A novel calcium current in dysgenic skeletal muscle.

Adams, Brett; Beam, Kurt G.

doi:10.1085/jgp.94.3.429

Cited by 73 publications

(59 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Analyses of skeletal muscle myotubes from the ␤ 1 -null mice dem- onstrates that the L-type Ca 2ϩ current is reduced 10-20-fold. Given our data show that the ␣ 1S subunit is absent or extremely low, the residual L-type current in the ␤ 1 -null cells could be the same as I dys , present in dysgenic cells (24). However, confirmation that this is true will require careful analyses of the residual currents in both dysgenic and ␤ 1 -null myotubes.…”

Section: Discussionmentioning

confidence: 63%

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α ₁ subunit and eliminates excitation-contraction coupling

Gregg

Messing

Strube

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

224

208

View full text Add to dashboard Cite

The multisubunit (␣ 1S , ␣ 2 ͞␦, ␤ 1 , and ␥) skeletal muscle dihydropyridine receptor transduces transverse tubule membrane depolarization into release of Ca 2؉ from the sarcoplasmic reticulum, and also acts as an L-type Ca 2؉ channel. The ␣ 1S subunit contains the voltage sensor and channel pore, the kinetics of which are modified by the other subunits. To determine the role of the ␤ 1 subunit in channel activity and excitation-contraction coupling we have used gene targeting to inactivate the ␤ 1 gene. ␤ 1 -null mice die at birth from asphyxia. Electrical stimulation of ␤ 1 -null muscle fails to induce twitches, however, contractures are induced by caffeine. In isolated ␤ 1 -null myotubes, action potentials are normal, but fail to elicit a Ca 2؉ transient. L-type Ca 2؉ current is decreased 10-to 20-fold in the ␤ 1 -null cells compared with littermate controls. Immunohistochemistry of cultured myotubes shows that not only is the ␤ 1 subunit absent, but the amount of ␣ 1S in the membrane also is undetectable. In contrast, the ␤ 1 subunit is localized appropriately in dysgenic, mdg͞mdg, (␣ 1S -null) cells. Therefore, the ␤ 1 subunit may not only play an important role in the transport͞insertion of the ␣ 1S subunit into the membrane, but may be vital for the targeting of the muscle dihydropyridine receptor complex to the transverse tubule͞sarcoplasmic reticulum junction.

show abstract

Section: Discussionmentioning

confidence: 63%

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α ₁ subunit and eliminates excitation-contraction coupling

Gregg

Messing

Strube

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

224

208

View full text Add to dashboard Cite

show abstract

“…The average amplitude of the maximum calcium transient measured at the end of the test pulse, was 0.12 -+ 0.02 AF/F for the same myotubes. Under the same conditions in normal myotubes, the maximum In addition to T type calcium current, some dysgenic myotubes express a low density of lays, an L type calcium current that differs from Isio~ in having faster activation kinetics and a higher sensitivity to dihydropyridines (Adams and Beam, 1989). Fig.…”

Section: Calcium Transients Related To Calcium Currents In Dysgenic Mmentioning

confidence: 92%

Calcium transients associated with the T type calcium current in myotubes.

Garcı́a

Beam

1994

The Journal of General Physiology

Self Cite

View full text Add to dashboard Cite

Immature skeletal muscle cells, both in vivo and in vitro, express a high density of T type calcium current and a relatively low density of the dihydropyridine receptor, the protein thought to function as the /slow calcium channel and as the voltage sensor for excitation-contraction coupling. Although the role of the voltage sensor in eliciting elevations of myoplasmic, free calcium (calcium transients) has been examined, the role of the T type current has not. In this study we examined calcium transients associated with the T type current in cultured myotubes from normal and dysgenic mice, using the whole cell configuration of the patch clamp technique in conjunction with the calcium indicator dye Fluo-3. In both normal and dysgenic myotubes, the T type current was activated by weak depolarizations and was maximal for test pulses to ~ -20 mV. In normal myotubes that displayed T type calcium current, the calcium transient followed the amplitude and the integral of the current at low membrane potentials (-40 to -20 mV) but not at high potentials, where the calcium transient is caused by SR calcium release. The amplitude of the calcium transient for a pulse to -20 mV measured at 15 ms after depolarization represented, on average, 4.26 ---0.68% (n = 19) of the maximum amplitude of the calcium transient elicited by strong, 15-ms test depolarizations. In dysgenic myotubes, the calcium transient followed the integral of the calcium current at all test potentials, in cells expressing only T type current as well as in cells possessing both T type current and the L type current Idys. Moreover, the calcium transient also followed the amplitude and time course of current in dysgenic myotubes expressing the cardiac, DHP-sensitive calcium channel. Thus, in those cases where the transient appears to be a consequence of calcium entry, it has the same time course as the integral of the calcium current. Inactivation of the T type calcium current with 1-s prepulses, or block of the current by the addition of amiloride (0.3-1.0 mM) caused a reduction in the calcium transient which was similar in normal and dysgenic myotubes. To allow calculation of expected changes of intracellular calcium in response to influx, myotubes were converted to a roughly spherical shape (myoballs) by adding 0.5 p,M colchicine to culture dishes of normal cells. Calcium currents and calcium transients recorded from myoballs were similar to those in normal myotubes. The calculated change in calcium due to influx through the T type channel disagreed with the measured calcium concentration, suggesting that additional calcium might be released from the SR or that calcium buffers in the pipette had not equilibrated with the interior of the cell.

