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Biomedical subjects

D E Cochrane

Publications and source records attributed to D E Cochrane.

45 records · Page 3Linked to original sources
PubMed1975

Calcium and stimulus-secretion coupling in the adrenal medulla: contrasting stimulating effects of the ionophores X-537A and A23187 on catecholamine output.

1. The ionophores X-537A and A23187, which are known to transfer calcuim across cell membranes, stimulated catecholamine release from perfused cat adrenal glands. 2. These stimulant effects persisted in the presence of hexamethonium and atropine and are therefore attributable to direct actions of the ionophores on the adrenal chromaffin cells. 3. Perfusion with calcium-free Locke abolished responses to A23187 and reduced those to X-537A. 4. Responses to X-537A were consistently large and comparable with those produced by 56 mM potassium. By contrast, responses to A23177, over the wide range of concentrations tested, were variable and much smaller. 5. That the two ionophores can stimulate through calcium-dependent mechanisms is considered fresh support for the calcium hypothesis of stimulus-secretion coupling. That they differ in effectiveness may mean that factors besides calcium are important. The greater potency of the less specific ionophore, X-537A, may be attributable to its ability to depolarize as well as carry calcuim, while the relatively small effects of A23187, a generally more effective ionophore for calcuim, may indicate that inward movement of calcium without a background of membrane perturbation such as may be produced by depolarization, is insufficient to elicit strong secretory responses.

Adrenal Medulla↗
PubMed1974

Calcium-induced extrusion of secretory granules (exocytosis) in mast cells exposed to 48-80 or the ionophores A-23187 and X-537A.

Isolated peritoneal mast cells from rats were observed, by phase contrast microscopy, to extrude secretory granules when exposed to 48/80 or the ionophores A-23187 and X-537A, which are known to facilitate transmembrane fluxes of calcium. These effects were abolished when the cells were treated with EDTA and suspended in a Ca-free environment. Ca-deprived cells exposed to any one of the three drugs promptly extruded granules when calcium, but not magnesium, was added to the incubation medium. Such Ca-evoked or Ca-dependent responses persisted when Na was omitted from the incubation medium and replaced with sucrose, choline, or K. The responses thus seem independent of possible shifts in the alkali metal ions. The results are considered support for the view that Ca influx mediates stimulus-secretion coupling and does so by initiating exocytosis.

Animals↗
PubMed1972

The interaction between caffeine and calcium in the desensitization of muscle postjunctional membrane receptors.

The interaction between caffeine and calcium on the rate of desensitization of muscle postjunctional membrane (PJM) receptors during the sustained application of 0.27 mM carbamylcholine (CARB) has been studied in vitro on the sartorius muscle of the frog. The rate of PJM repolarization with CARB added to the solution bathing the muscle or the recovery of the effective transmembrane resistance (EMR) during the microperfusion of CARB directly onto the end-plate region of individual fibers was used as an index of the rate of desensitization. Caffeine (1.5 mM) increased the rate of PJM repolarization with bulk application of CARB in a 1.8 or 10 mM calcium Ringer solution but had no effect on PJM repolarization in a calcium-deficient, 4 mM magnesium Ringer solution. For EMR measurements the preparation was rendered mechanically quiescent by repeated challenges with isotonic KCl during an exposure of several hours to a calcium-free, 4 mM magnesium-1 mM EGTA Ringer solution. In these fibers, the microperfusion of 0.27 mM CARB together with 1.8 mM calcium plus 1.5 mM caffeine significantly increased the rate of EMR recovery above that observed in the absence of caffeine. Raising the calcium concentration to 10 mM had a similar effect; however, no additional increase was noted by the inclusion of 1.5 mM caffeine. It is suggested that the major role of caffeine in PJM desensitization is to increase the calcium permeability of the surface membrane. The transmembrane movement of calcium and the consequent intracellular accumulation of calcium is seen as a critical factor in controlling the rate of PJM desensitization.

Animals↗
PubMedDate not supplied

Neurotensin elevates hematocrit and plasma levels of the leukotrienes, LTB4, LTC4, LTD4 and LTE4, in anesthetized rats.

