L-type calcium channel structure and function: Mechanisms of gating and signaling in skeletal muscle

A defining property of L-type channels is their potentiation by both dihydropyridine agonists and strong depolarization. In contrast, non L-type channels are potentiated by neither agonist nor depolarization, suggesting that these two processes may be linked. We have tested whether the mechanisms of agonist- and depolarization-induced potentiation in the cardiac L-type channel (α_1C) are linked. We found that the mutant L-type channel GFP-α_1C(TQ → YM), bearing the mutations T1066Y and Q1010M, was potentiated by strong depolarization but not agonist. Conversely, the chimeric channel GFP-CACC was potentiated by agonist but not strong depolarization. These data indicate that the mechanisms of agonist- and depolarization-induced potentiation of α_1C are distinct. Because neither GFP-CACC nor GFP-CCAA was potentiated by depolarization, no single repeat of the α _1C can be responsible for this process. GFP-CACC displayed a low open probability similar to that of α_1C, but could not support depolarization-induced potentiation, demonstrating that a relatively low open probability alone is not sufficient for depolarization-induced potentiation to occur. Thus, depolarization-induced potentiation may be a global channel property.; The II–III loop of the skeletal muscle L-type channel (α_1S) is responsible for bidirectional signaling interactions with the skeletal ryanodine receptor (RyR1): transmitting the orthograde, excitation-contraction (EC) coupling signal to RyR1 and receiving a retrograde, Ca2+ current-enhancing signal from RyR1. Previous reports argued for the importance of two distinct regions of the skeletal II-III loop (residues R681–L690 and residues L720–Q765, respectively), claiming for each a key function in DHPR-RyR1 communication. To address whether residues 720–765 of the α_1S II-III loop are sufficient to enable bi-directional signaling, we constructed a chimera (SkLM) having skeletal (Sk) α_1S sequence except for a II-III loop (L) from the highly divergent α_1M subunit of the house fly, Musca domestica (M). Whole-cell patch clamp analysis of SkLM expressed in dysgenic myotubes showed that this construct was unable to mediate bi-directional signaling despite normal surface expression levels and correct junctional targeting. Introducing α_1S residues L720–L764 into the Musca II-III loop of SkLM completely restored bi-directional signaling, indicating that this 45 residue “critical domain” is likely to be the only sequence of the α_1S II-III loop required for bi-directional coupling.