Frontiers in Biology

, Volume 5, Issue 6, pp 524–531 | Cite as

Advances in genomic study of cortical projection neurons



The mammalian neocortex gives rise to perception and initiates voluntary motor responses. The cortical laminae are comprised of six distinct cellular layers of local circuit neurons and projection neurons. To explore molecular identities of the distinct cortical projection neurons, discovery-orientated genomic approaches have been adopted. Microarray analysis of dissected cortical tissues has been applied to identify cortical layer markers. Early neuronal cells were sorted by FACS from GFPlabeled embryonic brains for gene expression profiling. Laser capture microdissection of retrograde-labeled projection neurons, when coupled with optimal RNA amplification technology, has become a valuable strategy for neuronal isolation and gene expression analysis in differentiated neurons. RNA sequencing technology is promising not only for the determination of gene expression, but also for discovery of posttranscriptional modifications of the complex neural system. There is no doubt that advances in genomic studies are opening up novel research avenues for our understanding of the cortical neuronal functions.


expression profiling pyramidal neurons RNA amplification cortex retrograde labeling 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Arlotta P, Molyneaux B J, Chen J, Inoue J, Kominami R, Macklis J D (2005). Neuronal subtype-specific genes that control corticospinal motor neuron development in vivo. Neuron, 45(2): 207–221CrossRefPubMedGoogle Scholar
  2. Bertone P, Gerstein M, Snyder M (2005). Applications of DNA tiling arrays to experimental genome annotation and regulatory pathway discovery. Chromosome Res, 13(3): 259–274CrossRefPubMedGoogle Scholar
  3. Böhm C, Newrzella D, Sorgenfrei O (2005). Laser microdissection in CNS research. Drug Discov Today, 10(17): 1167–1174CrossRefPubMedGoogle Scholar
  4. Bunney W E, Bunney B G, Vawter M P, Tomita H, Li J, Evans S J, Choudary P V, Myers R M, Jones E G, Watson S J, Akil H (2003). Microarray technology: a review of new strategies to discover candidate vulnerability genes in psychiatric disorders. Am J Psychiatry, 160(4): 657–666CrossRefPubMedGoogle Scholar
  5. Cahoy J D, Emery B, Kaushal A, Foo L C, Zamanian J L, Christopherson K S, Xing Y, Lubischer J L, Krieg P A, Krupenko S A, Thompson W J, Barres B A (2008). A transcriptome database for astrocytes, neurons, and oligodendrocytes: a new resource for understanding brain development and function. J Neurosci, 28(1): 264–278CrossRefPubMedGoogle Scholar
  6. Chen J G, Rasin M R, Kwan K Y, Sestan N (2005). Zfp312 is required for subcortical axonal projections and dendritic morphology of deeplayer pyramidal neurons of the cerebral cortex. Proc Natl Acad Sci USA, 102(49): 17792–17797CrossRefPubMedGoogle Scholar
  7. Chow N, Cox C, Callahan L M, Weimer J M, Guo L, Coleman P D (1998). Expression profiles of multiple genes in single neurons of Alzheimer’s disease. Proc Natl Acad Sci USA, 95(16): 9620–9625CrossRefPubMedGoogle Scholar
  8. Clément-Ziza M, Gentien D, Lyonnet S, Thiery J P, Besmond C, Decraene C (2009). Evaluation of methods for amplification of picogram amounts of total RNA for whole genome expression profiling. BMC Genomics, 10(1): 246CrossRefPubMedGoogle Scholar
