J. Gen. Physiol.
© The Rockefeller University Press
0022-1295/97/05/619/13 $2.00
Volume 109, Number 5, May 1997 619-631

Ca²⁺-induced Ca²⁺ Release in Chinese Hamster Ovary (CHO) Cells Co-expressing Dihydropyridine and Ryanodine Receptors

Norio Suda,^* Dorothee Franzius,^* Andrea Fleig,^* Seiichiro Nishimura, Matthias Bödding,^* Markus Hoth,^* Hiroshi Takeshima,^§ and Reinhold Penner^*

From the ^* Department of Membrane Biophysics, Max-Planck-Institut für biophysikalische Chemie, Am Fassberg, 37077 Göttingen, Germany;  Department of Medical Chemistry and Molecular Genetics, Kyoto University Faculty of Medicine, Kyoto 606, Japan; and ^§ Department of Pharmacology, Faculty of Medicine, University of Tokyo, Tokoyo 113, Japan

Combined patch-clamp and Fura-2 measurements were performed on chinese hamster ovary (CHO) cells co-expressing two channel proteins involved in skeletal muscle excitation-contraction (E-C) coupling, the ryanodine receptor (RyR)-Ca²⁺ release channel (in the membrane of internal Ca²⁺ stores) and the dihydropyridine receptor (DHPR)-Ca²⁺ channel (in the plasma membrane). To ensure expression of functional L-type Ca²⁺ channels, we expressed ₂, , and  DHPR subunits and a chimeric DHPR ₁ subunit in which the putative cytoplasmic loop between repeats II and III is of skeletal origin and the remainder is cardiac. There was no clear indication of skeletal-type coupling between the DHPR and the RyR; depolarization failed to induce a Ca²⁺ transient (CaT) in the absence of extracellular Ca²⁺ ([Ca²⁺]_o). However, in the presence of [Ca²⁺]_o, depolarization evoked CaTs with a bell-shaped voltage dependence. About 30% of the cells tested exhibited two kinetic components: a fast transient increase in intracellular Ca²⁺ concentration ([Ca²⁺]_i) (the first component; reaching 95% of its peak <0.6 s after depolarization) followed by a second increase in [Ca²⁺]_i which lasted for 5-10 s (the second component). Our results suggest that the first component primarily reflected Ca²⁺ influx through Ca²⁺ channels, whereas the second component resulted from Ca²⁺ release through the RyR expressed in the membrane of internal Ca²⁺ stores. However, the onset and the rate of Ca²⁺ release appeared to be much slower than in native cardiac myocytes, despite a similar activation rate of Ca²⁺ current. These results suggest that the skeletal muscle RyR isoform supports Ca²⁺-induced Ca²⁺ release but that the distance between the DHPRs and the RyRs is, on average, much larger in the cotransfected CHO cells than in cardiac myocytes. We conclude that morphological properties of T-tubules and/or proteins other than the DHPR and the RyR are required for functional "close coupling" like that observed in skeletal or cardiac muscle. Nevertheless, some of our results imply that these two channels are potentially able to directly interact with each other.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				R. Y. NAGARAJ, C. M. NOSEK, M. A. P. BROTTO, M. NISHI, H. TAKESHIMA, T. M. NOSEK, and J. MA Increased susceptibility to fatigue of slow- and fast-twitch muscles from mice lacking the MG29 gene Physiol Genomics, November 9, 2000; 4(1): 43 - 49. [Abstract] [Full Text] [PDF]

				M. Nishi, S. Komazaki, N. Kurebayashi, Y. Ogawa, T. Noda, M. Iino, and H. Takeshima Abnormal Features in Skeletal Muscle from Mice Lacking Mitsugumin29 J. Cell Biol., December 27, 1999; 147(7): 1473 - 1480. [Abstract] [Full Text] [PDF]

				S. Hoare, J. A. Copland, Z. Strakova, K. Ives, Y.-J. Jeng, M. R. Hellmich, and M. S. Soloff The Proximal Portion of the COOH Terminus of the Oxytocin Receptor Is Required for Coupling to Gq, but Not Gi. INDEPENDENT MECHANISMS FOR ELEVATING INTRACELLULAR CALCIUM CONCENTRATIONS FROM INTRACELLULAR STORES J. Biol. Chem., October 1, 1999; 274(40): 28682 - 28689. [Abstract] [Full Text] [PDF]

Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents