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M Tyhurst

Publications and source records attributed to M Tyhurst.

At least 19 recordsLinked to original sources
PubMed1987

Use of osmium etching procedures for studying the ultrastructure of isolated rat islets of Langerhans.

Variants of the osmium etching procedure, described by Tanaka et al. for the demonstration of the structure of organelles by scanning electron microscopy, have been utilized in studies of isolated rat islets of Langerhans. Use of a preliminary glutaraldehyde fixation followed by the osmium etching procedure originally described, allowed effective removal of the cytoplasmic ground substance, as demonstrated by transmission electron microscopy. High resolution scanning electron microscopy of similar preparations permitted studies of the interaction of cytoskeletal elements with individual organelles. In particular, direct interactions of insulin storage granules with cytoskeletal elements were clearly evident.

Animals↗
PubMed1986

The cytoskeleton and insulin secretion.

One of the central, unresolved problems in our understanding of insulin secretion is the way in which stimulus recognition and its associated metabolic events are translated into the mechanical processes of insulin-storage granule movement and extrusion from the cells by exocytosis. In the present article we have examined the structural organization of the B-cell cytoskeleton in detail and have reviewed how drugs that affect the cytoskeleton alter insulin secretion. Available information about the interactions of tubulin, actin, myosin, and actomyosin with insulin-secretory granules is summarized, and a tentative model is proposed to explain how stimulus-effector system coupling might be achieved.

Actin Cytoskeleton↗
PubMed1984

Effects of monensin on biosynthesis and intracellular processing of proinsulin.

Monensin, a specific sodium ionophore, has been shown to reduce glucose-induced proinsulin biosynthesis by 30% and to completely inhibit the intracellular conversion of proinsulin to insulin. Autoradiography of monensin-treated cells demonstrated the presence of large quantities of newly synthesized proteins in amorphous vesicles close to the Golgi complex of B cells. The results suggest profound effects of monensin on biosynthesis and intracellular processing of proinsulin.

Animals↗
PubMed1982

Actomyosin interactions with insulin-storage granules in vitro.

Interactions between actomyosin and insulin storage granules isolated from rat islets of Langerhans have been examined in a simple system in vitro, which allows comparison of the sedimentation of the granules in the presence of absence of actomyosin in various conditions. Actomyosin altered granule-sedimentation rates in a manner consistent with the binding of the granules of actomyosin filaments. This interaction was enhanced by addition of ATP (1.5 mM) but unaltered by addition of CaCl2, by calmodulin or by calmodulin in the presence of 10 microM-CaCl2. Addition of EGTA (0.1 mM), cyclic AMP (10 microM) of cytochalasin B (10 microgram/ml) were also without effects in these conditions. Pre-incubation of granules with phospholipase c did not affect granule-actomyosin interaction. Ultrastructural studies showed close contacts between the membranes of the granules and actomyosin filaments. The results indicate the possibility that actomyosin might provide the motile force for granule translocation during the insulin secretory process.

Actomyosin↗
PubMed1982

Effects of taxol and nocodazole on insulin secretion from isolated rat islets of Langerhans.

Taxol, a promotor of microtubule polymerization, and nocodazole, which induces microtubule depolymerization, used at concentrations known to be specific for these effects in other cell types, were each shown to inhibit glucose-stimulated insulin secretion from isolated rat islets of Langerhans. These findings suggest that the dynamic regulation of microtubule polymerization-depolymerization in pancreatic B cells may be important for insulin secretion via the microtubule-microfilamentous system.

Alkaloids↗
PubMed1980

Regulation of actin polymerizaton in rat islets of Langerhans.

A DNAase-inhibition assay was used to determine the proportions of globular (G-) and filamentous (F-) actin in islets of Langerhans after incubation in various conditions, or after subcellular fractionation. Stimulation of insulin secretion resulted in an ATP-dependent increase in the proportion of F-actin present; fractionation showed 80-90% of the actin to be present in the final supernatant.

1-Methyl-3-isobutylxanthine↗
PubMed1980

Problems in the use of polycarbonate diffusion chambers for syngeneic pancreatic islet transplantation in rats.

