In contrast to cardiac excitation-contraction coupling (ECC), the functional interaction between Dihydropyridine receptors (DHPRs) and the type 1 Ryanodine receptor (RyR1), in skeletal muscle (SkM) appears to be physical. Accordingly, SkM displays a characteristic ultrastructural association between four DHPRs (=tetrad) and one RyR1. What is the modus operandi of the unique tetradic structure?

Several cytoplasmic domains of the main DHPR subunit, a_1S, have been implicated in functional interactions with RyR1, among them the a_1S II-III loop and the I-II loop/b_1a complex. To elucidate the importance of the latter domain for ECC we: - mutated defined I-II loop positions of a_1S and expressed it in a_1S-null myotubes- designed a_1S-a_1S tandem-channel expression plasmids and selectively mutated one or both a_1S Fluorescence microscopy, electrical activation and SR Ca²⁺ release measurements reveal that:

- single/double residue exchanges at I-II loop positions presumably critical for the b_1a interaction do not interfere with a_1S trafficking and ECC

- triple residue exchanges/extended deletions within the I-II loop abolish functional a_1S expression

- the latter mutations are rescued by expression of an a_1S-a_1S tandem-channel as wt-mutant combination in a_1S-null myotubes

Our results suggest that:

- single/double residue exchanges within the a_1S-I-II loop, who virtually abolish the interaction with b in vitro, do not interfere with functional tetrad formation within intact myotubes

- since detrimental alterations in the I-II loop can be overcome by a wt a_1S in the tandem context, it could be that not all a_1S within a tetrad are required to be functional for operating the RyR1.