  9. Colosimo M E, Brown A, Mukhopadhyay S, Gabel C, Lanjuin A E, Samuel A D, Sengupta P (2004). Identification of thermosensory and olfactory neuron-specific genes via expression profiling of single neuron types. Curr Biol, 14(24): 2245–2251CrossRefPubMedGoogle Scholar
  10. Cui D, Dougherty K J, Machacek D W, Sawchuk M, Hochman S, Baro D J (2005). Divergence between motoneurons: gene expression profiling provides a molecular characterization of functionally discrete somatic and autonomic motoneurons. Physiol Genomics, 24(3): 276–289CrossRefPubMedGoogle Scholar
  11. Eberwine J (2001). Single-cell molecular biology. Nat Neurosci, 4(Suppl): 1155–1156CrossRefPubMedGoogle Scholar
  12. Eberwine J, Yeh H, Miyashiro K, Cao Y, Nair S, Finnell R, Zettel M, Coleman P (1992). Analysis of gene expression in single live neurons. Proc Natl Acad Sci USA, 89(7): 3010–3014CrossRefPubMedGoogle Scholar
  13. Emmert-Buck M R, Bonner R F, Smith P D, Chuaqui R F, Zhuang Z, Goldstein S R, Weiss R A, Liotta L A (1996). Laser capture microdissection. Science, 274(5289): 998–1001CrossRefPubMedGoogle Scholar
  14. Espina V, Wulfkuhle J D, Calvert V S, VanMeter A, Zhou W, Coukos G, Geho D H, Petricoin E F 3rd, Liotta L A (2006). Laser-capture microdissection. Nat Protoc, 1(2): 586–603CrossRefPubMedGoogle Scholar
  15. Evans S J, Choudary P V, Vawter M P, Li J, Meador-Woodruff J H, Lopez J F, Burke S M, Thompson R C, Myers R M, Jones E G, Bunney W E, Watson S J, Akil H (2003). DNA microarray analysis of functionally discrete human brain regions reveals divergent transcriptional profiles. Neurobiol Dis, 14(2): 240–250CrossRefPubMedGoogle Scholar
  16. Fox R M, Von Stetina S E, Barlow S J, Shaffer C, Olszewski K L, Moore J H, Dupuy D, Vidal M, Miller D M 3rd (2005). A gene expression fingerprint of C. elegans embryonic motor neurons. BMC Genomics, 6(1): 42Google Scholar
  17. Geschwind D H, Levitt P (2007). Autism spectrum disorders: developmental disconnection syndromes. Curr Opin Neurobiol, 17(1): 103–111CrossRefPubMedGoogle Scholar
  18. Ginsberg S D, Che S, Counts S E, Mufson E J (2006). Shift in the ratio of three-repeat tau and four-repeat tau mRNAs in individual cholinergic basal forebrain neurons in mild cognitive impairment and Alzheimer’s disease. J Neurochem, 96(5): 1401–1408CrossRefPubMedGoogle Scholar
  19. Ginsberg S D, Hemby S E, Lee V M, Eberwine J H, Trojanowski J Q (2000). Expression profile of transcripts in Alzheimer’s disease tangle-bearing CA1 neurons. Ann Neurol, 48(1): 77–87CrossRefPubMedGoogle Scholar
  20. Gong S, Zheng C, Doughty ML, Losos K, Didkovsky N, Schambra U B, Nowak N J, Joyner A, Leblanc G, Hatten M E, Heintz N (2003). A gene expression atlas of the central nervous system based on bacterial artificial chromosomes. Nature, 425(6961): 917–925CrossRefPubMedGoogle Scholar
  21. Gray P A, Fu H, Luo P, Zhao Q, Yu J, Ferrari A, Tenzen T, Yuk D I, Tsung E F, Cai Z, Alberta J A, Cheng L P, Liu Y, Stenman J M, Valerius M T, Billings N, Kim H A, Greenberg M E, McMahon A P, Rowitch D H, Stiles C D, Ma Q (2004). Mouse brain organization revealed through direct genome-scale TF expression analysis. Science, 306(5705): 2255–2257CrossRefPubMedGoogle Scholar
