulted in a twofold increase in Cd(2+) influx rate via the LTCC. Finally, a robust nimodipine-sensitive Cd(2+) influx was observed using a low extracellular Cd(2+) concentration (5 muM) in neurons and testicular slice cultures, attesting to the relevance of the LTCC pathway to heavy metal toxicity. Taken together, our results indicate that endogenously-expressed ZnT-1, by modulating LTCC, has a dual role: regulating calcium influx, and attenuating Cd(2+) and Zn(2+) permeation and toxicity in neurons and other cell types.

Growth of neurites toward neurite- neurite contact sites increases synaptic clustering and secretion and is regulated by synaptic activity.

The integrative properties of dendrites are determined by several factors, including their morphology and the spatio-temporal patterning of their synaptic inputs. One of the great challenges is to discover the interdependency of these two factors and the mechanisms which sculpt dendrites' fine morphological details. We found a novel form of neurite growth behavior in neuronal cultures of the hippocampus and cortex, when axons and dendrites grew directly toward neurite-neurite contact sites and crossed them, forming multi-neurite intersections (MNIs). MNIs were found at a frequency higher than obtained by computer simulations of randomly distributed dendrites, involved many of the dendrites and were stable for days. They were formed specifically by neurites originating from different neurons and were extremely rare among neurites of individual neurons or among astrocytic processes. Axonal terminals were clustered at MNIs and exhibited higher synaptophysin content and release capability than in those located elsewhere. MNI formation, as well as enhancement of axonal terminal clustering and secretion at MNIs, was disrupted by inhibitors of synaptic activity. Thus, convergence of axons and dendrites to form MNIs is a non-random activity-regulated wiring behavior which shapes dendritic trees and affects the location, clustering level and strength of their presynaptic inputs.