Melatonin modulates Ca 2+ mobilization and amylase release in response to cholecystokinin octapeptide in mouse pancreati
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ORIGINAL PAPER
Melatonin modulates Ca2+ mobilization and amylase release in response to cholecystokinin octapeptide in mouse pancreatic acinar cells Patricia Santofimia-Castaño & Deborah Clea Ruy & Ginés M. Salido & Antonio González
Received: 18 March 2013 / Accepted: 12 June 2013 # University of Navarra 2013
Abstract In the present work, we have evaluated the effect of an acute addition of melatonin on cholecystokinin octapeptide (CCK-8)-evoked Ca2+ signals and amylase secretion in mouse pancreatic acinar cells. For this purpose, freshly isolated mouse pancreatic acinar cells were loaded with fura-2 to study intracellular free Ca2+ concentration ([Ca2+]c). Amylase release and cell viability were studied employing colorimetric methods. Our results show that CCK-8 evoked a biphasic effect on amylase secretion, finding a maximum at a concentration of 0.1 nM and a reduction of secretion at higher concentrations. Preincubation of cells with melatonin (1 μM–1 mM) significantly attenuated enzyme secretion in response to high concentrations of CCK-8. Stimulation of cells with 1 nM CCK-8 led to a transient increase in [Ca2+]c, followed by a decrease towards a constant level. In the presence of 1 mM melatonin, stimulation of cells with CCK-8 resulted in a smaller [Ca2+]c peak response, a faster rate of decay of [Ca2+]c and lower values for the steady state of [Ca2+]c, compared with the effect of CCK-8 alone. Melatonin also reduced the oscillatory pattern of Ca2+ mobilization P. Santofimia-Castaño : G. M. Salido : A. González (*) Department of Physiology (Cell Physiology Research Group), University of Extremadura, Avenida Universidad s/n, 10003 Caceres, Spain e-mail: [email protected] D. C. Ruy Facultade de Agronomia & Medicina Veterinaria, Universidade de Brasilia, 70900-100 Brasilia, Federal District, Brazil
evoked by a physiological concentration of CCK8 (20 pM), and completely inhibited Ca2+ mobilization induced by 10 pM CCK-8. On the other hand, Ca2+ entry from the extracellular space was not affected in the presence of melatonin. Finally, melatonin alone did not change cell viability. We conclude that melatonin, at concentrations higher than those found in blood, might regulate exocrine pancreatic function via modulation of Ca2+ signals. Keywords Pancreas . Calcium . Amylase secretion . Cholecystokinin . Melatonin . Cell viability Abbreviations [Ca2+]c Cytosolic free Ca2+ concentration CCK-8 Cholecystokinin octapeptide EGTA Ethylene glycol-bis (2-aminoethylether)-N,N,N′N′tetraacetic acid ER Endoplasmic reticulum Fura-2/AM Fura-2 acetoxymethyl ester
Introduction The exocrine pancreas is highly specialized for the synthesis, storage and exocrine secretion of digestive enzymes. Exocrine pancreatic secretion is activated by nutrients, and is subject to neurohormonal regulation and to interdigestive (resting) periods [19, 32, 33, 35]. A rise in
P. Santofimia-Castaño et al.
cytosolic free Ca2+ concentration ([Ca2+]c) is an important early signal by which physiological secretagogues like cholecystokinin, acting in a co
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