Author(s): Patricia Santofimia-CastaÃÂ±o, Andreas Schmid, Ines Anderie, Miguel FernÃÂ¡ndez-Bermejo, GinÃÂ©s M. Salido, and Antonio GonzÃÂ¡lez
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.
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