show abstract

“…Primary cultures of myotubes were prepared from newborn dysgenic mice as described previously (15). Approximately 1 week after plating, plasmids carrying cDNA for wild-type or mutant DHPRs (0.1-0.2 g/l) were microinjected into single nuclei.…”

Section: Methodsmentioning

confidence: 99%

Excitation-Contraction Coupling Is Not Affected by Scrambled Sequence in Residues 681–690 of the Dihydropyridine Receptor II-III Loop

Proenza¹,

Wilkens²,

Beam³

2000

Journal of Biological Chemistry

View full text Add to dashboard Cite

A peptide corresponding to residues 681-690 of the II-III loop of the skeletal muscle dihydropyridine receptor ␣ 1 subunit (DHPR, ␣ 1S ) has been reported to activate the skeletal muscle ryanodine receptor (RyR1) in vitro. Within this region of ␣ 1S , a cluster of basic residues, Arg 681 -Lys 685 , was previously reported to be indispensable for the activation of RyR1 in microsomal preparations and lipid bilayers. We have used an intact ␣ 1S subunit with scrambled sequence in this region of the II-III loop (␣ 1S -scr) to test the importance of residues 681-690 and the basic motif for skeletal-type excitationcontraction (EC) coupling and retrograde signaling in vivo. When expressed in dysgenic myotubes (which lack endogenous ␣ 1S ), ␣ 1S -scr restored calcium currents that were indistinguishable, in current density and voltage dependence, from those restored by wild-type ␣ 1S . The scrambled DHPR also rescued skeletal-type EC coupling, as indicated by electrically evoked contractions in the presence of 0.5 mM Cd 2؉ and 0.1 mM La 3؉ . Furthermore, the release of intracellular Ca 2؉ , as assayed by the indicator dye, Fluo-3, had similar kinetics and voltage dependence for ␣ 1S and ␣ 1S -scr. These data suggest that residues 681-690 of the ␣ 1S II-III loop are not essential in muscle cells for normal functioning of the DHPR, including skeletal-type EC coupling and retrograde signaling. Excitation-contraction (EC)1 coupling in skeletal and cardiac muscle involves a functional interaction between dihydropyridine receptors (DHPRs), voltage-gated L-type calcium channels in the sarcolemma, and ryanodine receptors (RyRs), calcium release channels in the sarcoplasmic reticulum membrane. The mechanism of EC coupling differs in skeletal and cardiac muscle. In cardiac muscle, calcium influx through the pore-forming subunit of the cardiac DHPR (␣ 1C ) activates RyRs (1). However, in skeletal muscle, EC coupling is independent of the entry of extracellular Ca 2ϩ (2) and may result instead from a mechanical coupling between the skeletal DHPR ␣ 1 subunit (␣ 1S ) and the skeletal muscle RyR isoform (RyR1). Expression of ␣ 1S /␣ 1C chimeras in dysgenic myotubes (which lack endogenous ␣ 1 subunits) has established that skeletal-type EC coupling depends upon skeletal sequence within the putative cytoplasmic region between repeats II and III (II-III loop, amino acids 666 -791 (3)). Chimeric DHPRs in which smaller segments of the skeletal DHPR were substituted into the cardiac DHPR subsequently identified residues 720 -765 within the II-III loop as critical for activation of skeletal-type EC coupling (4, 5). Moreover, this same critical region is essential for "retrograde signaling," whereby RyR1 enhances the current density of ␣ 1S (6). On the other hand, observations in vitro indicate that a different region of the II-III loop, residues 671-690 ("peptide A"), is important for activation of RyR1, as indicated by ryanodine binding, single channel activity, and calcium release (7-9). Within peptide A, residues 681-690 have been identifi...

show abstract

A novel calcium current in dysgenic skeletal muscle.

Cited by 73 publications

References 25 publications

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α ₁ subunit and eliminates excitation-contraction coupling

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α ₁ subunit and eliminates excitation-contraction coupling

Calcium transients associated with the T type calcium current in myotubes.

Excitation-Contraction Coupling Is Not Affected by Scrambled Sequence in Residues 681–690 of the Dihydropyridine Receptor II-III Loop

Contact Info

Product

Resources

About

A novel calcium current in dysgenic skeletal muscle.

Cited by 73 publications

References 25 publications

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α 1 subunit and eliminates excitation-contraction coupling

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α 1 subunit and eliminates excitation-contraction coupling

Calcium transients associated with the T type calcium current in myotubes.

Excitation-Contraction Coupling Is Not Affected by Scrambled Sequence in Residues 681–690 of the Dihydropyridine Receptor II-III Loop

Contact Info

Product

Resources

About

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α ₁ subunit and eliminates excitation-contraction coupling

Absence of the β subunit ( cchb1 ) of the skeletal muscle dihydropyridine receptor alters expression of the α ₁ subunit and eliminates excitation-contraction coupling