Two phases of zymogen granule lifetime in mouse pancreas: ghost granules linger after exocytosis of contents

Different cell types show widely divergent mechanisms and kinetics of exocytosis. We investigated these processes in pancreatic acinar cells by using video-rate 2-photon microscopy to image entry of extracellular dye into individual zymogen granules undergoing exocytosis. Fluorescence signals display two distinct phases; an initial peak that then decays over several seconds to a prolonged plateau. Several observations suggest that the first component reflects the binding of dye to the granule contents and their subsequent release into the acinar duct. These observations include: the peak/plateau fluorescence ratio differs between different dyes; the initial fluorescence decay mirrors the loss of granule contents as monitored by differential interference contrast microscopy; and the fall in vesicular fluorescence is accompanied by a rise in fluorescence in the adjacent duct lumen. We thus propose the use of extracellular fluorescent probes as a convenient means to monitor the kinetics of loss of proteinaceous content from secretory granules. In pancreatic acinar cells the fusion pore remains open much longer than required to ensure secretion of the granule contents, and instead the persistent empty 'ghost-granule' may act as a conduit to which secondary granules can fuse and release their contents by compound exocytosis.     

Peripheral spin exchange nuclear spin resonance angiography: Studies of the effect of blood flow on contrast

Sequential two-dimensional (2-D) inflow MR-angiography (SDSMRA) utilizes the bloodflow-induced changes in the MR-signal for intraluminal vessel contrast. To assess the effect of the peripheral arterial flowpulse on the average intraluminal MR-signalintensity (MIMRSI) seven healthy subjects, mean age 31 +/- 4 years, were studied using color Doppler flow mapping and SDSMRA. In each subject the regional MIMRSI was measured in 21 vascular segments and correlated with the regional flowpulse in a simple regression model. MIMRSI ranged from 915 +/- 299 in the right profunda femoris to 1378 +/- 283 in the distal abdominal aorta: distally, the trifurcation it was, with the exception of the posterior tibial artery in three subjects (MIMRSI = 545 +/- 64), not measurable. The regional differences in the peripheral flowpulse were without statistically measurable effect on MIMRSI. MIMRSI was best correlated to the systolic forward flow in 14, to the backward flow in 3, and to the mean forward flow in 1 vascular segment, respectively. The mean correlation coefficient (r) was 0.56, ranging from r = 0.11 in the right popliteal artery to r = 0.93 in the right superficial femoral artery. In the distal abdominal aorta no positive correlation between the blood flow and MIMRSI was determined. The regional differences in the peripheral arterial flow-pulse have no significant effect on MIMRSI in healthy men. MIMRSI correlates best with the systolic forward flow. However, on the average, 44% of the MIMRSI appear to be related to flow-independent yet unidentified factors. Optimal design of SDSMRA pulse sequences will require a better characterization of flow-dependent and flow-independent MIMRSI parameter.     

Detection of antibodies to human immunodeficiency virus by latex agglutination with recombinant antigen.

Recombinant human immunodeficiency virus (HIV) env antigen was attached to polystyrene particles, and these complexes were used to develop the first latex agglutination assay for antibodies to HIV. A total of 95 positive and 116 negative human serum samples were assayed for antibodies to HIV by latex agglutination, and results were compared with those of a commercial enzyme immunoassay. Latex agglutination was also compared with, and found to be completely concordant with, Western blot (immunoblot) analysis with virion antigens.     

Erholungsfähigkeit der Niere nach zwei- oder dreistündiger Ischämie und Resektion der gesunden Niere,kontrolliert während einer neunmonatigen Erholungszeit

Zusammenfassung Die linke Kaninchenniere wurde einer ischämischen Belastung von 2 oder 3 Std ausgesetzt und die rechte Niere 21 Tage nach der Belastung entfernt. Während einer neunmonatigen Erholungszeit bestimmten wir im Katheterharn Gefrierpunktsdepression, Eiweiß, Glucose, Harnstoff, Ammoniak, Natrium, Kalium, Chlor und im Plasma den Rest-N. Das Experiment wurde durch einen Konzentrierungsversuch abgeschlossen und die Nieren nach Tötung der Tiere histologisch untersucht.Die Nierenfunktion erholt sich in wenigen Tagen nach der Resektion der gesunden Niere vollständig ohne Ausfälle von Einzelfunktionen. Gefrierpunktsdepression und Konzentrierungsversuch zeigen eine fast normale Anpassungsbreite der belasteten Nieren. Es kommt in keinem Fall zur Retention harnpflichtiger Substanzen. Anzeichen für die Ausbildung einer Schrumpfniere sind nicht vorhanden.Die histologische Untersuchung der Nieren zeigt bis auf den Ausfall einzelner Nephrone einen nahezu normalen Befund. Die Erholung der Nierenzellen ist morphologisch und funktionell vollständig, obwohl der Schwund an RNS bei gleicher Belastung 40–45% beträgt.Außer der für die Niere früher festgestellten Unabhängigkeit der Erholungsfähigkeit von der anaeroben Glykolyse übersteht der Informationsbestand der Niere unter Erhaltung des Bauplans ebenfalls einen absoluten Sauerstoffmangel von 3 Std ohne Schaden.Mit 3 TextabbildungenDer Deutschen Forschungsgemeinschaft danken wir für großzügige Förderung unserer Untersuchungen.Auszugsweise vorgetragen auf der 28. Tagung der Deutschen Physiologischen Gesellschaft, Juni 1963 in Köln.     

