Protein kinase C activates capacitative calcium entry in the insulin secreting cell line RINm5F

Abstract

This study examines the calcium store-regulated (capacitative) calcium influx pathway in the endocrine pancreatic cell line RINmSF, utilizing thapsigargin. After preincubation of the cells with the phorbol ester TPA, thapsigargin induced a sustained elevation of cytosolic calcium as well as a sustained stimulation of manganese entry, the latter being used to assess calcium influx. Thapsigargin given alone provoked a smaller and only transient elevation of cytosolic calcium and stimulation of manganese entry. The protein kinase C inhibitor staurosporine antagonized the effect of the phorbol ester. Verapamil, nifedipine, or measures to hyperpolarize the cells exerted no inhibitory actlon against this effect, which excludes an involvement of voltage-dependent calcium channels. In conclusion, our data shows for the first time that protein kinase C stimulation activates the capacitative calcium influx pathway of endocrine pancreatic insulin-producing cells.

Molecular cloning, functional expression, and signal transduction of the GIP‐receptor cloned from a human insulinoma

“…Loading of cells (48 h after transfection) with the fluorescent calcium indicator fura-2 was performed as described [14,15]. Measurements of the [CaZ+]cyt concentration and calibration of the data were performed essentially as described previously [14,15], using a modified KrebsRinger bicarbonate buffer containing 4 mM glucose and 1.5 mM CaC12.…”

“…Measurements of the [CaZ+]cyt concentration and calibration of the data were performed essentially as described previously [14,15], using a modified KrebsRinger bicarbonate buffer containing 4 mM glucose and 1.5 mM CaC12. The measurements were performed at 37°C.…”

“…The measurements were performed at 37°C. Since CHL cells displayed dye efflux during the measurements, we corrected the data as described previously [14] using either addition of manganese and the manganese chelator diethylenetrianminepenta acetic acid (DTPA) in the beginning and at the end of each measurement, or performing the measurements in the presence of 250/IM sulfinpyrazone and adding manganese and DTPA.…”

Protein Kinase C Activates Store-operated Ca2+Channels in Human Glomerular Mesangial Cells

“…Depending on the specifically tested cell and the experimental conditions (8,(22)(23)(24), variable results have been reported on the modulation of SOC by PKC. For example, PKC decreases Ca 2ϩ influx through SOC in Xenopus oocytes (8) and thyroid FRTL-5 cells (24) but activates SOC in RINm5F cells (22), mouse pancreatic acinar cells (23), bovine adrenocortical cells (25), and NIH 3T3 cells with submaximal depletion of internal stores (26).…”

“…For example, PKC decreases Ca 2ϩ influx through SOC in Xenopus oocytes (8) and thyroid FRTL-5 cells (24) but activates SOC in RINm5F cells (22), mouse pancreatic acinar cells (23), bovine adrenocortical cells (25), and NIH 3T3 cells with submaximal depletion of internal stores (26). In many of these studies, Ca 2ϩ -sensitive fluorescence was used to evaluate Ca 2ϩ influx in response to depleting internal Ca 2ϩ stores.…”