Electroporation of the Hindbrain to Trace Axonal Trajectories and Synaptic Targets in the Chick Embryo

Overview

Journal J Vis Exp

Specialties Biology
General Medicine

Date 2013 Jun 11

PMID 23748440

Citations 6

Authors

Ayelet Kohl

Yoav Hadas

Avihu Klar

Dalit Sela-Donenfeld

Affiliations

Soon will be listed here.

Abstract

Electroporation of the chick embryonic neural tube has many advantages such as being quick and efficient for the expression of foreign genes into neuronal cells. In this manuscript we provide a method that demonstrates uniquely how to electroporate DNA into the avian hindbrain at E2.75 in order to specifically label a subset of neuronal progenitors, and how to follow their axonal projections and synaptic targets at much advanced stages of development, up to E14.5. We have utilized novel genetic tools including specific enhancer elements, Cre/Lox - based plasmids and the PiggyBac-mediated DNA transposition system to drive GFP expression in a subtype of hindbrain cells (the dorsal most subgroup of interneurons, dA1). Axonal trajectories and targets of dA1 axons are followed at early and late embryonic stages at various brainstem regions. This strategy contributes advanced techniques for targeting cells of interest in the embryonic hindbrain and for tracing circuit formation at multiple stages of development.

Citing Articles

Hindbrain boundaries as niches of neural progenitor and stem cells regulated by the extracellular matrix proteoglycan chondroitin sulphate.

Hutchings C, Nuriel Y, Lazar D, Kohl A, Muir E, Genin O Development. 2024; 151(4).

PMID: 38251863 PMC: 10911165. DOI: 10.1242/dev.201934.

Axonal Projection Patterns of the Dorsal Interneuron Populations in the Embryonic Hindbrain.

Hirsch D, Kohl A, Wang Y, Sela-Donenfeld D Front Neuroanat. 2022; 15:793161.

PMID: 35002640 PMC: 8738170. DOI: 10.3389/fnana.2021.793161.

Temporal-specific roles of fragile X mental retardation protein in the development of the hindbrain auditory circuit.

Wang X, Kohl A, Yu X, Zorio D, Klar A, Sela-Donenfeld D Development. 2020; 147(21).

PMID: 32747436 PMC: 7473654. DOI: 10.1242/dev.188797.

Neural stem cells deriving from chick embryonic hindbrain recapitulate hindbrain development in culture.

Peretz Y, Kohl A, Slutsky N, Komlos M, Varshavsky S, Sela-Donenfeld D Sci Rep. 2018; 8(1):13920.

PMID: 30224755 PMC: 6141497. DOI: 10.1038/s41598-018-32203-w.

Control of axon guidance and neurotransmitter phenotype of dB1 hindbrain interneurons by Lim-HD code.

Kohl A, Marquardt T, Klar A, Sela-Donenfeld D J Neurosci. 2015; 35(6):2596-611.

PMID: 25673852 PMC: 6605615. DOI: 10.1523/JNEUROSCI.2699-14.2015.

References

Avraham O, Hadas Y, Vald L, Hong S, Song M, Klar A . Motor and dorsal root ganglion axons serve as choice points for the ipsilateral turning of dI3 axons. J Neurosci. 2010; 30(46):15546-57. PMC: 6633670. DOI: 10.1523/JNEUROSCI.2380-10.2010. View

Lumpkin E, Collisson T, Parab P, Omer-Abdalla A, Haeberle H, Chen P . Math1-driven GFP expression in the developing nervous system of transgenic mice. Gene Expr Patterns. 2003; 3(4):389-95. DOI: 10.1016/s1567-133x(03)00089-9. View

Lumsden A . The cellular basis of segmentation in the developing hindbrain. Trends Neurosci. 1990; 13(8):329-35. DOI: 10.1016/0166-2236(90)90144-y. View

Alsina B, Vu T . Visualizing synapse formation in arborizing optic axons in vivo: dynamics and modulation by BDNF. Nat Neurosci. 2001; 4(11):1093-101. DOI: 10.1038/nn735. View

Avraham O, Hadas Y, Vald L, Zisman S, Schejter A, Visel A . Transcriptional control of axonal guidance and sorting in dorsal interneurons by the Lim-HD proteins Lhx9 and Lhx1. Neural Dev. 2009; 4:21. PMC: 2704203. DOI: 10.1186/1749-8104-4-21. View

