Transcriptional responses of cultured rat sympathetic neurons during BMP-7-induced dendritic growth.

Michelle M Garred,Michael M Wang,Pamela J Lein,Christina A Harrington,Xin Guo,Ferdinando Di Cunto

doi:10.1371/journal.pone.0021754

Abstract

BackgroundDendrites are the primary site of synapse formation in the vertebrate nervous system; however, relatively little is known about the molecular mechanisms that regulate the initial formation of primary dendrites. Embryonic rat sympathetic neurons cultured under defined conditions extend a single functional axon, but fail to form dendrites. Addition of bone morphogenetic proteins (BMPs) triggers these neurons to extend multiple dendrites without altering axonal growth or cell survival. We used this culture system to examine differential gene expression patterns in naïve vs. BMP-treated sympathetic neurons in order to identify candidate genes involved in regulation of primary dendritogenesis.Methodology/Principal FindingsTo determine the critical transcriptional window during BMP-induced dendritic growth, morphometric analysis of microtubule-associated protein (MAP-2)-immunopositive processes was used to quantify dendritic growth in cultures exposed to the transcription inhibitor actinomycin-D added at varying times after addition of BMP-7. BMP-7-induced dendritic growth was blocked when transcription was inhibited within the first 24 hr after adding exogenous BMP-7. Thus, total RNA was isolated from sympathetic neurons exposed to three different experimental conditions: (1) no BMP-7 treatment; (2) treatment with BMP-7 for 6 hr; and (3) treatment with BMP-7 for 24 hr. Affymetrix oligonucleotide microarrays were used to identify differential gene expression under these three culture conditions. BMP-7 significantly regulated 56 unique genes at 6 hr and 185 unique genes at 24 hr. Bioinformatic analyses implicate both established and novel genes and signaling pathways in primary dendritogenesis.Conclusions/SignificanceThis study provides a unique dataset that will be useful in generating testable hypotheses regarding transcriptional control of the initial stages of dendritic growth. Since BMPs selectively promote dendritic growth in central neurons as well, these findings may be generally applicable to dendritic growth in other neuronal cell types.

Highlights

The shape of the dendritic arbor determines the total synaptic input a neuron can receive [1,2,3], and influences the types and distribution of these inputs [4,5,6]
bone morphogenetic proteins (BMPs)-7 triggers dendritogenesis in cultured sympathetic neurons via transcriptional mechanisms
To determine the critical period when transcriptional changes required for the dendritic response to BMP-7 occurred, transcription was inhibited by adding actinomycin-D to cultures at varying times after BMP-7 addition

Summary

Introduction

The shape of the dendritic arbor determines the total synaptic input a neuron can receive [1,2,3], and influences the types and distribution of these inputs [4,5,6]. Altered patterns of dendritic growth and plasticity are associated with impaired neurobehavioral function in experimental models [7], and are thought to contribute to clinical symptoms observed in both neurodevelopmental disorders [8,9,10] and neurodegenerative diseases [11,12,13]. Addition of bone morphogenetic proteins (BMPs) triggers these neurons to extend multiple dendrites without altering axonal growth or cell survival We used this culture system to examine differential gene expression patterns in naıve vs BMP-treated sympathetic neurons in order to identify candidate genes involved in regulation of primary dendritogenesis

Methods

Results

Discussion

Conclusion