Ryanodine Action at Calcium Release Channels. 2. Relation to Substituents of the Cyclohexane Ring

Ryanodine (1) and dehydroryanodine (2) are equipotent probes for the ryanodine receptor (ryr) of calcium release channels and differ only in 9_eq-methyl for 1 and 9,21-methylene for 2. Ryanoids 1 and 2 are used here to prepare novel modifications of the cyclohexane substituents to determine their effects on ryr activity and selectivity. 10-Oxo-1 when reacted with carbonyl and other reagents gave 13 C-10 derivatives including the epi-amine and epi-4-azidobenzoyl hydrazide as a candidate affinity probe. Four derivatives of 2 included the Δ⁸-10-hydroxy and Δ⁸-10-oxo compounds. Defunctionalization of the cyclohexane ring of 2 or its 4,6-ethylboronate was achieved in part by controlled periodate oxidation of the 9,21-diol to the 21-nor-9-oxo compounds. These in turn provided access to the 9_ax- and 9_eq-hydroxy derivatives and to the 21-nor-10-deoxy-9-oxo compound which was converted to 21-nor-10-deoxy-1 and 10-deoxy-2 along with the epimeric 10-deoxy-9-hydroxy compounds. Ryanoids of similar potency to 1 as inhibitors of [³H]-1 binding in mouse brain, rabbit skeletal muscle, and canine ventricle ryr preparations and in a rat cardiac contractility assay (inhibition of mechanical response to electrical stimulation) are epi-1 and the 10-epi-amino, 10-epi-methoxyamino, and 10-epi-azidobenzoyl hydrazide derivatives and 10-deoxydehydroryanodine. With a few exceptions the potency of the ryanoids at the cardiac ryr correlates well with their inhibition of cardiac contractility, indicating that the activity is associated with stabilizing the calcium release channel in a subconducting state, thereby uncoupling the excitation−contraction process.