KATP channels depress force by reducing action potential amplitude in mouse EDL and soleus muscle.

Although ATP-sensitive K+ (KATP) channel openers depress force, channel blockers have no effect. Furthermore, the effects of channel openers on single action potentials are quite small. These facts raise questions as to whether 1) channel openers reduce force via an activation of KATP channels or via some nonspecific effects and 2) the reduction in force by KATP channels operates by changes in amplitude and duration of the action potential. To answer the first question we tested the hypothesis that pinacidil, a channel opener, does not affect force during fatigue in muscles of Kir6.2-/- mice that have no cell membrane KATP channel activity. When wild-type extensor digitorum longus (EDL) and soleus muscles were stimulated to fatigue with one tetanus per second, pinacidil increased the rate at which force decreased, prevented a rise in resting tension, and improved force recovery. Pinacidil had none of these effects in Kir6.2-/- muscles. To answer the second question, we tested the hypothesis that the effects of KATP channels on membrane excitability are greater during action potential trains than on single action potentials, especially during metabolic stress such as fatigue. During fatigue, M wave areas of control soleus remained constant for 90 s, suggesting no change in action potential amplitude for half of the fatigue period. In the presence of pinacidil, the decrease in M wave areas became significant within 30 s, during which time the rate of fatigue also became significantly faster compared with control muscles. It is therefore concluded that, once activated, KATP channels depress force and that this depression involves a reduction in action potential amplitude.

Modulation of integrin-mediated intercellular adhesion during the interaction of thymocytes with stromal cells expressing VLA-4 and LFA-1 ligands.

Mature peripheral T cells closely regulate their intercellular interactions by modulating integrin adhesion functions. The ability of members of the integrin family to mediate intercellular adhesion is dependent on signals from within the cells (inside-out signaling) that increase the avidity of integrins for their ligands. These changes in avidity are independent of the quantitative changes on the number of receptors, and there is evidence to suggest that phosphorylation events play a predominant role in the regulation of the avidity state of the integrins. Whether such regulatory mechanisms are operative during T cell development had hitherto been an opened question. In the present work, we have used an in vitro adhesion assay between thymocytes and target cells expressing VLA-4 and LFA-1 counter ligands to determine how thymocytes can discriminate between integrin-specific signals during T cell development. Our findings are that VLA-4, but not LFA-1, is constitutively expressed in its high-avidity state during the early stages of T cell development, and that the high-avidity state of thymocytes for VCAM-1-expressing cells is closely regulated by signaling through protein kinase C and protein tyrosine kinase pathways. At later stages of development, mature thymocytes prior to leaving the thymus turn off both VLA-4 and LFA-1 adhesion functions. Our results show that the low-affinity state of integrins on peripheral mature T cells is established before mature thymocytes leave the thymus. Only when mature T cells recognize antigenic peptides in the context of major histocompatibility complex in the periphery will they turn on the adhesion function of VLA-4 and/or LFA-1 integrins.

Long-term intraperitoneal deferoxamine for hemochromatosis.

Intraperitoneal deferoxamine is a well established treatment for aluminum accumulation syndrome in patients with end-stage renal disease receiving peritoneal dialysis, but the use of intraperitoneal deferoxamine has not been described outside of the setting of chronic renal failure. We present here a case of secondary hemochromatosis, complicated by cirrhosis and cardiomyopathy, in which a chronic peritoneal dialysis catheter was used both to treat ascites and to deliver parenteral deferoxamine for iron overload. Daily urinary iron excretion was similar to that achieved when using standard routes of deferoxamine administration. Over a 2-year period, reversal of both the biochemical indicators and the clinical manifestations of iron overload was accomplished.

Human alpha(1,3/1,4)-fucosyltransferases discriminate between different oligosaccharide acceptor substrates through a discrete peptide fragment.

