Modeling Neurotransmitter Specification in Neural Progenitor Cells

William Fisher1,2, Adele Doyle1,2,3

1 Neuroscience Research Institute, 2College of Creative Studies, 3Center for BioEngineering University of California Santa Barbara

Figure 1. Individual Neurotransmitter Specification Networks. These figures summarize the molecular interactions involved in individual neurotransmitter specification, based on the meta analysis of the studies referenced below. The type of protein is indicated by node color and the molecule type is indicated by node shape. See legend (methods section) for details.

IntroductionAbstract

Results

References

Discussion and Conclusions • Based on this meta analysis of experimental data, many molecules and molecule types are

needed to coordinate neurotransmitter (NT) identity (Fig. 1).• Many regulatory molecules can affect the expression of multiple neurotransmitters. This

molecular regulation may occur via changes in biosynthetic enzyme expression or as molecular switches. (Fig. 2-4)

• Ongoing work will incorporate the available quantitative data into a mathematical model to describe NT expression. Matlab-based simulations using this model will be used to generate experimentally-testable predictions about the control of NT expression.

• This work may ultimately help direct differentiation of neurons towards particular neurotransmitter phenotypes for use in neural stem cell therapies.

Figure 3. Shared Signaling. Building individual models from the literature highlighted molecules that regulate multiple different neurotransmitter fates. This figure summarizes transcription factors known to affect expression of multiple neurotransmitters.

Although numerous molecular markers exist to track neuronal differentiation in vivo and in vitro, the molecular networks that control neurotransmitter expression are not well understood. Discerning how neural progenitor cells spontaneously make this fate decision can improve our ability to controllably generate neurons expressing specific neurotransmitters. To understand the regulatory networks that control neurotransmitter identity in nascent neurons, we synthesized data from previously published studies to create putative molecular interaction networks describing the onset of expression of eight well-known neurotransmitters. These preliminary interaction networks suggest that the expression and interaction of several transcription factors influence specification of progenitor cells into particular neuron subtypes. Signaling regulators of embryonic development including growth factors and retinoic acid are also associated with neurotransmitter fate choice. These models also highlighted molecules that may act as switches in fate choice between two or more neurotransmitter phenotypes, such as GABAergic and Glutamatergic neurons. Finally, this metadata analysis revealed substantial shared signaling between circuits responsible for different neurotransmitter fates. Ongoing work is focused on translating the qualitative regulatory networks into predictive mathematical models of neurotransmitter specification, suitable for in vitro testing and validation. A quantitative predictive model of the regulatory networks responsible for neurotransmitter fate decisions could ultimately help generate neurons expressing specific neurotransmitters to aid patients with neurodegenerative diseases.

Figure 4. Fate Switches. A subset of molecules regulates expression of more than one neurotransmitter (left panel: GABAergic/Glutamatergic, right panel: GABAergic/Cholinergic). This meta analysis suggests these molecules may participate as switches (e.g., Ascl1) to control neurotransmitter expression, and the subsequent neuron subtype. In the cases identified, expression of the switch molecule above a threshold concentration represses expression of one neurotransmitter while driving the expression of another.

Materials and Methods

• Stem cell therapies targeting specific neuron subtypes (i.e., neurotransmitter phenotype) have the potential to improve treatment of neurodegenerative diseases [CITE].

• Currently there exist numerous protocols to direct the differentiation of neurons from progenitor cells [CITE], as well a limited number of reports describing molecules that are necessary or sufficient to specify particular neurotransmitter types [CITE]. However, it is not known how these varied protocols and reported influential molecules work together as a regulatory circuit to govern neuron subtype fate.

• Therefore, to better understand the regulatory architecture of neuron subtype specification, we have integrated data from published studies that describe the interactions of one or a few molecules in driving a particular neurotransmitter fate into a consensus model for individual neurotransmitter expression.

Figure 2. Biosynthetic Pathways. Melatonergic fate is defined by expression of enzymes (red bars) required for biosynthesis of respective neurotransmitters (NT). In some cases, molecules regulate NT expression by acting on these enzymes. Noradrenergic and adrenergic fates are not shown separately here because molecules regulating these NTs are selected based on their effect on the single key biosynthetic enzyme (either DBH or PNMT, respectively).

• Searched PubMed and Web of Science for reviews and peer-reviewed primary literature describing specification of neurotransmitter (NT) fates. Recorded search prompts, dates, and sources.

• Next, we identified experimental evidence supporting the role of each cited molecular marker in NT fate specification, as well as: • molecule type, molecular

mechanism, and interactions with other molecules in network

• Created qualitative network diagrams for each NT summarizing the putative regulatory molecules and their interactions.

