Levels of SR proteins involved in calcium binding and release are unchanged in failing dilated cardiomyopathy. In contrast, protein levels of calcium ATPase involved in SR calcium uptake are reduced in the failing myocardium. Moreover, SR calcium ATPase is decreased relative to its inhibitory protein, phospholamban.(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic electro-stimulation of fast-twitch rabbit muscle with the frequency pattern received by a slow-twitch muscle induces a progressive transformation of the sarcoplasmic reticulum. After 2 days stimulation activities of Ca2'-dependent ATPase and of Ca2' transport begin to decrease, and are paralleled by a progressive decrease in CaZi-dependent and CaZi, Mg2' -dependent phosphoprotein formation, reduced rate of dephosphorylation and a rearrangement of the electrophoretic polypeptide and phosphoprotein patterns. These findings suggest a transformation of the sarcoplasmic reticulum to resemble that of a slow-twitch muscle. This transformation is paralleled by increase in time-to-peak of twitch contraction and half relaxation time and occurs before conversion of the myosin light chain pattern is observed. The parallel time course of changes in contractile properties of stimulated muscle and the molecular and functional properties of the sarcoplasmic reticulum emphasizes the definitive role of the latter in determining the twitch characteristics of fast and slow twitch muscles.Sarcoplasmic reticulum has been mainly characterized (for recent reviews see [1,2]) in fast-twitch fibres, since they predominate the fibre populations of most vertebrate muscles. A small number of reports deals with properties of sarcoplasmic reticulum isolated from muscles composed mainly of slow-twitch fibres [3-151. According to these studies, marked differences exist between the sarcoplasmic reticulum in the two fibre types. The present study was undertaken in order to examine whether transformation of fast-twitch muscle as induced by chronic electrostimulation affects the sarcoplasmic reticulum and to which extent changes of the latter can be correlated to the time course of transformation of contractile properties. Changes induced by electro-stimulation should also substantiate the differences existing between the sarcoplasmic reticulum in fast-twitch and slow-twitch muscles [I 51. It has been shown that many properties of fast-twitch muscles can be changed towards those of slow-twitch muscles by indirect stimulation at a frequency pattern resembling that of a motoneuron innervating a slow-twitch muscle [16- Enzymes. ATPase (EC 3.6.1.3); lactate dehydrogenase (EC 1.1.1.27); pyruvate kinase (EC 2.7.1.40); glycogen phosphorylase (EC 2.4.1.1).Symbol. d, days.321. Early stimulation induced changes occur in contractile properties whereas alterations in the myosin are detectable only after prolonged stimulation [21,28,331. Early changes of the sarcoplasmic reticulum have, however, been observed in SrCter's [22,26,29] and our laboratory [25,31,34] and it appeared interesting to study these changes more in detail in order to gain information on their role in the transformation of the overall contractile properties. MATERIALS AND METHODS Stimulation and Physiological MeasurementsMale adult rabbits of the strain 'Weisse Wiener' were stimulated unilaterally via externalized electrodes implanted in close vicinity to the left lateral popliteal ne...

Subfractionation of sarcoplasmic reticulum from fast‐twitch and slow‐twitch rabbit skeletal muscles was performed on a sucrose density gradient. Vesicle fractions were characterized by: measurement of (Ca2+,Mg2+)‐dependent (extra) ATPase, Mg2+ ‐dependent (basal) ATPase, Ca2+ uptake characteristics, polypeptide patterns in sodium dodecylsulphate polyacrylamide gel electro‐phoreses, phosphoprotein formation and electronmicroscopy of negatively stained samples. In fast‐twitch muscle, low and high density vesicles were separated. The latter showed high activity of (Ca2+, Mg2+)‐dependent ATPase, negligible activity of Mg2+‐dependent ATPase, high initial rate and high capacity of Ca2+ uptake, high amount of phosphorylated 115000‐Mr polypeptide, and appeared morphologically as thin‐walled vesicles covered with particles of 4 nm in diameter. Low density vesicles had little (Ca2+, Mg2+)‐dependent ATPase but high Mg2+‐dependent ATPase. Although the initial rate of Ca2+ uptake was markedly lower, the total capacity of uptake was comparable with that of high density vesicles. Phosphorylated 115000‐Mr polypeptide was detectable at low concentrations. Instead, 57000 and 47000‐Mr polypeptides were characterized as forming stable phosphoproteins in the presence of ATP and Mg2+. Negatively stained, these vesicles appeared to have smooth surfaces. It is suggested that low density vesicles represent a Ca2+ sequestering system different from that of high density vesiclés and that Mg2+‐dependent (basal) ATPase as well as the 57000 and 47000‐Mr polypeptides are part of the Ca2+ transport system within the low density vesicles. According to the results from slow‐twitch muscle, Ca2+ sequestration by the sarcoplasmic reticulum functions in this muscle type only through the low density vesicles.

Calreticulin binds Zn 2+ with the relatively high affinit)/low capacity. To determine the location of the Zn 2+ binding site in calreticulin different domains of the protein were expressed in E. coli, using the glutathione S-transferase fusion protein system, and their Zn2+-dependent interaction with Zn2+-lDA-agarose were determined. Three distinct domains were used in this study: the N + P-domain (the first 290 residues); the N-domain (residues 1-182) and the proline-rich P-domain (residues 180-273). The N + P-domain bound to the Zn2+-lDA-agarose and were eluted with an increasing concentration of imidazole. The N-domain also bound 65Zn2+ as measured by the overlay method. The P-domain did not interact with the ZnZ+-IDA-agarose and it did not bind any detectable amount of Zu 2+. Chemical modification of calreticulin with diethyl pyrocarbonate indicated that rise out of seven histidines were protected in the presence of Zn 2+ but they were modified by diethyl pyrocarbonate in the absence of Zn 2+ suggesting that these residues may be involved in Zn 2+ binding to ealreticulin. We conclude that Zn 2+ binding sites in calreticulin are localized to the N-domain of the protein, region that is not involved in Ca 2+ binding to calreticulin.

This report presents a 46-year-old man who was treated for hypertension with the angiotensin-converting-enzyme (ACE) inhibitor enalapril. After 3 years of continuous treatment he presented with jaundice and progressive liver failure that continued despite withdrawal of the medication. The patient was taking no other medication. All known causes of acute liver failure could be excluded indicating a drug-induced liver damage after long-term treatment with enalapril. Analysis of liver biopsies revealed a pathomorphological pattern comparable to than observed in severe halothane hepatitis. Serological studies including T-cell stimulation with enalapril and a broad spectrum of tests for autoimmunity including autoantibodies against calreticulin, the major Ca2+ and Zn2+ binding protein of the endoplasmic reticulum and suggested to be involved in the pathogenesis of halothane hepatitis were negative. Thus, the mechanism of enalapril-induced liver injury remains obscure. Liver failure progressed and finally led to orthotopic liver transplantation. To our knowledge, this is the longest duration of chronic treatment with an ACE inhibitor before liver failure occurred. In addition, liver failure progressed despite withdrawal of the medication. It is concluded that even after long-term treatment with an ACE inhibitor liver failure may be induced. Therefore, regular monitoring of liver enzymes should be considered.