Approximately 60-70% of the total fiber calcium was localized in the terminal cisternae (TC) in resting frog muscle as determined by electron-probe analysis of ultrathin cryosections . During a 1 .2 s tetanus, 59% (69 mmol/kg dry TC) of the calcium content of the TC was released, enough to raise total cytoplasmic calcium concentration by -1 mM . This is equivalent to the concentration of binding sites on the calcium-binding proteins (troponin and parvalbumin) in frog muscle . Calcium release was associated with a significant uptake of magnesium and potassium into the TC, but the amount of calcium released exceeded the total measured cation accumulation by 62 mEq/kg dry weight . It is suggested that most of the charge deficit is apparent, and charge compensation is achieved by movement of protons into the sarcoplasmic reticulum (SR) and/or by the movement of organic co-or counterions not measured by energy dispersive electron-probe analysis . There was no significant change in the sodium or chlorine content of the TIC during tetanus. The unchanged distribution of a permeant anion, chloride, argues against the existence of a large and sustained transSR potential during tetanus, if the chloride permeability of the in situ SR is as high as suggested by measurements on fractionated SR .The calcium content of the longitudinal SR (LSR) during tetanus did not show the LSR to be a major site of calcium storage and delayed return to the TC . The potassium concentration in the LSR was not significantly different from the adjacent cytoplasmic concentration. Analysis of small areas of I-band and large areas, including several sarcomeres, suggested that chloride is anisotropically distributed, with some of it probably bound to myosin . In contrast, the distribution of potassium in the fiber cytoplasm followed the water distribution . The mitochondrial concentration of calcium was low and did not change significantly during a tetanus. The TIC of both tetanized and resting freeze-substituted muscles contained electron-lucent circular areas . The appearance of the TIC showed no evidence of major volume changes during tetanus, in agreement with the estimates of unchanged (-72%) water content of the TIC obtained with electron-probe analysis .The release of Ca from and its subsequent return to the triadic portion of the sarcoplasmic reticulum (SR) (28,80,82) are the major determinants of the contractile cycle of striated muscle (for review, see reference 24) . Since the demonstration of the SR as the ATP-dependent relaxing factor (57), a wealth of information has been accumulated about the kinetics and THE JOURNAL OF CELL BIOLOGY " VOLUME 90 SEPTEMBER 1981 577-594 © The Rockefeller University Press -0021-9525/81/09/0577/18 $1 .00 mechanisms of calcium uptake by the SR (e.g., 41,48,63,104,106, and for review, see references 62, 102). In contrast, comparatively little is known about the mechanism of release and associated ion movements, largely because isolated SR preparations do not lend themselves to reproduction of the ph...

In contrast to skeletal muscle isoforms of myosin binding protein C (MyBP-C), the cardiac isoform has 11 rather than 10 fibronectin or Ig modules (modules are identified as C0 to C10, NH2 to COOH terminus), 3 phosphorylation sites between modules C1 and C2, and 28 additional amino acids rich in proline in C5. Phosphorylation between C1 and C2 increases maximum Ca-activated force (Fmax), alters thick filament structure, and increases the probability of myosin heads on the thick filament binding to actin on the thin filament. Unphosphorylated C1C2 fragment binds to myosin, but phosphorylation inhibits the binding. MyBP-C also binds to actin. Using two types of immunoprecipitation and cosedimentation, we show that fragments of MyBP-C containing C0 bind to actin. In low concentrations C0-containing fragments bind to skinned fibers when the NH2 terminus of endogenous MyBP-C is bound to myosin, but not when MyBP-C is bound to actin. C1C2 fragments bind to skinned fibers when endogenous MyBP-C is bound to actin but not to myosin. Disruption of interactions of endogenous C0 with a high concentration of added C0C2 fragments produces the same effect on contractility as extraction of MyBP-C, namely decrease in Fmax and increase in Ca sensitivity. These results suggest that cardiac contractility can be regulated by shifting the binding of the NH2 terminus of MyBP-C between actin and myosin. This mechanism may have an effect on diastolic filling of the heart.

Ca ions can influence the contraction of cardiac muscle by activating kinases that specifically phosphorylate the myofibrillar proteins myosin-binding protein C (MyBP-C) and the regulatory light chain of myosin (RLC). To investigate the possible role of Ca-regulated phosphorylation of MyBP-C on contraction, isolated quiescent and rhythmically contracting cardiac trabeculae were exposed to different concentrations of extracellular Ca and then chemically skinned to clamp the contractile system. Maximum Ca-activated force (F(max)) was measured in quiescent cells soaking in 1) 2.5 mM Ca for 120 min, 2) 1.25 mM for 120 min, or 3) 1.25 mM for 120 min followed by 10 min in 7.5 mM, and 4) cells rhythmically contracting in 2.5 mM for 20 min. F(max) was, respectively, 21.5, 10.5, 24.7, and 32.6 mN/mm(2). Changes in F(max) were closely associated with changes in the degree of phosphorylation of MyBP-C and occurred at intracellular concentrations of Ca below levels associated with phosphorylation of RLC. Monophosphorylation of MyBP-C by a Ca-regulated kinase is necessary before beta-adrenergic stimulation can produce additional phosphorylation. These results suggest that Ca-dependent phosphorylation of MyBP-C modulates contractility by changing thick filament structure.

A B S T R A C T Treatment of rat ventricular cells with 10 mM EGTA makes the sarcolemma highly permeable to small ions and molecules without removing its restriction of the diffusion of larger molecules or inactivating all of its enzymatic functions. These hyperpermeable cardiac cells have been used to study the regulation of the range of concentration of Ca over which activation of the contractile proteins occurs (Ca sensitivity). The Ca sensitivity can be varied from three-to sixfold without any significant alteration in the general shape of the relation between force and Ca concentrations. Although cyclic nucleotides in concentrations of 10 -9 to 10 -5 M do not influence Ca sensitivity, in the presence of a phosphodiesterase inhibitor, cGMP increases and cAMP decreases Ca sensitivity. Treatment of the hyperpermeable cells with a nonionic detergent raises Ca sensitivity as does removal of the phosphate donor by complete substitution of CTP for ATP. These data indicate that Ca sensitivity is probably modulated by a cAMPdependent phosphorylation that decreases Ca sensitivity. The sarcolemma is required for this reaction to take place. The effect of this reaction is antagonized by a cGMP-dependent reaction occurring inside the cell. Studies involving the perfusion of the heart with and without epinephrine before the exposure to EGTA indicate that epinephrine can regulate this system of control of Ca sensitivity. The functional considerations of this regulatory system are discussed.

Inductively coupled plasma (ICP) etch rates for GaN are reported as a function of plasma pressure, plasma chemistry, rf power, and ICP power. Using a Cl2/H2/Ar plasma chemistry, GaN etch rates as high as 6875 Å/min are reported. The GaN surface morphology remains smooth over a wide range of plasma conditions as quantified using atomic force microscopy. Several etch conditions yield highly anisotropic profiles with smooth sidewalls. These results have direct application to the fabrication of group-III nitride etched laser facets.