Benzodiazepines and Benzothiazepines as Modulators of the Sarcoplasmic Reticulum Calcium ATP-ase and Ryanodine Receptors in Striated Muscle

Abstract

We have reported that CGP-37157, a benzothiazepine (BZT) derivative of clonazepam utilized as a blocker of the mitochondrial Na+/Ca2+ exchanger, also activates ryanodine receptors (RyRs) and inhibits the sarcoplasmic reticulum (SR) Ca2+-stimulated ATPase (SERCA). We extended the studies to other BZT (e.g., clonazepan, diltiazem) as well as to benzodiazepines (BZD; e.g. diazepan, lorazepan). We aimed to determine if these drug classes have as a common trait the ability to modulate RyRs and/or SERCA. The effects of BZD and BZT on RyRs activity were tested in SR microsomes with a Ca2+ leak assay as well as after reconstituting RyRs into lipid bilayers. The agents tested had variable potency to increase RyR-mediated Ca2+ leak from skeletal SR microsomes. As an example, while diazepam significantly increased RyR activity, most of the others (clonazepam, lorazepam) had minor effects at high doses. In contrast, diltiazem produced moderate inhibition. Planar bilayer studies confirmed the leak observations with both cardiac and skeletal RyR. In the presence of ruthenium red, most agents decreased the rate of loading which indicates inhibitory effects on SERCA activity. The effects of BZT and BZD on loading correlated with a decrease in ATPase activity of SERCA-enriched skeletal SR fractions. In summary, most BZT and BZD utilized in therapeutics or as pharmacological tools have an inhibitory action on SERCA. In contrast, they show a variety of effects on RyRs ranging from inhibition to activation. Hence, the pharmacological action of BZT and BZD on cellular Ca2+ homeostasis reported in the literature of cardiac and skeletal muscle as well as other non-muscle systems may require taking into consideration the contributions of all drug-sensitive intracellular Ca2+ transporters. (Supported by NIH-GM078665 and AHA-MWA 12180038).