Adrenergic: [1] Tai TC, Journ of Neurochem, 2001 [2] Tai TC Journ of Neurochem, 2009 [3] Tai TC Journ of Neurochem, 2010 [4] Tai TC, Molec Pharm, 2010 [5] Wong DL, Annals New York Acad Sci, 2006 [6] Her S, Molec Pharm, 2003 [7] Evinger MJ, Journ Molec Neurosci, 2005 [8] Rodriguez-Flores JL, Mammal Genome, 2010Noradrenergic: [1] Pattyn A, Molec and Cell Neurosci, 2000 [2] Howard M, Dev Bio, 2000 [3] Guo S, Neuron, 1999 [4] Kim HS, Journ of Neurochem, 2001 [5] Qian Y, Genes and Dev, 2001 [6] Hirsch MR, Development, 1998 [7] Tsarovina K, The Journ of Neurosci, 2010 [8] Benjanirut C, Journ of Bio Chem, 2005 [9] Hsieh M, Journ of Necurochem, 2005 [10] Rylich J, Journ Bio Chem, 2003 [11] Liu H, Dev Bio, 2005 Cholinergic: [1] Bisonette, Stem Cells, 2011 [2] Golden, Exp Neurolog, 2003 [3] Manabe, Cell Death and Diff, 2007 [4] Nilbratt, Journ Cell Molec Med, 2009 [5] Asbreuk, Neurosci, 2002 [6] Lopes, PNAS, 2012 [7] Cho, PLoS Genetics, 2014 [8] Fragkouli, Dev, 2009 [9] The Joun of Neurosci, 2010 [10] Zhao, Dev Bio, 2014 Dopaminergic: [1] Kwon, Stem Cell Res, 2014 [2] Rousa, Stem Cell, 2006 [3] Mesman, PLoS One, 2014 [5] Robler, Neurosci, 2010 [6] Anderson, Cell, 2006 [7] Veenvliet, Dev, 2013 [9] Frank M, Dev, 2009 [10] Chung, PNAS, 2011 [11] Zi, Stem Cells, 2012 [12] Doi, Stem Cell Rep, 2014 [13] Nakatani, Dev Bio, 2009 [14] Sakane, Dev neurobio, 2013 [15] Omodei, Dev, 2008 [16] Rabe, Neurodev, 2012 [17] Liu, Cell Res, 2012 [19] Anderson, Exp Cell Res, 2006 [20] Lin, Dev Bio, 2009 [21] Jacobs, Dev, 2007 GABA/Glutamatergic: [2] Sellers, Neuro Dev, 2014 [3] Hori, Neuroplast, 2012 [5] Yamada, The Journ of Neurosci, 2014 [6] Puelles, The Journ of Neurosci, 2006 [7] Blum, Cereb Cort, 2011 [8] Nakatani, Dev, 2007 [9] Chen, PLoS One, 2012 [10] Roybon, Cereb Cort, 2009 [11] Pillai, Dev, 2007 [12] Xiang, Somat and Motor Res, 2012 [13] Batista, Dev Bio, 2008 [14] Cheng, Nat Neurosci, 2005 [15] Pozas, Neuron, 2005 [16] Canty, the Journ of Neurosci, 2009 [18] Poitras, Dev, 2007 Serotonergic: [1] Brisco, Nature, 1999 [2] Carcagno, The Journ of Neurosci, 2014 [3] Cheng, Journ of Neurosci, 2003 [4] Hendricks, Cell Press, 2003 [5] Jacob, nat Neurosci, 2007 [6] Pattyn, Nat Neurosci, 2004 [8] Cheng CW, Dev Dynamics, 2007 [9] Song NN, PLoS One, 2011 [10] Dolmazon V, Stem Cells and Dev, 2011 [11] Horst HS, Molec and Cell Neurosci, 2004 [12] Craven S, Dev, 2003 [13] Todd KJ, PLoS One, 2012 [14] O’Reilly KC, Exp Bio and Med, 2007 [15] Jacob J, Dev, 2009 [16] Terahoka H, Journ of Neurobio, 2009 Histaminergic: [1] Blandina, Sys Neurosci, 2012 [2] Sundvik, Journ of Neurosci, 2013 [3] Valko, Annals of Neurology, 2014 [4] Molina-Hernandez, Journ of Neurochem, 2008 [5] Molyva, Pharm Rep, 2014 [6] Miyoshi, Journ of Pharm Neurosci, 2006 [7] Otani, Neurosci, 2008 Melatonergic: [1] Maronde, The Journ of Neurosci, 1999 [2] Dinet, Exp Eye Res, 2006 [3] Li, PNAS, 1998 [4] Bernard, Journ of Neurochem, 2001

Regulatory ComplexTranscription FactorsEnzymeCellular Signaling ProteinCell Adhesion MoleculeGrowth FactorsTransporter Protein

Small Molecules

Weaker Evidenced InteractionUpregulationRepressionPhosphorylation

HexamerTrimer

Dimer

Legend