MRI of carcinoid tumors: spectrum of appearances in the gastrointestinal tract and liver.

The purpose of this study was to evaluate the spectrum of appearances of gastrointestinal carcinoid tumors at magnetic resonance imaging (MRI) and to elucidate patterns of appearances of carcinoid liver metastases on precontrast and postgadolinium images. The MR examinations of 29 patients (11 men, 18 women; age range, 33-87 years) with histologically confirmed gastrointestinal carcinoid tumors, representing our complete 9.5 years of experience with this entity, were retrospectively reviewed. Twelve patients had MR examinations prior to resection or biopsy of the primary tumor (preoperative group); 17 patients were imaged postsurgically (postoperative group). All MR studies were performed at 1.5 T and comprised T1-weighted spoiled gradient echo (SGE), T2-weighted fat-suppressed turbo spin echo, HASTE, and serial postgadolinium T1-weighted SGE sequences without and with fat suppression. Morphology, signal intensity, and contrast enhancement of primary tumors and of metastases to the mesentery, peritoneum, and liver were evaluated. Primary tumors were visualized in 8 of 12 patients and best demonstrated on postgadolinium T1-weighted fat-suppressed images. The appearance of primary tumors was a nodular mass originating from the bowel wall (4 of 12 patients) or regional uniform bowel wall thickening (4 of 12 patients) with moderate intense enhancement on postgadolinium images. In 4 of 12 patients the primary tumor was prospectively not seen. Mesenteric metastases, seen in eight patients, presented as nodular masses and were associated with mesenteric stranding in seven patients. A total of 156 liver metastases were evaluated in 16 patients. On precontrast T1- and T2-weighted images, 117 metastases (75%) were hypointense and hyperintense, respectively. A total of 146 metastases (94%) were hypervascular, showing moderate intense enhancement during the hepatic arterial phase, and 9 metastases (6%) were hypovascular. Twenty-three metastases (15%) were visible only on immediate postgadolinium images. MRI is able to demonstrate findings in carcinoid tumors, including the primary tumor, mesenteric metastases, and liver metastases. Liver metastases are commonly hypervascular and may be demonstrable only on immediate postgadolinium images.

Action of mercurials on the active and passive transport properties of sarcoplasmic reticulum.

The effect of Hg2+ and CH3-Hg+ on the passive and active transport properties of the Ca2+-Mg2+-ATPase-rich fraction of skeletal sarcoplasmic reticulum (SR) is reported. The agents abolish active transport, at 10(-5) and 10(-4) M concentrations, respectively. Addition of the mercurials was also shown to release actively accumulated Ca2+. The mercurials increase the passive Ca2+ and Mg2+ permeability in the absence of ATP at the same concentrations at which they inhibit transport. It is proposed that both effects are the result of direct binding of the mercurials to the SH groups of the Ca2+-Mg2+-ATPase pump, altering the conformational equilibria of the pump. The agents were also shown to increase the passive KCl permeability. The SR preparation consists of two vesicle populations with respect to K+ permeability, one with rapid KCl equilibration faciliated by a monovalent cation channel function and one with slow KCl equilibration. The mercurials increase the rates of KCl equilibration in both fractions, but produce higher rates in the fraction containing the channel function. The results are discussed in terms of pump and channel function and are compared with results for the electrical behavior of the CA2+-Mg2+-ATPase and other SR proteins in black lipid membranes, as presented by others.

Measurement of surface potential and surface charge densities of sarcoplasmic reticulum membranes.

