Comparison of the properties of active Ca2+ transport by the islet-cell endoplasmic reticulum and plasma membrane

The properties of active or ATP-dependent calcium transport by islet-cell endoplasmic reticulum and plasma membrane-enriched subcellular fractions were directly compared. These studies indicate that the active calcium transport systems of the two membranes are fundamentally distinct. In contrast to calcium uptake by the endoplasmic reticulum-enriched fraction, calcium uptake by islet-cell plasma membrane-enriched vesicles exhibited a different pH optimum, was not sustained by oxalate, and showed an approximate 30-fold greater affinity for ionized calcium. A similar difference in affinity for calcium was exhibited by the Ca²⁺-stimulated ATPase activities which are associated with these islet-cell subcellular fractions. Consistent with the effects of calmodulin on calcium transport, calmodulin stimulated Ca²⁺-ATPase in the plasma membranes, but did not increase calcium-stimulated ATPase activity in the endoplasmic reticulum membranes. The physiological significance of the differences observed in calcium transport by the endoplasmic reticulum and plasma membrane fractions relative to the regulation of insulin secretion by the islets of Langerhans is discussed.     

Characteristics and determinants of osmotic lysis in chromaffin granules

(1) Using isolated bovine chromaffin granules, we demonstrate that osmotic lysis is not a random process and establish the osmotic pressure dependence of osmotic lysis in chromaffin granules, the so-called osmotic fragility curve. (2) We show by measuring the release of constituents of the granule core and correlating these with changes in spectroscopic parameters (turbidity and endogenous catecholamine fluorescence), that the latter can be safely used to measure lysis. (3) Within a particular granule population, noradrenaline granules lyse at higher osmolarities than adrenaline granules, suggesting a higher core osmolarity of the noradrenaline granules. (4) The size distribution of chromaffin granules as a function of lysis was determined by the use of whole mount electron microscopy. It is shown that the mean size of chromaffin granules decreases as a function of lysis. (5) On the basis of theoretical considerations three alternative models of the sequence of osmotic lysis in chromaffin granules are proposed. The experimental results best support a model which postulates that during partial osmotic lysis, granule membranes reseal into smaller vesicles after graded release of contents. The osmotic fragility would represent several cycles of lysis and resealing and would not be a reflection of the distribution of osmotic pressures in the granule population.     

3',5'-Cyclic Adenosine Monophosphate- and Ca2+-Calmodulin-Dependent Endogenous Protein Phosphorylation Activity in Membranes of the Bovine Chromaffin Secretory Vesicles: Identification of Two Phosphorylated Components as Tyrosine Hydroxylase and Protein Kinase Regulatory Subunit Type II

Abstract: Membranes of the secretory vesicles from bovine adrenal medulla were investigated for the presence of the endogenous protein phosphorylation activity. Seven phosphoprotein bands in the molecular weight range of 250,000 to 30,000 were observed by means of the sodium dodecyl sulphate electrophoresis and autoradiography. On the basis of the criteria of molecular weight, selective stimulation of the phosphorylation by cyclic AMP (as compared with cyclic GMP) and immunoprecipitation by specific antibodies, band 5 (molecular weight 60,300) was found to represent the phosphorylated form of the secretory vesicle-bound tyrosine hydroxylase. The electrophoretic mobility, the stimulatory and inhibitory effects of cyclic AMP in presence of Mg²⁺ and Zn,²⁺ respectively, and immunoreactivity toward antibodies showed band 6 to contain two forms of the regulatory subunits of the type II cyclic AMP-dependent protein kinase, distinguishable by their molecular weights (56,000 and 52,000, respectively). Phosphorylation of band 7 (molecular weight 29,800) was stimulated about 2 to 3 times by Ca²⁺ and calmodulin in the concentration range of both agents believed to occur in the secretory tissues under physiological conditions.     

