The mechanism of Ca2' release from sarcoplasmic reticulum, which triggers contraction in skeletal muscle, remains the key unresolved problem in excitation-contraction coupling. Recently, we have described the isolation of purified fractions referable to terminal and longitudinal cisternae of sarcoplasmic reticulum. Junctional terminal cisternae are distinct in that they have a low net energized Ca2+ loading, which can be enhanced 5-fold or more by addition of ruthenium red. The loading rate, normalized for calcium pump protein content, then approaches that of longitudinal cisternae of sarcoplasmic reticulum. We now find that the ruthenium red-enhaniced Ca2+ loading rate can be blocked by the previous addition of ryanodine. The inhibition constant is in the nanomolar range (20-180 nM). Ryanodine and ruthenium red have no effect on the Ca21 loading rate of longitudinal cisternae. Direct binding studies with [3HJryano-dine localized the receptors to the terminal cisternae and not to longitudinal cisternae. Scatchard analysis of the binding data gives a dissociation constant for ryanodine in the range of the drug action on the terminal cisternae ("100 nM range) with approximately 4 to 20 pmol bound per mg of protein.Ryanodine is known to be toxic in animals, leading to irreversible muscle contractures. These studies provide evidence on the mode of action of ryanodine and its localization to the terminal cisternae. The low concentration at which the drug is effective appears to account for its toxicity. Ryanodine locks the Ca2+ release channels in the "open state," so that Ca2+ is not reaccumulated and the muscle fiber cannot relax.

Distribution and metabolic fate of radiolabeled hexadecylphosphocholine (He-PC) has been studied in mice. It is demonstrated that He-PC is well-absorbed from the intestinal tract, intravenous (IV) and oral administration lead to similar distributions throughout the body, the highest accumulation of radioactivity occurs in liver, lung and kidney, and the metabolic products are radioactive choline, phosphocholine and 1,2-diacylphosphatidylcholine. The occurrence of these metabolites indicates that phospholipases C and D may be involved in He-PC breakdown.

Hexadecylphosphocholine (He-PC) is a new compound synthesized according to the minimal structural requirements deducted from studies with other ether lipids. In vitro studies on He-PC revealed remarkable antineoplastic activity on HL60, U937, Raji and K562 leukemia cell lines. In addition, He-PC, applied orally, showed a superior effect in the treatment of dimethylbenzanthracene-induced rat mammary carcinomas when compared to intravenously administered cyclophosphamide. After oral application He-PC was well absorbed from the intestine and metabolized in the liver by phospholipases C and D. During a 5-week treatment no hematotoxic effects were detected. In a clinical pilot study on breast cancer patients with widespread skin involvement, topically applied He-PC showed skin tumor regressions without local or systemic side effects.

Two new aspects of mammalian porin are presented. First, by affinity chromatography we show that channel active human or bovine porin reversibly bind the stilbene-disulfonate group of the chloride channel blocker 4,4'-diisothiocyanatostilbene-2,2'-disulfonate (DIDS). The procedure is suitable for further purification of porin after enrichment by ion exchange chromatography and shows a yield of 24.3%. The data support our recent proposal that VDAC forms part of the ORDIC channel complex which is affected in cystic fibrosis. Second, a purification scheme for mammalian porin is given starting with direct solubilisation of ground bovine skeletal muscle to avoid breaking up tissue. About 130 mg of channel active "Porin 31BM" are enriched from 946 g muscle tissue. Concerning its apparent molecular mass, primary structure, channel activity, channel conductance and voltage dependence the molecule shows high similarity to human porin. "Porin 31BM" is furthermore labelled by antibodies raised against human B lymphocyte derived "Porin 31HL".