  22. Greenberg S A (2001). DNA microarray gene expression analysis technology and its application to neurological disorders. Neurology, 57(5): 755–761PubMedGoogle Scholar
  23. Harel N Y, Strittmatter S M (2006). Can regenerating axons recapitulate developmental guidance during recovery from spinal cord injury? Nat Rev Neurosci, 7(8): 603–616CrossRefPubMedGoogle Scholar
  24. Heintz N (2000). Analysis of mammalian central nervous system gene expression and function using bacterial artificial chromosome-mediated transgenesis. Hum Mol Genet, 9(6): 937–943CrossRefPubMedGoogle Scholar
  25. Hoerder-Suabedissen A, Wang W Z, Lee S, Davies K E, Goffinet A M, Rakić S, Parnavelas J, Reim K, Nicolić M, Paulsen O, Molnár Z (2009). Novel markers reveal subpopulations of subplate neurons in the murine cerebral cortex. Cereb Cortex, 19(8): 1738–1750CrossRefPubMedGoogle Scholar
  26. Jiang Y M, Yamamoto M, Kobayashi Y, Yoshihara T, Liang Y, Terao S, Takeuchi H, Ishigaki S, Katsuno M, Adachi H, Niwa J, Tanaka F, Doyu M, Yoshida M, Hashizume Y, Sobue G (2005). Gene expression profile of spinal motor neurons in sporadic amyotrophic lateral sclerosis. Ann Neurol, 57(2): 236–251CrossRefPubMedGoogle Scholar
  27. Johnson M B, Kawasawa Y I, Mason C E, Krsnik Z, Coppola G, Bogdanović D, Geschwind D H, Mane S M, State M W, Sestan N (2009). Functional and evolutionary insights into human brain development through global transcriptome analysis. Neuron, 62(4): 494–509CrossRefPubMedGoogle Scholar
  28. Kamme F, Salunga R, Yu J, Tran D T, Zhu J, Luo L, Bittner A, Guo H Q, Miller N, Wan J, Erlander M (2003). Single-cell microarray analysis in hippocampus CA1: demonstration and validation of cellular heterogeneity. J Neurosci, 23(9): 3607–3615PubMedGoogle Scholar
  29. Khaitovich P, Muetzel B, She X, Lachmann M, Hellmann I, Dietzsch J, Steigele S, Do H H, Weiss G, Enard W, Heissig F, Arendt T, Nieselt-Struwe K, Eichler E E, Pääbo S (2004). Regional patterns of gene expression in human and chimpanzee brains. Genome Res, 14(8): 1462–1473CrossRefPubMedGoogle Scholar
  30. Kurn N, Chen P, Heath J D, Kopf-Sill A, Stephens KM, Wang S (2005). Novel isothermal, linear nucleic acid amplification systems for highly multiplexed applications. Clin Chem, 51(10): 1973–1981CrossRefPubMedGoogle Scholar
  31. Leamey C A, Glendining K A, Kreiman G, Kang N D, Wang K H, Fassler R, Sawatari A, Tonegawa S, Sur M (2007). Differential gene expression between sensory neocortical areas: potential roles for Ten_m3 and Bcl6 in patterning visual and somatosensory pathways. Cereb Cortex, 18(1): 53–66CrossRefPubMedGoogle Scholar
  32. Lein E S, Hawrylycz M J, Ao N, Ayres M, Bensinger A, Bernard A, Boe A F, Boguski M S, Brockway K S, Byrnes E J, Chen L, Chen L, Chen TM, Chin M C, Chong J, Crook B E, Czaplinska A, Dang C N, Datta S, Dee N R, Desaki A L, Desta T, Diep E, Dolbeare T A, Donelan M J, Dong H W, Dougherty J G, Duncan B J, Ebbert A J, Eichele G, Estin L K, Faber C, Facer B A, Fields R, Fischer S R, Fliss T P, Frensley C, Gates S N, Glattfelder K J, Halverson K R, Hart M R, Hohmann J G, Howell M P, Jeung D P, Johnson R A, Karr P T, Kawal R, Kidney J M, Knapik