Regulatory volume decrease (RVD) in α-cells isolated from rat pancreatic islets was examined using video-imaging methods. Rats were humanely killed by stunning and cervical dislocation. Pancreatic islets were isolated by collagenase digestion, and the islets dispersed into single cells in Ca2+-free medium (Miley et al. 1997). α-Cells were selected from the islet cell population on the basis of volume, i.e. < 0.8 pl (see Majid et al. 2001).
Cells were bathed in isotonic, Hepes-buffered solutions (302 mosmol (kg H2O)-1). On exposure to hypotonic solutions (197 mosmol (kg H2O)-1 by removal of NaCl), relative cell volume increased to a maximum of 1.30 ± 0.04 (mean ± S.E.M.) in nine α-cells. Cell volume then decreased over the remainder of the 15 min exposure to the hypotonic solution, i.e. they exhibited a RVD. The volume recovery (maximum volume – minimum volume) was 0.21 ± 0.02 in control conditions. The RVD in α-cells was inhibited by 10 mM R-(+)-([2-n-Butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro1-oxo-1H-inden-5-yl]oxy)acetic acid (DIOA; a K+-Cl– cotransporter inhibitor, Shen et al. 2000). The volume recovery in the presence of DIOA was 0.12 ± 0.03 (n = 6; significantly less than control by Student’s t test for unpaired data, P < 0.05). The RVD in pancreatic β-cells, which involves K+ and Cl– channels, was not affected by 10 mM DIOA (P > 0.1). These data suggest that K+-Cl– cotransporters contribute to the RVD in pancreatic α-cells.
The relative volume of eight α-cells increased from 0.49 ± 0.08 pl in control conditions, to a maximum of 0.52 ± 0.03 pl (n = 8) when they were exposed to isotonic solutions containing 10 mM DIOA for 15 min (P < 0.05 by paired t test). Cell volume did not change in time-matched isotonic control experiments (P > 0.1). These data suggest that K+-Cl– cotransporters may be active in α-cells at normal cell volumes in isotonic solutions.
In conclusion, the data are consistent with the expression of K+-Cl– cotransporters in pancreatic α-cells.
All procedures accord with current UK legislation.