Neurotensin interacts with carbachol, secretin, and caerulein in the stimulation of the exocrine pancreas of the rat in vitro

In the present investigation the effect of neurotensin on pancreatic secretion of isolated pancreatic lobules from the rat was examined. We found a dose- and time-dependent stimulation of amylase release beginning with a concentration of 10(-9) M neurotensin. This response was potentiated by the cholinergic agonist carbachol, the gastrointestinal peptide secretin, and the CCK analogue caerulein. As we found neurotensin-immunoreactive nerves within the pancreas and as neurotensin-like immunoreactivity is present in the circulation (found previously), neurotensin may well be a further peptide taking part in the regulation of exocrine pancreatic secretion either as a hormone or a neurotransmitter. Neurotensin would then cooperate with cholinergic mechanisms, secretin, and CCK.     

The role of the Asp-32 residue of cholecystokinin in gastric acid secretion and gastrin receptor recognition

The aspartic acid residue at the penultimate position is known to be essential for the hormonal activity of CCK and gastrin on gastric acid secretion. This residue was successively replaced by beta-aspartic acid, beta-alanine, and glutamic acid in the C-terminal heptapeptide of CCK 27-33. The analogues obtained were tested on rat gastric acid secretion and for recognition by gastrin receptors. The replacement by beta-aspartic or beta-alanine decreased gastric secretion and gastrin receptor recognition. In contrast, replacement by glutamic acid affected these two parameters less. The nature of the N-blocking group (Boc or Z) also influenced these activities, Boc derivatives being more potent than Z derivatives. The results were compared to those previously obtained on pancreatic secretion and on stimulation of gall bladder contraction where the modifications were found capable of differentiating between cholecystokinin, pancreozymin and gastrin activities.     

Survival of the parasitic protozoan, Babesia bigemina, in blood cooled at widely different rates to -196 degrees C

Survival of the parasitic protozoan, Babesia bigemina, in blood cooled at widely different rates to ? 196°C. International Journal for Parasitology4: 169–172. The infectivity of Babesia bigemina in blood containing 2 m DMSO was tested in 99 cattle after the blood had been cooled to ? 196°C at eight rates ranging from 0·73–3070°C/min. Blood cooled at each rate was infective; 95 of the recipients became infected, the exceptions being four of the seven cattle inoculated with blood cooled at 3070° C/min. The infectivity of blood cooled at 39, 82 and 212°C/min was higher than that of blood cooled at slower or faster rates. Least depression of infectivity occurred at 82°C/min.     

Apokrine Sekretion der peritrophischen Membran von Chironomus thummi piger Str. (Diptera)

Zusammenfassung Am Beispiel der Larven von Chironomus thummi piger werden die Zellen, die die peritrophische Membran abscheiden, licht- und elektronenmikroskopisch untersucht. Es liegen zwei Zelltypen vor, die einerseits durch ihren Reichtum an granulären E.R.-Schläuchen (ER-Zellen), andererseits durch ihren hohen Gehalt an Mitochondrien (M-Zellen) charakterisiert sind. Die ER-Zellen zeigen Veränderungen, die in vielerlei Hinsicht der klassischen Auffassung der apokrinen Sekretion entsprechen und sich nicht in ein modernes Schema der Sekretionsmorphologie einordnen lassen. Die Zellen gehen im Verlauf der Sekretabgabe weder völlig zugrunde, noch bleiben sie vollständig erhalten. Die M-Zellen, die im Gegensatz zu den ER-Zellen keine Sekretionsgranula besitzen, weisen ähnliche Umwandlungen auf. Das fertige Sekretionsprodukt — die peritrophische Membran — entstammt einmal vorgeformten Sekretionsgranula, zum anderen der umgewandelten, abgeschnürten oberen Zellhälfte. Die peritrophische Membran besteht aus zwei Schichten, wovon die lumenseitige, auf Längsschnitten quergestreifte Lage (Wabentextur) vermutlich aus den Sekretionsgranula hervorgeht und die andere längsgefaserte Lage auf die Umwandlung von Zytoplasma zurückzuführen sein dürfte. Die Mikrovilli der Sekretionszellen sind in keiner Weise für die Strukturierung der Wabentextur verantwortlich.