The IV injection of neurotensin (NT) into anesthetized rats produced a marked increase in hematocrit, labored breathing and peripheral blood stasis with cyanosis. This effect could also be produced by the NT-related peptides, neuromedin-N and xenopsin; however, it was not observed when nine other biologically active peptides, including bradykinin and substance P, were tested. Associated with these responses were increases in the plasma levels of histamine (measured radioenzymatically) and the leukotrienes, LTB4, LTC4, LTD4, and LTE4 (measured by RIA and HPLC). The increment in hematocrit after varying doses of NT correlated to the increase in plasma levels of LTC4. Histamine and LTC4 were both capable of elevating hematocrit when given IV; however, LTC4 was approximately 1000 times more potent than histamine and active doses of histamine elevated LTC4 levels. Furthermore, the effects of NT on plasma LTC4 and hematocrit were reduced by pretreating animals with antagonists to histamine and serotonin. Pretreatment with the specific mast cell degranulating agent, compound 48/80, also blocked NT's ability to elevate plasma levels of histamine, LTB4 and LTC4 and prevented the increased hematocrit and cyanosis. These results indicate that NT-related peptides are very potent and specific stimulators of leukotriene release and that this action is mediated by mast cells and associated with loss of plasma volume and blood stasis. A working hypothesis is that histamine, released from mast cells in response to NT, stimulates LTC4 production by other cells.

Anesthesia, General↗
PubMedDate not supplied

Rapid degradation of neurotensin by stimulated rat mast cells.

A RIA towards neurotensin (NT) using C-terminal- and N-terminal-specific antisera was used to study degradation of this tridecapeptide by isolated rat mast cells. Incubation of NT (10 microM) with peritoneal or pleural mast cells resulted in a rapid loss of NT immunoreactivity (iNT), as measured by C-terminal-directed antiserum, with little effect on N-terminal iNT. The rate of the reaction was faster with pleural cells (T1/2, 30 s) than with peritoneal cells (T1/2, 180 s) and was greater than 10-fold slower in the presence of metabolic poisons. The enzyme(s) involved is most likely released from the cells during secretion, as NT was degraded by media conditioned by compound 48/80-stimulated mast cells 40-60 times faster than by media from unstimulated cells. This degradation by conditioned media was concentration dependent, pH dependent, and temperature sensitive. HPLC analyses indicated a near stoichiometric conversion of NT to NT(1-12) (66%) and NT(1-11) (34%) after incubation for 10-30 s with conditioned media. By 30 min only NT(1-11) and NT(1-10) were present. Phenanthroline (1 mM), an inhibitor of carboxypeptidase, prevented the loss of C-terminal iNT and the generation of NT(1-12) and NT(1-11). While NT(1-12) was effective in releasing histamine from mast cells in vitro and increasing vascular permeability in vivo, NT(1-11) was not. These results suggest that carboxypeptidase-like enzyme(s) could modulate the level and form of NT-related peptides in various states involving activation of mast cells.

Amino Acid Sequence↗
PubMedDate not supplied

Stimulated rat mast cells generate histamine-releasing peptide from albumin.

Media conditioned by compound 48/80-stimulated rat mast cells generated immunoreactive histamine-releasing peptide (HRP) when incubated at physiological pH with bovine serum albumin and the carboxypeptidase inhibitor, O-phenanthroline. The generation of immunoreactive HRP (IR-HRP) was time (after 3 h the concentration of IR-HRP was 20 nM), temperature, and pH dependent and was prevented by omitting albumin, by using media conditioned by nonstimulated mast cells, or by pretreatment of mast cells with disodium cromoglycate, an inhibitor of mast cell secretion. The amount of IR-HRP generated increased linearly with the number of mast cells stimulated and varied directly with the concentration of conditioned media. After removal of the media from stimulated mast cells, the remaining cell pellet retained its ability to generate IR-HRP for up to 8 h. Stimulation of mast cells by either neurotensin or substance P, or of sensitized cells by anti-IgE serum, also produced conditioned media that generated IR-HRP. The amount of IR-HRP formed by various conditioned media or by stimulated cell pellets was dependent upon the concentration of O-phenanthroline used. Including the chymase inhibitor, chymostatin, prevented the formation of IR-HRP in a dose-dependent manner. HPLC analysis showed four peaks of IR-HRP. The major one coeluted with synthetic HRP. These results indicate that the peptide, HRP, can be generated by stimulated mast cells incubated in the presence of albumin. They suggest that a chymase-like enzyme secreted by the mast cell is able to cleave albumin to yield HRP.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