Cytosolic Ca2+ spikes evoked by the thiol reagent thimerosal in both intact and internally perfused single pancreatic acinar cells.
Cytosolic calcium signals evoked by the sulphydryl-group-oxidising agent, thimerosal, have been investigated in acutely isolated pancreatic acinar cells. Two techniques were employed for the assessment of the cytosolic free-calcium concentration ([Ca2+]i): measurement of calcium-dependent chloride and non-specific cation currents (whole-cell patch-clamp recording) and microfluorimetry (fura-2). Thimerosal (0.5-100 microM) evoked repetitive spikes in both chloride and cation currents as seen by patch-clamp recording, and in [Ca2+]i as seen by microfluorimetry, with a latency of 1-3 min. The response increased in magnitude over time and was not reversed on removal of thimerosal. The thimerosal-induced spikes were reversibly blocked by 2 mM dithiothreitol and by 20 mM caffeine. Inclusion of heparin (200 micrograms/ml) in the pipette solution blocked the thimerosal-induced spikes. The calcium spikes continued after the removal of extracellular calcium; however, low concentrations of thimerosal (0.5-5 microM) were unable to initiate a current response in the absence of external calcium. High concentrations of thimerosal (50-100 microM) could initiate spikes without extracellular calcium. Thimerosal, at concentrations that failed to produce an independent effect, potentiated the acetylcholine-evoked oscillations in [Ca2+]i. We conclude that thimerosal is able to mobilise calcium from an intracellular store; the blockade by heparin may indicate that thimerosal exerts an action on the inositol trisphosphate pathway. The dependence on extracellular calcium for initiation, but not for continuation of the thimerosal-induced calcium spikes suggests that thimerosal may have the additional effect of inhibiting the plasma membrane calcium ATPase.
Thorn
P
P
MRC Secretory Control Research Group, University of Liverpool, UK.
Brady
P
P
Llopis
J
J
Gallacher
D V
DV
Petersen
O H
OH
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D002118:Calcium / Q000502:physiology*; D015220:Calcium Channels; D002469:Cell Separation; D003600:Cytosol / Q000502:physiology*; D004594:Electrophysiology; D006493:Heparin / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D051379:Mice; D010179:Pancreas / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D010477:Perfusion; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors; D018160:Receptors, Cytoplasmic and Nuclear; D013849:Thimerosal / Q000494:pharmacology*
D015220:Calcium Channels; D053496:Inositol 1,4,5-Trisphosphate Receptors; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D013849:Thimerosal; D006493:Heparin; D002118:Calcium