Transfection of mammalian cells with connexins and measurement of voltage sensitivity of their gap junctions

Cristiane del Corsso, Miduturu Srinivas, Marcia Urban-Maldonado, Alonso P. Moreno, Alfredo G. Fort, Glenn I. Fishman, David C. Spray

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

Vertebrate gap junction channels are formed by a family of more than 20 connexin proteins. These gap junction proteins are expressed with overlapping cellular and tissue specificity, and coding region mutations can cause human hereditary diseases. Here we present a summary of what has been learned from voltage clamp studies performed on cell pairs either endogenously expressing gap junctions or in which connexins are exogenously expressed. General protocols presented here are currently used to transfect mammalian cells with connexins and to study the biophysical properties of the heterologously expressed connexin channels. Transient transfection is accomplished overnight with maximal expression occurring at about 36 h; stable transfectants normally can be generated within three or four weeks through colony selection. Electrophysiological protocols are presented for analysis of voltage dependence and single-channel conductance of gap junction channels as well as for studies of chemical gating of these channels.

Original languageEnglish (US)
Pages (from-to)1799-1809
Number of pages11
JournalNature Protocols
Volume1
Issue number4
DOIs
StatePublished - Nov 1 2006

    Fingerprint

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

del Corsso, C., Srinivas, M., Urban-Maldonado, M., Moreno, A. P., Fort, A. G., Fishman, G. I., & Spray, D. C. (2006). Transfection of mammalian cells with connexins and measurement of voltage sensitivity of their gap junctions. Nature Protocols, 1(4), 1799-1809. https://doi.org/10.1038/nprot.2006.266