Change in the Characteristics of Ca2+ Signaling in Pancreatic Acinar Cells in Culture

Patricia Santofimia-Castaño,Antonio González,Ginés M Salido,Ines Anderie,Andreas Schmid,Miguel Fernández-Bermejo

doi:10.11131/2014/101058

Abstract

In this study we studied Ca2+ responses to different stimuli in pancreatic acinar cells subjected to culture conditions. Cells were isolated from adult mice pancreas and were subjected to culture conditions along one week. Changes in intracellular free Ca2+ concentration were monitored by single cell fluorescence analysis of fura-2-loaded cells. Mitochondrial distribution was analyzed by confocal microscopy study of MitoTracker Green FM-loaded cells. Expression of amylase-containing cytoplasmic vesicles was analyzed by confocal microscopy study of cells transfected with a plasmid encoding amylase linked to a green fluorescent protein. Cell viability was analyzed employing the AlamarBlue test. Our results show that pancreatic cells in culture retain a mitochondrial network and amylase-positive vesicles. However, cells dropped their ability to mobilize Ca2+ in response to activation of cell membrane receptors. Ca2+ mobilization in response to the sarcoendoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin was not altered. Cell viability was not affected by treatment with cholecystokinin, but was reduced in the presence of thapsigargin or hydrogen peroxide. We conclude that primary culture of pancreatic cells may be a suitable model to be used in studies where the involvement of mechanisms linked to the activation of specific cell membrane receptors is not required.

Highlights

The main function of the exocrine pancreas is the secretion of digestive enzymes, conveyed in Cl− rich fluid
In order to get a thorough knowledge of the physiology of the exocrine pancreas, and due to the importance of the pathological processes affecting this tissue, an array of studies have been performed employing freshly isolated pancreatic acinar cells or cell lines in culture, mostly tumoral
In the present study we have evaluated whether primary cultures of pancreatic acinar cells, obtained from adult mice, maintain the characteristics of Ca2+ signaling in response to several stimuli, to assess whether they may be a suitable model to study pancreatic physiology

Summary

Introduction

The main function of the exocrine pancreas is the secretion of digestive enzymes, conveyed in Cl− rich fluid. This process occurs an alkaline HCO−3 and via mechanisms which depend on the generation of Ca2+ signals in response to secretagogues [1]. In vitro models of exocrine pancreas are useful for the study of Ca2+ signaling, pancreatic differentiation, secretion mechanisms or cell injury for example. In order to get a thorough knowledge of the physiology of the exocrine pancreas, and due to the importance of the pathological processes affecting this tissue (inflammation, cancer, etc.), an array of studies have been performed employing freshly isolated pancreatic acinar cells or cell lines in culture, mostly tumoral. It would be interesting tofind a cellular

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