R H, Kuan C L, Lake J H, Laramee A R, Larsen K D, Lau C, Lemon T A, Liang A J, Liu Y, Luong L T, Michaels J, Morgan J J, Morgan R J, Mortrud M T, Mosqueda N F, Ng L L, Ng R, Orta G J, Overly C C, Pak T H, Parry S E, Pathak S D, Pearson O C, Puchalski R B, Riley Z L, Rockett H R, Rowland S A, Royall J J, Ruiz M J, Sarno N R, Schaffnit K, Shapovalova N V, Sivisay T, Slaughterbeck C R, Smith S C, Smith K A, Smith B I, Sodt A J, Stewart N N, Stumpf K R, Sunkin S M, Sutram M, Tam A, Teemer C D, Thaller C, Thompson C L, Varnam L R, Visel A, Whitlock R M, Wohnoutka P E, Wolkey C K, Wong V Y, Wood M, Yaylaoglu M B, Young R C, Youngstrom B L, Yuan X F, Zhang B, Zwingman T A, Jones A R (2007). Genome-wide atlas of gene expression in the adult mouse brain. Nature, 445(7124): 168–176CrossRefPubMedGoogle Scholar
  33. Liang WS, Dunckley T, Beach T G, Grover A, Mastroeni D, Ramsey K, Caselli R J, Kukull W A, McKeel D, Morris J C, Hulette C M, Schmechel D, Reiman E M, Rogers J, Stephan D A (2008). Altered neuronal gene expression in brain regions differentially affected by Alzheimer’s disease: a reference data set. Physiol Genomics, 33(2): 240–256CrossRefPubMedGoogle Scholar
  34. Lobo M K, Karsten S L, Gray M, Geschwind D H, Yang X W (2006). FACS-array profiling of striatal projection neuron subtypes in juvenile and adult mouse brains. Nat Neurosci, 9(3): 443–452CrossRefPubMedGoogle Scholar
  35. Lombardino A J, Hertel M, Li X C, Haripal B, Martin-Harris L, Pariser E, Nottebohm F (2006). Expression profiling of intermingled longrange projection neurons harvested by laser capture microdissection. J Neurosci Methods, 157(2): 195–207CrossRefPubMedGoogle Scholar
  36. Low L K, Cheng H J (2006). Axon pruning: an essential step underlying the developmental plasticity of neuronal connections. Philos Trans R Soc Lond B Biol Sci, 361(1473): 1531–1544CrossRefPubMedGoogle Scholar
  37. Luo L, Salunga R C, Guo H, Bittner A, Joy K C, Galindo J E, Xiao H, Rogers K E, Wan J S, Jackson M R, Erlander M G (1999). Gene expression profiles of laser-captured adjacent neuronal subtypes. Nat Med, 5(1): 117–122CrossRefPubMedGoogle Scholar
  38. Magdaleno S, Jensen P, Brumwell C L, Seal A, Lehman K, Asbury A, Cheung T, Cornelius T, Batten DM, Eden C, Norland SM, Rice D S, Dosooye N, Shakya S, Mehta P, Curran T (2006). BGEM: an in situ hybridization database of gene expression in the embryonic and adult mouse nervous system. PLoS Biol, 4(4): e86CrossRefPubMedGoogle Scholar
  39. Marguerat S, Bähler J (2010). RNA-seq: from technology to biology. Cell Mol Life Sci, 67(4): 569–579CrossRefPubMedGoogle Scholar
  40. Mirnics K, Korade Z, Arion D, Lazarov O, Unger T, Macioce M, Sabatini M, Terrano D, Douglass K C, Schor N F, Sisodia S S (2005). Presenilin-1-dependent transcriptome changes. J Neurosci, 25(6): 1571–1578CrossRefPubMedGoogle Scholar
  41. Mirnics K, Middleton F A, Marquez A, Lewis D A, Levitt P (2000). Molecular characterization of schizophrenia viewed by microarray analysis of gene expression in prefrontal cortex. Neuron, 28(1): 53–67CrossRefPubMedGoogle Scholar
  42. Molyneaux B J, Arlotta P, Hirata T, Hibi M, Macklis J D (2005). Fezl is required for the birth and specification of corticospinal motor neurons. Neuron, 47(6): 817–831CrossRefPubMedGoogle Scholar
