The Zn2+-transporting Pathways in Pancreatic β-Cells

Abstract

In pancreatic ␤-cells

Investigation of Transport Mechanisms and Regulation of Intracellular Zn2+ in Pancreatic α-Cells

“…The expression of Zip5 mRNA in the whole mouse pancreas was also observed (73,74). Our previous work using quantitative real time PCR demonstrates a number of Zip transcripts in mouse insulinoma ␤-cells (MIN6), mouse islets, and mouse brain (32). In this work we have identified in glucagon-producing ␣-TC cells the expression of a number of genes of the ZIP and ZnT families (Fig.…”

“…Fluorescent Visualization of Zn 2ϩ Accumulation in ␣-TC Cells-Our previous results showed the partial co-localization of FluoZin-3 and a mitochondrial marker in ␤-cells (32). In this study ␣-TC cells stained simultaneously with FluoZin-3 AM and the mitochondrial marker dye MitoTracker Red (Fig.…”

“…MIP-GFP mice were a gift from Dr. M. Hara, University of Chicago, Chicago, IL (41). Islets were isolated as described previously (32). To obtain dispersed cells, isolated islets were incubated for 10 min in Ca 2ϩ -free phosphate-buffered solution supplemented with 2 mM EGTA, 3 mM glucose, 100 units/ml penicillin, and 100 g/ml streptomycin.…”

“…For excitation, a xenon lampbased DeltaRam high speed monochromator from Photon Technology International (PTI, Lawrenceville, NJ) was used. For control of the monochromator and video camera, as well as for analysis of fluorescent imaging, ImageMaster 3 software (PTI) was used as we have described previously (32 measurements, coverslips with ␣-TC cells, or dispersed islet cells, were loaded with 2 M FluoZin-3 AM for 50 min in incubation buffer in the presence of 2 mM glucose at 37°C and 5% CO 2 , 95% air. Then coverslips with ␣-TC cells or islet cells were subsequently incubated for 10 min in the same buffer without dye.…”

“…The resulting cDNA was used for amplification in quantitative real time PCR (qPCR). qPCR was performed as described previously (32). Primers were designed using Primer Express version 2.0 software (Applied Biosystems, Foster City, CA).…”