Activation of the cardiac ATP-sensitive K+ channel by ER-001533, a newly synthesized vasorelaxant.

Effects of ER-001533 (ER), a newly synthesized vasorelaxant, on the membrane currents were examined in single ventricular cells of guinea pigs. The patch-clamp technique was used in the "whole-cell" and "inside-out" patch configurations. In the whole-cell clamp condition, ER induced a time-independent K(+)-dominant current, which was inhibited by glibenclamide (1-3 microM), suggesting that ER activated the cardiac ATP-sensitive K+ channel (KATP). To elucidate the mechanism of ER-mediated KATP channel activation, the drug was applied to the inside-out patches before and after channel "run-down." Since nucleotide diphosphates could induce the channel openings after complete run-down, effects of the drug on the nucleotide diphosphate-induced channel openings were also examined. Before run-down, ER activated the KATP channel only in the presence of ATP. ER shifted the relation between [ATP]i and the channel activity to the right in a concentration-dependent fashion without a significant alteration of the slope. After channel run-down, ER did not affect the channel openings either in the absence or in the presence of UDP. However, ER could relieve the ATP-gamma-S inhibition of the UDP-induced channel openings. Thus, we conclude that ER antagonizes the inhibitory effect of ATP on the KATP channel in a competitive manner, thereby enhancing the channel openings.     

Bronchoalveolar lavage cell counts as a predictor of short term outcome in pulmonary sarcoidosis.

Sixty seven patients with biopsy proven pulmonary sarcoidosis were prospectively studied to determine whether single point bronchoalveolar lavage cell counts were a useful indicator of functional outcome and whether repeated lavage helped in management. The mean follow up period was 25 (range 13-37) months. No patient was having corticosteroid treatment at the time of initial bronchoalveolar lavage. "High intensity alveolitis" (lymphocyte count greater than or equal to 28%) was present at the initial lavage in 42 patients. These patients showed a significant improvement in their pulmonary function and chest radiographs over the follow up period whereas patients with "low intensity alveolitis" did not. Of the 42 patients with high intensity alveolitis, 31 had chronic sarcoidosis (duration over two years, mean 80 months). These patients showed a significant improvement in FVC but not in TLCO. Corticosteroids resulted in greater functional and radiological improvement in the patients with high intensity alveolitis than in those with low intensity alveolitis. Repeat bronchoalveolar lavage in 34 patients, mean 8.4 months after the original lavage, showed a weak inverse relation between a reduced lymphocyte count and change in forced vital capacity and isotope uptake on a gallium scan. These correlations were too weak to make repeated cell counts useful in management. Our results suggest that high intensity alveolitis may be a favourable prognostic factor for lung function in pulmonary sarcoidosis, even in patients with chronic disease, but that repeat lavage adds little to the management of the individual patient.     

Inhibitability and enhanceability of basophil histamine release in asthmatic and normal subjects

Circulating human basophils contain histamine, a potent mediator of inflammation. Previous in vitro studies have shown that histamine 'releasability' in asthmatic subjects differs from normal subjects but have not evaluated possible differences in the immunopharmacological control of the release of this mediator which might account for these differences. The purpose of the present study was to examine the immunopharmacologic control of basophil histamine release in 14 asthmatics and 10 normal subjects who were characterized by pulmonary function tests, allergic status (skin tests and serum IgE levels) and nonspecific airways reactivity to methacholine and histamine. Basophils were stimulated with anti-IgE, and the inhibitory effects of the H2 agonist, dimaprit, and dibutyryl cyclic AMP (dbcAMP), as well as the enhancing properties of 5-hydroperoxyeicosatetraenoic acid (5-HPETE) and indomethacin on the modulation of histamine release, were investigated. Although no statistically significant differences were seen in the percent histamine release triggered by anti-IgE in these two groups, enhancement of histamine release by 5-HPETE was more consistent in the asthmatic subjects (10 of 10) than in control subjects (6 of 8). The percent increase in histamine release produced by 5-HPETE in asthmatic subjects averaged 3.9 +/- 1.3% using 0.03 micrograms anti-IgE/ml and 4.8 +/- 3.2% using 0.1 microgram anti-IgE/ml (p less than 0.002, Wilcoxon's signed rank test), and averaged 3.0 +/- 4.3 and 3.1 +/- 5.3%, respectively, in control subjects (p greater than 0.10).(ABSTRACT TRUNCATED AT 250 WORDS)     

Study of circulating immune complex size in systemic lupus erythematosus.