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On the apocrine secretion in the formation of the peritrophic membrane of chironomus thummi piger Str

Summary Taking the larvae of Chironomus thummi piger as an example, the cells secreting the peritrophic membrane have been investigated with the light- and electron-microscope. Two cell-types can be distinguished which in one case are characterized by abundant rough E.R.-tubules (ER-cells) and in the other case by large quantities of mitochondria (M-cells). The ER-cells undergo changes which in many respects correspond to the individual stages of the classic apocrine secretion, and thus do not fit into a modern scheme of the morphology of secretion. In the course of the discharge of the secretory product the cell neither becomes completely necrotic nor remains totally intact. The M-cells, which do not contain secretion granules like the ER-cells, show similar changes. The final product — the peritrophic membrane — is formed on the one hand by membrane bound secretion granules and on the other hand by the transformed and pinched off upper half of the cell. The peritrophic membrane consists of two layers, the one of which, facing the lumen of the midgut and exhibiting a honey-comb-texture, presumably is formed by the secretion granules, whereas the other layer arises from transformed cytoplasm. The microvilli of the secretory cells are not responsible for the formation of the honey-comb-pattern.

Die Untersuchungen wurden mit Unterstützung durch die Max-Planck-Gesellschaft und die Deutsche Forschungsgemeinschaft durchgeführt.     

Cyproheptadine metabolites inhibit proinsulin and insulin biosynthesis and insulin release in isolated rat pancreatic islets

The contribution of drug metabolites to cyproheptadine (CPH)-induced alterations in endocrine pancreatic -cells was investigated by examining the inhibitory activity of CPH and its biotransformation products, desmethylcyproheptadine (DMCPH), CPH-epoxide and DMCPH-epoxide, on hormone biosynthesis and secretion in pancreatic islets isolated from 50-day-old rats. Measurement of (pro)insulin (proinsulin and insulin) synthesis using incorporation of ³H-leucine showed that DMCPH-epoxide, DMCPH and CPH-epoxide were 22, 10 and 4 times, respectively, more potent than CPH in inhibiting hormone synthesis. The biosynthesis of (pro)insulin was also inhibited by CPH and DMCPH-epoxide in islets isolated from 21-day-old rat fetuses. The inhibitory action of CPH and its metabolites was apparently specific for (pro)insulin, and the synthesis of other islet proteins was not affected. Other experiments showed the metabolites of CPH were active in inhibiting glucose-stimulated insulin secretion but were less potent than the parent drug in producing this effect. CPH and its structurally related metabolites, therefore, have differential inhibitory activities on insulin synthesis and release. The observation that CPH metabolites have higher potency than CPH to inhibit (pro)insulin synthesis, when considered with published reports on the disposition of the drug in rats, indicate that CPH metabolites, particularly DMCPH-epoxide, are primarily responsible for the insulin depletion observed when the parent compound is given to fetal and adult animals.Abbreviations CPH cyproheptadine - CPH-epoxide cyproheptadine-10-11-epoxide - DMCPH desmethylcyproheptadine - DMCPH-epoxide desmethylcyproheptadine-10,11-epoxide - HPLC high-performance liquid chromatography - KBB Krebs biocarbonate buffer Recipient of a Society of Toxicology Predoctoral Research Fellowship.Present address: Department of Biochemistry, The University of Hong Kong, Hong Kong.     

Age and growth of Lutjanus kasmira (Forskål) in Hawaiian waters

The growth of Hawaiian taape, Lutjanus kasmira , was studied by examining otoliths and by analysing length-frequency distribution. Annual hyaline and opaque markings were visible in whole mounts of sagittae, which were verified by enumeration of daily increments with a scanning electron microscope (SEM) and through marginal increment analysis. The von Bertalanffy growth curve was fitted to the data, resulting in: where t. l . is total length (cm) and t is age (years). SEM observations revealed that the slowgrowth hyaline zones were composed of daily increments too small (0.4–0.8 μm) to be resolved optically. Thus, age estimates derived by numerically integrating otolith growth rate data obtained with a light microscope showed a negative bias, resulting in overestimation of growth rates. Parameter estimates obtained from three different types of length-frequency analysis were also unstable. This was due, at least in part, to differences in the size composition of fish sampled with different fishing gears and from different depths.
The growth rate registered in Hawaii falls within the reported growth coefficients of lutjanids, whereas it is one of the highest in the Pacific and clearly higher than a deep-water lutjanid species growth in Hawaii. Probably, this high growth rate may have been enhanced by the relative lack of competitors in the depauperate Hawaiian marine fish community.