The binding of the anionic fluorescent probe 1-anilino-8-naphthalene-sulfonate (ANS-) was used to estimate the surface potential of fragmented sarcoplasmic reticulum (SR) derived from rabbit skeletal muscle. The method is based on the observation that ANS- is an obligatory anion whose equilibrium constant for binding membranes is proportional to the electrostatic function of membrane surface potential, exp(e psi o/kT), where psi o is the membrane surface potential, e is the electronic charge, and kT has its usual meaning. The potential measured is characteristic of the ANS- bindings of phosphatidylcholine head groups and is about one-third as large as the average surface potential predicted by the Gouy-Chapman theory. At physiological ionic strength the surface potentials, measured by ANS-, referred to as the aqueous phase bathing the surface, were in the range -10 to -15 mV. This was observed for the outside and inside surfaces of the Ca2+-ATPase-rich fraction of the SR and for both surfaces of the SR fraction rich in acidic Ca2+ binding proteins. The inside and outside surfaces were differentiated on the basis of ANS- binding kinetics observed in stopped-flow rapid mixing experiments. A mechanism by which changes in Ca2+ concentration could give rise to an electrostatic potential across the membrane and possibly result in changes in Ca2+ permeability. The dependence of the surface potential on the monovalent ion concentration in the medium was used together with the Gouy-Chapman theory to determine the lower limits for the surface charge density for the inside and outside surfaces of the two types of SR. Values for the Ca2+-ATPase rich SR fraction were between 2.9 X 10(3) and 3.8 X 10(3) esu/cm2, (0.96 X 10(-6) and 1.26 X 10(-6) C/cm2) with no appreciable transmembrane asymmetry. A small amount of asymmetry was observed in the values for the inside and outside surfaces of the fraction rich in acidic binding proteins which were ca. 6.6 X 10(3) and ca. 2.2 X 10(3) esu/cm2 (2.2 X 10(-6) and 0.73 X 10(-6) C/cm). The values could be accounted for by the known composition of negatively-charged phospholipids in the SR. The acidic Ca2+ binding proteins were shown to make at most a small contribution to the surface charge, indicating that their charge must be located at least several tens of A from the membrane surface. The experiments gave evidence for a Donnan effect on the K+ distribution in the fraction rich in acidic binding proteins. This could be accounted for by the known concentration of acidic binding proteins in this SR fraction. The equilibrium constant for ANS- was shown to be more sensitive to changes in the divalent cation concentration than to changes in the monovalent cation concentration, as predicted by the Gouy-Chapman theory. Use of these findings together with the stopped-flow rapid mixing techniques constitutes a method for rapid and continuous monitoring of changes in ion concentrations in the SR lumen.

The pH dependence and the binding equilibria of the calcium indicator--arsenazo III.

The underlying principles of binding equilibria of arsenazo III with Ca2+ and Mg2+ are presented. Ca2+ and Mg2+ can bind arsenazo III in several different protonated forms depending on pH. The binding affinities of these different protonated forms of arsenazo III with Ca2+ increase in the order of H4A4- less than H3A5- less than H2A6- and with Mg2+, H4A4- congruent to H3A5- less than H2A6-. Arsenazo III is not membrane bound. The sensitivity ratio of arsenazo III with Ca2+ to arsenazo III with Mg2+ is close to two orders of magnitude. Arsenazo III and its complexes are extremely sensitive to pH changes. With 5 microM arsenazo III, the minimum detectable amount of Ca2+ can be as low as 0.08 microM. Contrary to current belief, we found that Mg2+ can bind to arsenazo III in a slightly acidic medium. Potential applications of arsenazo III to the study of membrane Ca2+ transport are also discussed.

Divalent cation-induced aggregation of chromaffin granule membranes.