The molecular size of circulating immune complexes in patients with systemic lupus erythematosus was determined by the C1q solid-phase assay after the sera were fractionated by sucrose-gradient ultracentrifugation. Circulating immune complexes in patients with membranous glomerulonephritis were uniformly large, sedimenting exclusively above 19S, whereas the immune complexes in patients with cerebritis were small, at or just above 7S. In lupus patients with diffuse proliferative glomerulonephritis and patients without renal involvement, immune complexes of both large and small sizes were found. Of patients without renal involvement, more circulating immune complexes were associated with active disease (n = 22, prevalence = 82%, mean level = 24 standard deviations) than with inactive disease (n = 17, prevalence = 41%, mean level = 41%, mean level = 6 . 5 standard deviations). In patients with clinical evidence for renal involvement, circulating immune complexes were detected in all of five patients with membranous glomerulonephritis, in 88% of 17 patients with diffuse proliferative glomerulonephritis and in one of four patients with mesangial nephritis. Thus, in addition to the finding of an overall positive correlation between disease activity and circulating immune complex levels, circulating immune complexes of certain general molecular size ranges appear to be associated with different clinical manifestations of systemic lupus erythematosus.     

Differential staining of human alpha beta and gamma delta T cells by the fluorescein conjugate of an anti-CD3 monoclonal antibody.

The enumeration of total T cells, an important function of the clinical immunology laboratory, utilizes antibodies to CD3, the macromolecular complex associated with the antigen-specific receptors of T cells. We compared the ability of some commonly employed commercial anti-CD3 reagents to stain human peripheral blood lymphocytes. Surprisingly, the fluorescein isothiocyanate (FITC) conjugate of Coulter clone T3 (FITC-T3) stained most T cells brightly, but selectively stained gamma delta T cells very dimly or not at all. In contrast, the other anti-CD3 reagents studied (FITC-Leu 4, PE-T3, PE-Leu 4, and indirectly labelled T3 and Leu 4) stained all T cells equivalently. Dual-colour flow cytometric analysis with FITC-T3 and PE-Leu 4 readily demonstrated a FITC-T3-/PE-Leu 4+ population of T cells. This unique population stained dimly or not at all with a combination of anti-CD4 and anti-CD8 monoclonal antibodies and positively with the pan-gamma delta T cell antibody TCR delta 1. Moreover, an excellent correlation was found between the number of FITC-T3-/PE-Leu 4+ cells and the number of TCR delta 1+ cells in 32 normal individuals. Thus, the FITC-T3-/PE-Leu 4+ phenotype accurately marks all gamma delta T cells. In contrast to FITC-T3, both PE-conjugated and unconjugated T3 stained gamma delta T cells brightly. Therefore, T3 binds to an epitope present on all T cells, but fluoresceinylation specifically attenuates this antibody's ability to bind to gamma delta T cells. These findings indicate that the use of FITC-T3 can result in a significant and variable underestimation of peripheral blood T cell number and demonstrate further that the CD3 complexes of human alpha beta and gamma delta T cells are significantly different.     

Extrasynaptic receptors on cell bodies of neurons in central nervous system of the leech.

1. A systematic study has been made of the sensitivity of identified sensory and motoneurons in the leech central nervous system to chemical transmitter substances. 2. The following substances elicited responses from the cell bodies of individual neurons: acetylcholine, 5-hydroxytryptamine, gamma-aminobutyric acid, glutamic acid, glycine, dopamine, and norepinephrine. Since the cell bodies of leech neurons are free of synapses, the receptors that give rise to these responses are extrasynaptic. 3. Sensory and motoneurons of different function had characteristic complements of extrasynaptic receptors. For example, mechanosensory cells responding to light touch, to pressure, and to noxious stimuli could be distinguished by their responses to iontophoretically applied compounds. For one of these modalities (nociceptive), neurons with different receptive fields but otherwise similar properties had markedly distinct extrasynaptic receptors. The possible significance of extrasynaptic receptors is discussed.     

Chronic amitriptyline administration increases serotonin transporter binding sites in the hippocampus of aged rats

The effects of ageing and of chronic antidepressant treatment upon 5-HT transporter sites ([3H]paroxetine binding) in the rat hippocampus was examined. [3H]paroxetine binding to transporter sites was decreased with ageing in the hippocampus of control rats (38% decrease in dentate gyrus and CA4). Amitriptyline (10 mg/kg, i.p.) had no significant effect on [3H]paroxetine binding in 10 months old rats, but increased binding sites in 24 months rats in all hippocampal subregions (greatest increase of 109% in CA1 compared to saline controls). These data indicate an age-related decrease in hippocampal serotonin transporter sites and upregulation of these sites following 10 weeks of amitriptyline. The observed increase in transporter sites following amitriptyline may contribute to the general lower effectiveness of tricyclic antidepressants with ageing.