Loperamide mobilizes intracellular Ca2+ stores in insulin-secreting HIT-T15 cells.
1 We have investigated the effects of loperamide on intracellular Ca(2+) stores and membrane K(+) channels in insulin-secreting hamster insulinoma (HIT-T15) cells. 2 In cell-attached patch-clamp mode, loperamide (3-250 micro M) activated large single-channel currents. The loperamide-activated currents were tentatively identified as Ca(2+)-activated K(+) channel (K(Ca)) currents based on their single-channel conductance (145 pS), apparent reversal potential, and insensitivity to tolbutamide. Smaller single-channel currents with a conductance (32 pS) indicative of adenosine triphosphate-sensitive K(+) channels (K(ATP) channels) were also recorded, but were insensitive to loperamide. 3 Surprisingly, the loperamide-activated currents persisted in the absence of extracellular Ca(2+). Yet under these conditions, we still measured loperamide-induced Ca(2+) increases. These effects are dose dependent. Loperamide had no effects in the inside-out patch configuration, suggesting that loperamide does not directly activate the channels with large conductance, but does so secondarily to release of Ca(2+) from intracellular stores. 4 Carbachol (100 micro M), an agonist of muscarinic receptors, which mediates IP(3)-dependent intracellular Ca(2+) release, enhanced the effects of loperamide on K(Ca) channels. 5 Both the putative K(Ca) currents and Ca(2+) signals induced by loperamide (with '0' [Ca(2+)](o)) were abolished when the intracellular Ca(2+) stores had been emptied by pretreating the cells with either carbachol or thapsigargin, an endoplasmic reticulum Ca(2+)-ATPase inhibitor that blocks reuptake of calcium. 6 These data indicate that loperamide in insulin-secreting beta-cells evokes intracellular Ca(2+) release from IP(3)-gated stores and activates membrane currents that appear to be carried by K(Ca), rather than K(ATP) channels.
He
Li-Ping
LP
Department of Pharmacology, Georgetown University, 3900 Reservoir Rd., NW, Washington DC 20057, USA. lipinghe40@hotmail.com
Mears
David
D
Atwater
Illani
I
Rojas
Eduardo
E
Cleemann
Lars
L
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D045744:Cell Line, Tumor; D018999:Charybdotoxin / Q000494:pharmacology; D006224:Cricetinae; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007340:Insulinoma / Q000378:metabolism*; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism; D008139:Loperamide / Q000494:pharmacology*; D018721:Muscarinic Agonists / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D024681:Potassium Channels, Calcium-Activated / Q000037:antagonists & inhibitors / Q000187:drug effects; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism
D015220:Calcium Channels; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007328:Insulin; D018721:Muscarinic Agonists; D024681:Potassium Channels, Calcium-Activated; D018160:Receptors, Cytoplasmic and Nuclear; D018999:Charybdotoxin; D008139:Loperamide; D002217:Carbachol; D002118:Calcium