Article date: May 2003
By: Li‐Ping He, David Mears, Illani Atwater, Eduardo Rojas, Lars Cleemann in Volume 139, Issue 2, pages 351-361
We have investigated the effects of loperamide on intracellular Ca2+ stores and membrane K+ channels in insulin‐secreting hamster insulinoma (HIT‐T15) cells.
In cell‐attached patch‐clamp mode, loperamide (3–250 μM) activated large single‐channel currents. The loperamide‐activated currents were tentatively identified as Ca2+‐activated K+ channel (KCa) 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 (KATP channels) were also recorded, but were insensitive to loperamide.
Surprisingly, the loperamide‐activated currents persisted in the absence of extracellular Ca2+. Yet under these conditions, we still measured loperamide‐induced Ca2+ 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 Ca2+ from intracellular stores.
Carbachol (100 μM), an agonist of muscarinic receptors, which mediates IP3‐dependent intracellular Ca2+ release, enhanced the effects of loperamide on KCa channels.
Both the putative KCa currents and Ca2+ signals induced by loperamide (with ‘0’ [Ca2+]o) were abolished when the intracellular Ca2+ stores had been emptied by pretreating the cells with either carbachol or thapsigargin, an endoplasmic reticulum Ca2+‐ATPase inhibitor that blocks reuptake of calcium.
These data indicate that loperamide in insulin‐secreting β‐cells evokes intracellular Ca2+ release from IP3‐gated stores and activates membrane currents that appear to be carried by KCa, rather than KATP channels.
We have investigated the effects of loperamide on intracellular Ca2+ stores and membrane K+ channels in insulin‐secreting hamster insulinoma (HIT‐T15) cells.
In cell‐attached patch‐clamp mode, loperamide (3–250 μM) activated large single‐channel currents. The loperamide‐activated currents were tentatively identified as Ca2+‐activated K+ channel (KCa) 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 (KATP channels) were also recorded, but were insensitive to loperamide.
Surprisingly, the loperamide‐activated currents persisted in the absence of extracellular Ca2+. Yet under these conditions, we still measured loperamide‐induced Ca2+ 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 Ca2+ from intracellular stores.
Carbachol (100 μM), an agonist of muscarinic receptors, which mediates IP3‐dependent intracellular Ca2+ release, enhanced the effects of loperamide on KCa channels.
Both the putative KCa currents and Ca2+ signals induced by loperamide (with ‘0’ [Ca2+]o) were abolished when the intracellular Ca2+ stores had been emptied by pretreating the cells with either carbachol or thapsigargin, an endoplasmic reticulum Ca2+‐ATPase inhibitor that blocks reuptake of calcium.
These data indicate that loperamide in insulin‐secreting β‐cells evokes intracellular Ca2+ release from IP3‐gated stores and activates membrane currents that appear to be carried by KCa, rather than KATP channels.
British Journal of Pharmacology (2003) 139, 351–361. doi:10.1038/sj.bjp.0705263
DOI: 10.1038/sj.bjp.0705263
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