Muller T, Anlag K, Wildner H, Britsch S, Treier M, Birchmeier C . The bHLH factor Olig3 coordinates the specification of dorsal neurons in the spinal cord. Genes Dev. 2005; 19(6):733-43. PMC: 1065726. DOI: 10.1101/gad.326105. View

Rose M, Ahmad K, Thaller C, Zoghbi H . Excitatory neurons of the proprioceptive, interoceptive, and arousal hindbrain networks share a developmental requirement for Math1. Proc Natl Acad Sci U S A. 2010; 106(52):22462-7. PMC: 2799716. DOI: 10.1073/pnas.0911579106. View

Lu Y, Lin C, Wang X . PiggyBac transgenic strategies in the developing chicken spinal cord. Nucleic Acids Res. 2009; 37(21):e141. PMC: 2790877. DOI: 10.1093/nar/gkp686. View

Gardzinski P, Lee D, Fei G, Hui K, Huang G, Sun H . The role of synaptotagmin I C2A calcium-binding domain in synaptic vesicle clustering during synapse formation. J Physiol. 2007; 581(Pt 1):75-90. PMC: 2075219. DOI: 10.1113/jphysiol.2006.127472. View

10.

Liu Z, Li H, Hu X, Yu L, Liu H, Han R . Control of precerebellar neuron development by Olig3 bHLH transcription factor. J Neurosci. 2008; 28(40):10124-33. PMC: 2590637. DOI: 10.1523/JNEUROSCI.3769-08.2008. View

11.

Marin F, Puelles L . Morphological fate of rhombomeres in quail/chick chimeras: a segmental analysis of hindbrain nuclei. Eur J Neurosci. 1995; 7(8):1714-38. DOI: 10.1111/j.1460-9568.1995.tb00693.x. View

12.

Nowack A, Yao J, Custer K, Bajjalieh S . SV2 regulates neurotransmitter release via multiple mechanisms. Am J Physiol Cell Physiol. 2010; 299(5):C960-7. PMC: 2980308. DOI: 10.1152/ajpcell.00259.2010. View

13.

Kohl A, Hadas Y, Klar A, Sela-Donenfeld D . Axonal patterns and targets of dA1 interneurons in the chick hindbrain. J Neurosci. 2012; 32(17):5757-71. PMC: 6703607. DOI: 10.1523/JNEUROSCI.4231-11.2012. View

14.

Avraham O, Zisman S, Hadas Y, Vald L, Klar A . Deciphering axonal pathways of genetically defined groups of neurons in the chick neural tube utilizing in ovo electroporation. J Vis Exp. 2010; (39). PMC: 2902355. DOI: 10.3791/1792. View

15.

Vogel J, Mobius C, Kuschinsky W . Early delineation of ischemic tissue in rat brain cryosections by high-contrast staining. Stroke. 1999; 30(5):1134-41. DOI: 10.1161/01.str.30.5.1134. View

16.

Leal-Ortiz S, Waites C, Terry-Lorenzo R, Zamorano P, Gundelfinger E, Garner C . Piccolo modulation of Synapsin1a dynamics regulates synaptic vesicle exocytosis. J Cell Biol. 2008; 181(5):831-46. PMC: 2396795. DOI: 10.1083/jcb.200711167. View

17.

Storm R, Cholewa-Waclaw J, Reuter K, Brohl D, Sieber M, Treier M . The bHLH transcription factor Olig3 marks the dorsal neuroepithelium of the hindbrain and is essential for the development of brainstem nuclei. Development. 2008; 136(2):295-305. DOI: 10.1242/dev.027193. View

18.

Wang J, Miller E, Simmons G, Miller T, Tabashnik B, Park Y . piggyBac-like elements in the pink bollworm, Pectinophora gossypiella. Insect Mol Biol. 2009; 19(2):177-84. DOI: 10.1111/j.1365-2583.2009.00964.x. View

19.

Niwa H, Yamamura K, Miyazaki J . Efficient selection for high-expression transfectants with a novel eukaryotic vector. Gene. 1991; 108(2):193-9. DOI: 10.1016/0378-1119(91)90434-d. View

20.

Clarke J, Lumsden A . Segmental repetition of neuronal phenotype sets in the chick embryo hindbrain. Development. 1993; 118(1):151-62. DOI: 10.1242/dev.118.1.151. View