Abstract
Exogenous expression of genes in mammalian neurons represents a substantial experimental challenge because of the low efficiency of commercially available liposomal transfection reagents for nondividing cells and considerable toxicity of viral transfection systems. In this chapter, we discuss application of the ℌbiolisticℍ; particle delivery system for heterologous expression of genes in primary neuron cultures. The method is based on the direct introduction of cDNA of interest into the nucleus by penetration with DNA-coated gold particles. With this approach, cDNA expression is independent of cell cycling and proliferation and is similar to intranuclear microinjection, with both avoiding cDNA delivery through the cytosol. Examples of successful transfection using PDS of rat superior cervical ganglion and trigeminal ganglion neurons are discussed.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsSpringer Nature is developing a new tool to find and evaluate Protocols. Learn more
References
Hille, B., Beech, D. J., Bernheim, L., Mathie, A., Shapiro, M. S., and Wollmuth, L. P. (1995) Multiple G-protein-coupled pathways inhibit N-type Ca2+ channels of neurons. Life Sci. 56, 989–992.
Gamper, N. and Shapiro, M. S. (2003) Calmodulin mediates Ca2+-dependent modulation of M-type K+ channels. J. Gen. Physiol. 122, 17–31.
Gamper, N., Stockand, J. D., and Shapiro, M. S. (2003) Subunit-specific modulation of KCNQ potassium channels by Src tyrosine kinase. J. Neurosci. 23, 84–95.
Gamper, N., Reznikov, V., Yamada, Y., Yang, J., and Shapiro, M. S. (2004) Phosphatidylinositol 4,5-bisphosphate signals underlie receptor-specific Gq/11-mediated modulation of N-type Ca2+ channels. J. Neurosci. 24, 10,980–10,992.
Patwardhan, A., Berg, K. A., Akopain, A. N., et al. (2005) Bradykinin-induced functional competence and trafficking of the delta opioid receptor in trigeminal nociceptors. J. Neurosci. 25, 8825–8832.
Malin, S. A. and Nerbonne, J. M. (2000) Elimination of the fast transient in superior cervical ganglion neurons with expression of KV4.2W362F: molecular dissection of IA. J. Neurosci. 20, 5191–5199.
Malin, S. A. and Nerbonne, J. M. (2001) Molecular heterogeneity of the voltagegated fast transient outward K+ current, I(Af), in mammalian neurons. J. Neurosci. 21, 8004–8014.
Bernheim, L., Beech, D. J., and Hille, B. (1991) A diffusible second messenger mediates one of the pathways coupling receptors to calcium channels in rat sympathetic neurons. Neuron 6, 859–867.
Price, T. J., Patwardhan, A., Akopian, A. N., Hargreaves, K. M., and Flores, C. M. (2004) Modulation of trigeminal sensory neuron activity by the dual cannabinoidvanilloid agonists anandamide, N-arachidonoyl-dopamine and arachidonyl-2-chloroethylamide. Br. J. Pharmacol. 141, 1118–1130.
Shaw, G. (1996) The pleckstrin homology domain:an intriguing multifunctional protein module. Bioessays 18, 35–46.
Varnai, P. and Balla, T. (1998) Visualization of phosphoinositides that bind pleckstrin homology domains:calcium-and agonist-induced dynamic changes and relationship to myo-[3H]inositol-labeled phosphoinositide pools. J. Cell. Biol. 143, 501–510.
Hirose, K., Kadowaki, S., Tanabe, M., Takeshima, H., and Iino, M. (1999) Spatiotemporal dynamics of inositol 1,4,5-trisphosphate that underlies complex Ca2+ mobilization patterns. Science 284, 1527–1530.
Nash, M. S., Young, K. W., Willars, G. B., Challiss, R. A., and Nahorski, S. R. (2001) Single-cell imaging of graded Ins(1,4,5)P3 production following G-proteincoupled-receptor activation. Biochem. J. 356, 137–142.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2006 Humana Press Inc.
About this protocol
Cite this protocol
Gamper, N., Shapiro, M.S. (2006). Exogenous Expression of Proteins in Neurons Using the Biolistic Particle Delivery System. In: Stockand, J.D., Shapiro, M.S. (eds) Ion Channels. Methods in Molecular Biology™, vol 337. Humana Press. https://doi.org/10.1385/1-59745-095-2:27
Download citation
DOI: https://doi.org/10.1385/1-59745-095-2:27
Publisher Name: Humana Press
Print ISBN: 978-1-58829-576-7
Online ISBN: 978-1-59745-095-9
eBook Packages: Springer Protocols