Gene Networks Activated by Specific Patterns of Action Gene Networks Activated by Specific Patterns of Action Potentials in Dorsal Root Ganglia Neurons Potentials in Dorsal Root Ganglia Neurons

Gene regulatory networks underlie the long-term changes in cell specification, growth of synaptic connections, and adaptation that occur throughout neonatal and postnatal life. Here we show that the transcriptional response in neurons is exquisitely sensitive to the temporal nature of action potential firing patterns. Neurons were electrically stimulated with the same number of action potentials, but with different inter-burst intervals. We found that these subtle alterations in the timing of action potential firing differentially regulates hundreds of genes, across many functional categories, through the activation or repression of distinct transcriptional networks. Our results demonstrate that the transcriptional response in neurons to environmental stimuli, coded in the pattern of action potential firing, can be very sensitive to the temporal nature of action potential delivery rather than the intensity of stimulation or the total number of action potentials delivered. These data identify temporal kinetics of action potential firing as critical components regulating intracellular signalling pathways and gene expression in neurons to extracellular cues during early development and throughout life. Adaptation in the nervous system in response to external stimuli requires synthesis of new gene products in order to elicit long lasting changes in processes such as development, response to injury, learning, and memory 1 . Information in the environment is coded in the pattern of action-potential firing, therefore gene transcription must be regulated by the pattern of neuronal firing. Such transcriptional regulation is unique to neurons. The mechanisms that could be responsible for regulating gene expression by the temporal pattern of neuronal firing are largely unknown. Whether such regulation is limited to a few genes having special properties or extends to large numbers of genes by virtue of the dynamic response of intracellular signalling networks and cellular systems influencing transcription is unknown. How responsive the transcriptome is to very subtle alterations in patterns of action potentials remains an open, important question. There is very little data on the genomic response to the temporal features of action potential firing, where timing of action potential bursting is variable and the total delivery of action potentials over a given time period constant. Pharmacological manipulations, e.g., kainate 2,3 , pilocarpine 4 , NMDA 5,6 , KCl 7 , bicuculline 8 and BDNF 7 have been used to induce membrane depolarization followed by transcriptome analysis. These studies have detailed differential mRNA abundance, transcription factor regulation, regulatory sequences, chromatin occupancy of gene regulatory sites, and gene networks activated or repressed in neurons upon stimulation. However, a pharmacological approach induces stimulation that lacks temporal control of action potential firing patterns, which is critical during neural development and plasticity 6,8,9 . In contrast, altering the pattern of the same number of action potentials by using electrical stimulation produces a more naturalistic stimulus required to address the question of whether the abundance of RNAs and transcriptional networks are differentially sensitive to neuronal firing patterns. We hypothesized that the temporal dynamics of action potentials would regulate the expression of hundreds of genes across broad functional categories 10 . In order to test the hypothesis that global mRNA abundance in neurons is dependent upon the specific pattern of action potential firing, mouse DRG neurons were electrically stimulated in cell culture with two distinct patterns of activation. These two patterns delivered the same total number of action potentials, but they were grouped into two different durations of firing at 10 Hz with different inter-burst 1 Nervous System Development and Plasticity Section, the Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), NIH, Bethesda, MD, USA. 2 Department of Pathology, New York Medical College, Valhalla, NY, USA. 3 DP Purpura Department of Neuroscience, Albert Einstein College of Medicine, New York, NY, USA. * These authors contributed equally to this work. Correspondence and requests for materials should be addressed to R.D. F. (email: fieldsd