  43. Molyneaux B J, Arlotta P, Menezes J R L, Macklis J D (2007). Neuronal subtype specification in the cerebral cortex. Nat Rev Neurosci, 8(6): 427–437CrossRefPubMedGoogle Scholar
  44. Mufson E J, Counts S E, Ginsberg S D (2002). Gene expression profiles of cholinergic nucleus basalis neurons in Alzheimer’s disease. Neurochem Res, 27(10): 1035–1048CrossRefPubMedGoogle Scholar
  45. O’Leary D D, Chou S J, Sahara S (2007). Area patterning of the mammalian cortex. Neuron, 56(2): 252–269CrossRefPubMedGoogle Scholar
  46. O’Leary D D, Koester S E (1993). Development of projection neuron types, axon pathways, and patterned connections of the mammalian cortex. Neuron, 10(6): 991–1006CrossRefPubMedGoogle Scholar
  47. Pietersen C Y, Lim M P, Woo T U (2009). Obtaining high quality RNA from single cell populations in human postmortem brain tissue. J Vis Exp, 30(30): 1444PubMedGoogle Scholar
  48. Polleux F, Ince-Dunn G, Ghosh A (2007). Transcriptional regulation of vertebrate axon guidance and synapse formation. Nat Rev Neurosci, 8(5): 331–340CrossRefPubMedGoogle Scholar
  49. Rossner M J, Hirrlinger J, Wichert S P, Boehm C, Newrzella D, Hiemisch H, Eisenhardt G, Stuenkel C, von Ahsen O, Nave K A (2006). Global transcriptome analysis of genetically identified neurons in the adult cortex. J Neurosci, 26(39): 9956–9966CrossRefPubMedGoogle Scholar
  50. Roy N S, Benraiss A, Wang S, Fraser R A, Goodman R, Couldwell WT, Nedergaard M, Kawaguchi A, Okano H, Goldman S A (2000). Promoter-targeted selection and isolation of neural progenitor cells from the adult human ventricular zone. J Neurosci Res, 59(3): 321–331CrossRefPubMedGoogle Scholar
  51. Rudnicki M, Eder S, Schratzberger G, Mayer B, Meyer T W, Tonko M, Mayer G (2004). Reliability of t7-based mRNA linear amplification validated by gene expression analysis of human kidney cells using cDNA microarrays. Nephron, Exp Nephrol, 97(3): e86–e95CrossRefGoogle Scholar
  52. Sandberg R, Yasuda R, Pankratz D G, Carter T A, Del Rio J A, Wodicka L, Mayford M, Lockhart D J, Barlow C (2000). Regional and strainspecific gene expression mapping in the adult mouse brain. Proc Natl Acad Sci USA, 97(20): 11038–11043CrossRefPubMedGoogle Scholar
  53. Schulze A, Downward J (2000). Analysis of gene expression by microarrays: cell biologist’s gold mine or minefield? J Cell Sci, 113(Pt 23): 4151–4156PubMedGoogle Scholar
  54. Sugino K, Hempel C M, Miller M N, Hattox A M, Shapiro P, Wu C, Huang Z J, Nelson S B (2006). Molecular taxonomy of major neuronal classes in the adult mouse forebrain. Nat Neurosci, 9(1): 99–107CrossRefPubMedGoogle Scholar
  55. Tudor M, Akbarian S, Chen R Z, Jaenisch R (2002). Transcriptional profiling of a mouse model for Rett syndrome reveals subtle transcriptional changes in the brain. Proc Natl Acad Sci USA, 99(24): 15536–15541CrossRefPubMedGoogle Scholar
  56. van Gelder R N, von Zastrow M E, Yool A, Dement W C, Barchas J D, Eberwine J H (1990). Amplified RNA synthesized from limited quantities of heterogeneous cDNA. Proc Natl Acad Sci USA, 87(5): 1663–1667CrossRefPubMedGoogle Scholar
