Journal article
Ca2+ Current and Charge Movements in Skeletal Myotubes Promoted by the β-Subunit of the Dihydropyridine Receptor in the Absence of Ryanodine Receptor Type 1
Biophysical journal, Vol.84(2), pp.942-959
02/2003
DOI: 10.1016/S0006-3495(03)74911-X
PMCID: PMC1302672
PMID: 12547776
Abstract
The
β
-subunit of the dihydropyridine receptor (DHPR) enhances the Ca
2+
channel and voltage-sensing functions of the DHPR. In skeletal myotubes, there is additional modulation of DHPR functions imposed by the presence of ryanodine receptor type-1 (RyR1). Here, we examined the participation of the
β
-subunit in the expression of L-type Ca
2+
current and charge movements in RyR1 knock-out (KO),
β
1 KO, and double
β
1/RyR1 KO myotubes generated by mating heterozygous
β
1 KO and RyR1 KO mice. Primary myotube cultures of each genotype were transfected with various
β
-isoforms and then whole-cell voltage-clamped for measurements of Ca
2+
and gating currents. Overexpression of the endogenous skeletal
β
1a isoform resulted in a low-density Ca
2+
current either in RyR1 KO (36 ± 9 pS/pF) or in
β
1/RyR1 KO (34 ± 7 pS/pF) myotubes. However, the heterologous
β
2a variant with a double cysteine motif in the N-terminus (C3, C4), recovered a Ca
2+
current that was entirely wild-type in density in RyR1 KO (195 ± 16 pS/pF) and was significantly enhanced in double
β
1/RyR1 KO (115 ± 18 pS/pF) myotubes. Other variants tested from the four
β
gene families (
β
1a,
β
1b,
β
1c,
β
3, and
β
4) were unable to enhance Ca
2+
current expression in RyR1 KO myotubes. In contrast, intramembrane charge movements in
β
2a-expressing
β
1a/RyR1 KO myotubes were significantly lower than in
β
1a-expressing
β
1a/RyR1 KO myotubes, and the same tendency was observed in the RyR1 KO myotube. Thus,
β
2a had a preferential ability to recover Ca
2+
current, whereas
β
1a had a preferential ability to rescue charge movements. Elimination of the double cysteine motif (
β
2a C3,4S) eliminated the RyR1-independent Ca
2+
current expression. Furthermore, Ca
2+
current enhancement was observed with a
β
2a variant lacking the double cysteine motif and fused to the surface membrane glycoprotein CD8. Thus, tethering the
β
2a variant to the myotube surface activated the DHPR Ca
2+
current and bypassed the requirement for RyR1. The data suggest that the Ca
2+
current expressed by the native skeletal DHPR complex has an inherently low density due to inhibitory interactions within the DHPR and that the
β
1a-subunit is critically involved in process.
Details
- Title: Subtitle
- Ca2+ Current and Charge Movements in Skeletal Myotubes Promoted by the β-Subunit of the Dihydropyridine Receptor in the Absence of Ryanodine Receptor Type 1
- Creators
- Chris A Ahern - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706David. C Sheridan - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Weijun Cheng - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Lindsay Mortenson - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Priya Nataraj - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Paul Allen - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Michel De Waard - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Roberto Coronado - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706
- Resource Type
- Journal article
- Publication Details
- Biophysical journal, Vol.84(2), pp.942-959
- Publisher
- Biophysical Society
- DOI
- 10.1016/S0006-3495(03)74911-X
- PMID
- 12547776
- PMCID
- PMC1302672
- ISSN
- 0006-3495
- eISSN
- 1542-0086
- Language
- English
- Date published
- 02/2003
- Academic Unit
- Molecular Physiology and Biophysics; Iowa Neuroscience Institute
- Record Identifier
- 9984070482502771
Metrics
21 Record Views