Divalent cations induce the aggregation of chromaffin granule ghosts (CG membranes) at millimolar concentrations. Monovalent cations produce the same effect at 100-fold higher concentrations. The kinetics of the dimerization phase were followed by light-scattering changes observed in stopped-flow rapid mixing experiments. The rate constant for Ca2+-induced dimerization (kapp) is 0.86-1.0 x 10(9) M-1sec-1, based on the "molar" vesicle concentration. This value is close to the values predicted by theory for the case of diffusion-controlled reaction (7.02 x 10(9) M-1sec-1), indicating that there is no energy barrier to dimerization. Arrhenius plots between 10 degrees and 42 degrees C support this; the activation energy observed, +4.4 Kcal, is close to the value (4.6-4.8 Kcal) predicted for diffusion control according to theory. Artificial vesicles prepared from CG lipids were also found to have cation-induced aggregation, but the rates (values of kapp) were less than 1/100 as large as those with native CG membranes. Also, significant differences were found with respect to cation specificity. It is concluded that the slow rates are due to the low probability that the segments of membrane which approach will be matched in polar head group composition and disposition. Thus large numbers of approaches are necessary before matched segments come into aposition. The salient features of the chromaffin granule membrane aggregation mechanism are as follows: (a) In the absence of cations capable of shielding and binding, the membranes are held apart by electrostatic repulsion of their negatively charged surfaces. (b) The divalent and monovalent cation effects on aggregation are due to their ability to shield these charges, allowing a closer approach of the membrane surfaces. (c) The major determinants of the aggregation rates of CG membranes are proteins which protrude from the (phospholipid) surface of the membrane and serve as points of primary contact. Transmembrane contact between these proteins does not require full neutralization of the surface charge and surface potential arising from the negatively charged phospholipids. (d) After contact between proteins is established, the interaction between membranes can be strengthened through transmembrane hydrogen bonding of phosphatidyl ethanolamine polar head groups, divalent cation-mediated salt bridging, and segregation of phosphatidylcholine out of the region of contact.

High and low affinity Ca2+ binding to the sarcoplasmic reticulum: use of a high-affinity fluorescent calcium indicator.

The fluorescent calcium indicator, calcein, has been used as a high-affinity indicator of Ca2+ in the aqueous phase at physiological pH in the study of high-affinity calcium binding to sarcoplasmic reticulum (SR). The binding constant of the indicator at physiological pH is 10(3)-10(4) M-1 and increases with increasing pH. The binding mechanism of the indicator with Ca2+ and Mg2+ is described. Application of calcein as an aqueous indicator of Ca2+ binding to the SR at room temperature has revealed two classes of binding sites: one with high capacity and low affinity (ca. 820 nmol/mg protein, Kd = 1.9 mM), and another with low capacity and higher affinity (ca. 35 nmol/mg protein, Kd = 17.5 micronM). The divalent cation specificity of the low-affinity site is low and Ca2+/Mg2+ specificity of the high-affinity site is high. Quantitative studies of the bindings indicate that the high-affinity site residues in the Ca2+ ATPase (carrier) protein and represents complexation in the active site of the carrier and that the low-affinity site residues in the nonspecific acidic binding proteins. The contribution of Donnan equilibrium effects to the measured binding is shown to be insignificant. Stopped flow kinetic studies of Ca2+ passive binding with calcein and arsenazo III dyes have demonstrated that the binding to high-affinity site is very fast and that the overall binding reaction with the low-affinity site is slow, with a time course of about 4 s. Our analysis has shown that at least part of the low-affinity acidic proteins are within the SR matrix and that Ca2+ can reach them only by transversing the membrane via the Ca2+ carrier (Ca2+ ATPase). A model of the SR is proposed that accounts for several functional properties of the organelle in terms of its known protein composition and topological organization.

High-pressure spectral studies of mixed valence compounds of antimony.

In Cs(2)SbCl(6) and related compounds antimony appears as Sb(III) and Sb(V) in alternate halide octahedra. The optical spectrum contains "mixed valence" peaks assigned to Sb(III) --> Sb(V) transfer near 18 and 27 kK (cm(-1)). In addition there is a peak near 31 kK assigned to an internal transition on Sb(III) and one near 37 kK assigned to Sb(V), mixed with the absorption edge of the crystal. The mixed valence peaks shift strongly to lower energy with pressure (about 5 kK in 120 kbar), and decrease rapidly in integrated intensity, as does the Sb(III) peak near 31 kK. A new peak appears near 33-34 kK, tentatively assigned to Sb(IV). The ground state apparently transforms from Sb(III)-Sb(V) to Sb(IV)-Sb(IV) at high pressure. Similar behavior is observed for Cs(2)Sb(0.3)Sn(0.7)Cl(6) and (CH(3)CH(2)NH(3))(2)Sb(0.5)Sn(0.5)Cl(6).