Uptake of dehydroascorbic acid and ascorbic acid to isolated nerve terminals and secretory granules from ox neurohypophyses

When uptake of L-[14C]ascorbic acid ([14C]AA) to various organs in guinea-pigs was studied after intracardiac injection, the adenohypophysis, pars intermedia, and the neurohypophysis had an uptake per milligramme protein which was about half of the uptake to the adrenals. Adrenal uptake was 20 +/- 2.8 pmol mg-1 protein microCi-1 injected. The uptake to the different parts of the hypophysis was considerably higher than the uptake to pancreas, liver, kidney, spleen and other organs. When isolated nerve endings (neurosecretosomes) from ox neurohypophyses were incubated with a medium containing labelled dehydroascorbic acid ([14C]DHA), the uptake was much slower than when the medium contained labelled ascorbic acid. The uptake of [14C]DHA showed a linear dependence on concentration, and was not influenced by addition of Mg2+ and ATP. Addition of Mg2+ + ATP, omission of Ca2+ and Mg2+ or exchange of Na+ in the medium with K+ had no effect on the uptake of ascorbic acid. When isolated secretory granules from ox neurohypophyses were incubated with a medium containing [14C]DHA, uptake was considerably faster than the uptake when they were incubated in a medium containing [14C]AA. The uptake of dehydroascorbic acid was linear with the concentration in the medium and was not changed by addition of Mg2+ ATP. Addition of 10 mM NH4Cl or exchange of 120 mM K+ in the incubation medium with Na+ did not change the uptake of dehydroascorbic acid. The contents of copper, zinc, iron and cobalt were determined in isolated nerve endings (A) and membranes (B) as well as in lysate (C) from isolated neurosecretory granules. The results (in nmol mg-1 protein) were for Cu: (A): 0.25 +/- 0.01 (SEM), (B): 0.67 +/- 0.16, (C): 0.22 +/- 0.06; for Zn: (A): 0.53 +/- 0.13, (B): 6.97 +/- 0.75, (C): 1.8 +/- 0.53; and for Fe: (A): 15.6 +/- 1.9, (B): 6.92 +/- 0.32, (C): 3.15 +/- 0.43. In all preparations the cobalt content was below the detection limit (less than 5 pmol mg-1 protein).     

Polyamines in nerve terminals and secretory granules isolated from neurohypophyses.

In isolated nerve terminals from ox neurohypophyses the following concentrations of polyamines [pmol (microgram protein)-1 (mean +/- SEM)] were found: spermine: 2.07 +/- 0.14 (n = 3), spermidine: 0.22 +/- 0.01 (n = 4), putrescine: 0.20 +/- 0.01 (n = 4). In secretory granules isolated from the same tissue, the concentrations were: spermine: 0.57 +/- 0.02 (n = 3), spermidine: 0.07 +/- 0.04 (n = 3), putrescine: 0.13 +/- 0.04 (n = 3). After incubation of isolated nerve terminals with the polyamines, they were taken up as a function of time and concentration, approaching saturation at high concentrations. The kinetic parameters of their synthesizing enzyme, ornithine decarboxylase, in ox neurohypophyseal nerve terminals (apparent Km 0.75 mM and Vmax 22.5 pmol mg protein-1 h-1) were comparable to those previously found in cerebral cortex of rats. When isolated, hemilobes from rat neurohypophyses were incubated in a medium which contained spermidine (5 mM), and were stimulated by 56 mM K+, release of vasopressin was smaller than in control experiments. However, after removal of spermidine and after restimulation, 50 min after initial stimulation, the release was significantly elevated. It is suggested that polyamines may take part in modulation of vasopressin release.