Five different human alpha(1,3)-fucosyltransferase (alpha(1,3)-Fuc-T) genes have been cloned. Their corresponding enzymes catalyze the formation of various alpha(1,3)- and alpha(1,4)-fucosylated cell surface oligosaccharides, including several that mediate leukocyte-endothelial cell adhesion during inflammation. Inhibitors of such enzymes are predicted to operate as anti-inflammatory agents; in principle, the isolation or design of such agents may be facilitated by identifying peptide segment(s) within these enzymes that interact with their oligosaccharide acceptor substrates. Little is known, however, about the structural features of alpha(1,3)-Fuc-Ts that dictate acceptor substrate specificity. To begin to address this problem, we have created and functionally characterized a series of 21 recombinant alpha(1,3)-Fuc-T chimeras derived from three human alpha(1,3)-Fuc-Ts (Fuc-TIII, Fuc-TV, and Fuc-TVI) that maintain shared and distinct polypeptide domains and that exhibit common as well as idiosyncratic acceptor substrate specificities. The in vivo acceptor substrate specificities of these alpha(1,3)-Fuc-T chimeras, and of their wild type progenitors, were determined by characterizing the cell surface glycosylation phenotype determined by these enzymes, after expressing them in a mammalian cell line informative for the synthesis of four distinct alpha(1,3)- and alpha(1,4)-fucosylated cell surface oligosaccharides (Lewis x, sialyl Lewis x, Lewis a, and sialyl Lewis a). Our results indicate that as few as 11 nonidentical amino acids, found within a "hypervariable" peptide segment positioned at the NH2 terminus of the enzymes' sequence-constant COOH-terminal domains, determines whether or not these alpha(1,3)-Fuc-T can utilize type I acceptor substrates to form Lewis a and sialyl Lewis a moieties.

Mechanisms underlying the depressed production of interleukin-2 in spleen and lymph node cell cultures of mice infected with Mycobacterium bovis BCG.

Mice were infected intravenously with 1.0 mg of Mycobacterium bovis BCG. At various times thereafter, spleen and peripheral lymph node cells were stimulated with concanavalin A for 18 to 20 h, and their capacity to produce interleukin-2 (IL-2) was evaluated by means of a T-cell blast proliferation technique. A depression of IL-2 production that was complete in the spleen but partial in lymph node cell cultures occurred at 2 to 3 weeks and persisted till weeks 8 to 10 after infection. No direct evidence was found for an active suppressor mechanism depressing in vitro the production of IL-2. In spleen cell cultures the suppression of IL-2 production would result from a functional defect of the IL-2-producing T-cell subset, whereas in lymph node cell cultures the depression mainly results from a relative lack of IL-2-producing cells caused by an accumulation of immunoglobulin-positive and "null" cells. Spleen cells from BCG-infected mice maintained their capacity to acquire IL-2 receptors when activated by concanavalin A.

Identification of analogs of the hallucinogen cyclohexamine.

The hallucinogenic drug 1-phenylcyclohexylamine and its N-methyl-, N-ethyl-, N-propyl-,N-isopropyl-, and N-(beta-phenylethyl)-derivatives are identified by spectroscopic techniques. Reference mass and infrarred spectra and gas-liquid and thin layer chromatographic data are provided and discussed.

Spectroscopic and chromatographic identification of dimethylamphetamines.

The qualitative analysis of the 6 isomeric dimethylamphetamines (both methyl groups on the aromatic ring) is described. Their ultraviolet, mass, and proton magnetic resonance spectra are similar, but infrared spectra allow a positive identification to be made, and reference spectra are provided. Melting point and gas-liquid and thin layer chromatographic data are also presented.

Heterotypic exclusion between vesicular stomatitis viruses of the New Jersey and Indiana serotypes.

Co-infection of cells with vesicular stomatitis viruses of the Indiana and New Jersey serotypes results in interference. Using specifically-labelled immunofluorescent antibodies, it was demonstrated that within any one co-infected cell, one virus serotype replicated to the relative exclusion of the other serotype. This result was further substantiated by an examination of the virus serotypes released by infectious centres co-infected with both viruses. Dominance of one serotype over the other was shown to be a function of the relative multiplicity of the two viruses. Superinfection by the second serotype at a higher multiplicity resulted in dominance by the second virus during the early period (up to 1-5 h) post-infection. After this time, the minority virus was able to overcome this dominance. Dominance of the majority virus was also abolished by u.v; inactivation. Cell protein synthesis appeared to be less affected in cells infected with both serotypes than when infection was with a single serotype.

Application of trichloroacetyl isocyanate to NMR analysis of steroids of pharmaceutical interest I: corticosteroids and chemically related compounds.

The trichloroacetyl carbamates of 38 corticosteroids and chemically related compounds were prepared, and their NMR spectra in deuterochloroform were obtained. The effects of the introduction of a number of functional groups on the chemical shift of the carbamate proton signals were determined.

Identification of the N-methylated analogs of the hallucinogenic amphetamines and some isomers.

The drugs 2-, 3-, and 4-methoxy-N-methylamphetamine, 3-methoxy-4,5-methylenedioxy-N-methylamphetamine, and 3,4-methylenedioxy-N-methylamphetamine are identified by spectroscopic techniques. The ultraviolet and mass spectra of isomers are similar, but proton magnetic resonance and infrared spectra are distinctly different, and reference spectra and data are provided. Gas-liquid and thin layer chromatographic systems for the analysis are discussed.