  57. Vercelli A, Repici M, Garbossa D, Grimaldi A (2000). Recent techniques for tracing pathways in the central nervous system of developing and adult mammals. Brain Res Bull, 51(1): 11–28CrossRefPubMedGoogle Scholar
  58. Vernes S C, Newbury D F, Abrahams B S, Winchester L, Nicod J, Groszer M, Alarcón M, Oliver P L, Davies K E, Geschwind D H, Monaco A P, Fisher S E (2008). A functional genetic link between distinct developmental language disorders. N Engl J Med, 359(22): 2337–2345CrossRefPubMedGoogle Scholar
  59. Visel A, Thaller C, Eichele G (2004). an atlas of gene expression patterns in the mouse embryo. Nucleic Acids Res, 32 (90001 Database issue): 552 D–556 DCrossRefGoogle Scholar
  60. Watakabe A, Sugai T, Nakaya N, Wakabayashi K, Takahashi H, Yamamori T, Nawa H (2001). Similarity and variation in gene expression among human cerebral cortical subregions revealed by DNA macroarrays: technical consideration of RNA expression profiling from postmortem samples. Brain Res Mol Brain Res, 88(1–2): 74–82CrossRefPubMedGoogle Scholar
  61. Watson J D, Wang S, Von Stetina S E, Spencer WC, Levy S, Dexheimer P J, Kurn N, Heath J D, Miller D M 3rd (2008). Complementary RNA amplification methods enhance microarray identification of transcripts expressed in the C. elegans nervous system. BMC Genomics, 9(1): 84CrossRefPubMedGoogle Scholar
  62. Wilhelm J, Muyal J P, Best J, Kwapiszewska G, Stein M M, Seeger W, Bohle RM, Fink L (2006). Systematic comparison of the T7-IVT and SMART-based RNA preamplification techniques for DNA microarray experiments. Clin Chem, 52(6): 1161–1167CrossRefPubMedGoogle Scholar
  63. Wurmbach E, González-Maeso J, Yuen T, Ebersole B J, Mastaitis J W, Mobbs C V, Sealfon S C (2002). Validated genomic approach to study differentially expressed genes in complex tissues. Neurochem Res, 27(10): 1027–1033CrossRefPubMedGoogle Scholar
  64. Yamamori T, Rockland K S (2006). Neocortical areas, layers, connections, and gene expression. Neurosci Res, 55(1): 11–27CrossRefPubMedGoogle Scholar
  65. Yao F, Yu F, Gong L, Taube D, Rao D D, MacKenzie R G (2005). Microarray analysis of fluoro-gold labeled rat dopamine neurons harvested by laser capture microdissection. J Neurosci Methods, 143(2): 95–106CrossRefPubMedGoogle Scholar
  66. Yeo G W, Xu X, Liang T Y, Muotri A R, Carson C T, Coufal N G, Gage F H (2007). Alternative splicing events identified in human embryonic stem cells and neural progenitors. PLOS Comput Biol, 3(10): 1951–1967CrossRefPubMedGoogle Scholar
  67. Zapala M A, Hovatta I, Ellison J A, Wodicka L, Del Rio J A, Tennant R, Tynan W, Broide R S, Helton R, Stoveken B S, Winrow C, Lockhart D J, Reilly J F, Young W G, Bloom F E, Lockhart D J, Barlow C (2005). Adult mouse brain gene expression patterns bear an embryologic imprint. Proc Natl Acad Sci USA, 102(29): 10357–10362CrossRefPubMedGoogle Scholar
  68. Zhu H, Yang Y, Gao J, Tao H, Qu C, Qu J, Chen J (2010). Area dependent expression of ZNF312 in human fetal cerebral cortex. Neurosci Res, 68(1): 73–76CrossRefPubMedGoogle Scholar

Copyright information

© Higher Education Press and Springer-Verlag Berlin Heidelberg 2010

Authors and Affiliations

  1. 1.Key Laboratory of Visual Science, National Ministry of Health, and School of Optometry and OphthalmologyWenzhou Medical CollegeZhejiangChina

Personalised recommendations