Islets of Langerhans have been enclosed in polycarbonate diffusion chambers and transplanted intraperitoneally to syngeneic streptozotocin diabetic rats. Direct implantation of 1100--1400 islets in these chambers failed to reverse diabetes during a period of 12 weeks, and viable islet tissue was not recoverable at the end of this period. Islets placed in chambers which had been implanted 3--12 weeks previously similarly failed to lower blood glucose of diabetic recipients, as a result of lack of survival of the islets. Insulin infusion into chambers previously implanted in vivo, I125 insulin diffusion studies in chambers recovered 6--8 weeks after implantation, and scanning electron microscopy of the recovered membranes all indicated that the pores were not totally occluded. The failure of islet transplantation via chambers in this simple syngeneic model has discouraging implications for their use as a means of avoiding allograft rejection.

Animals↗
PubMed1979

Interaction between insulin-storage granules and F-actin in vitro.

Possible interactions between polymerized (F-) actin and insulin-storage granules from rat islets of Langerhans were examined in vitro by comparing the sedimentation of the granules in the presence of various actin concentrations. Actin in the concentration range 0.1--0.5 mg/ml produced a retardation in granule-sedimentation rates consistent with binding of the granules to the actin filaments. The interaction was increased by addition of ATP (2mM), but was decreased by CaCl2 (0.1 mM). Binding of granules to actin was unaffected by cyclic AMP or by preincubation of the granules with phospholipase C. Specificity of the interaction was confirmed by the use of depolymerized (G-) actin and of myosin to provide a solution of comparable viscosity; neither of these caused any alteration of granule sedimentation. Possible implications of this interaction of insulin-storage granules with actin for the mechanism of insulin secretion are briefly discussed.

Actins↗
PubMed1978

Role of microtubules in the intracellular transport of growth hormone.

Pulse-chase experiments utilising (3H)leucine have been used to study the effects of colchicine and vinblastine on intracellular transport and secretion of newly synthesised growth hormone from rat anterior pituitary fragments. Growth hormone was isolated from medium and fragments by polyacrylamide gel electrophoresis. When colchicine or vinblastine, which disrupt microtubules, were added immediately after pulse labelling, inhibition of the subsequent secretion of newly synthesised growth hormone was detected throughout the succeeding 5h. Similar inhibition was seen if the drugs were added after a 1h delay. However, if colchicine or vinblastine were added only after a 2h chase incubation, then no significant effect on subsequent release of labelled growth hormone was seen. The results suggest that these agents may inhibit the transport of newly formed growth hormone storage granules from the Golgi complex to the cytoplasmic pool. Microtubules do not appear to be involved in the mechanism of the final secretion of newly synthesised hormone by exocytosis.

Animals↗
PubMed1977

Secretory capacity and ultrastructure of rat pancreatic islets after preservation of pancreas in different conditions.

Experiments were performed with rat pancreas to investigate optimum conditions for obtaining and preserving the pancreas for subsequent isolation of islets that were viable by both functional and morphological criteria. After only 30 min of warm ischaemia, the yield and viability of islets that could be isolated were poor. However, if the pancreas was removed and placed immediately in a cold bicarbonate-buffered medium, which was supplemented with HEPES to maintain the pH and Trasylol to inhibit proteolytic activity, then viable islets could be isolated consistently after over 8 hr of storage. These results demonstrate that even short periods of warm ischaemia will render the pancreas unsuitable for islet isolation. Once the pancreas is placed in a suitable cold medium, however, considerable delay may be permissible without adversely affecting the viability of the islets which can be obtained. The same conditions might prove to be applicable when human pancreas is obtained for the purpose of transplantation into diabetic subjects.

Animals↗
PubMed1977

Distribution of anionic sites on surface of B cell granule and plasma membranes: a study using cationic ferritin.

The distribution of anionic sites on the membranes of rat pancreatic B cells and of their storage granules has been studied by the use of a visual probe of cationic ferritin. Membranes of isolated storage granules possessed a net negative charge which was apparently evenly distributed; the number of anionic sites was not markedly altered by prior incubation of the granules with neuraminidase or with 10(-5) to 2 X 10(-3) M calcium chloride. Distribution of charges along B cell plasma membranes was less uniform but was similarly unaffected by alterations of calcium concentration, or by neuraminidase treatment. However, during the fusion of plasma membrane and granule membrane which occurs in exocytosis, the emerging granule membrane was found to be devoid of anionic sites. The implications of these findings for the regulation of insulin secretion by exocytosis are discussed.

Animals↗