Evidence for a KATP Channel in Rough Endoplasmic Reticulum (rerKATP Channel) of Rat Hepatocytes.

Sajjad Salari,Remy Sauve,Maedeh Ghasemi,Reza Saghiri,Afsaneh Eliassi,Javad Fahanik-Babaei,Agustin Guerrero-Hernandez

doi:10.1371/journal.pone.0125798

Abstract

We report in a previous study the presence of a large conductance K+ channel in the membrane of rough endoplasmic reticulum (RER) from rat hepatocytes incorporated into lipid bilayers. Channel activity in this case was found to decrease in presence of ATP 100 µM on the cytoplasmic side and was totally inhibited at ATP concentrations greater than 0.25 mM. Although such features would be compatible with the presence of a KATP channel in the RER, recent data obtained from a brain mitochondrial inner membrane preparation have provided evidence for a Maxi-K channel which could also be blocked by ATP within the mM concentration range. A series of channel incorporation experiments was thus undertaken to determine if the ATP-sensitive channel originally observed in the RER corresponds to KATP channel. Our results indicate that the gating and permeation properties of this channel are unaffected by the addition of 800 nM charybdotoxin and 1 µM iberiotoxin, but appeared sensitive to 10 mM TEA and 2.5 mM ATP. Furthermore, adding 100 µM glibenclamide at positive potentials and 400 µM tolbutamide at negative or positive voltages caused a strong inhibition of channel activity. Finally Western blot analyses provided evidence for Kir6.2, SUR1 and/or SUR2B, and SUR2A expression in our RER fractions. It was concluded on the basis of these observations that the channel previously characterized in RER membranes corresponds to KATP, suggesting that opening of this channel may enhance Ca2+ releases, alter the dynamics of the Ca2+ transient and prevent accumulation of Ca2+ in the ER during Ca2+ overload.

Highlights

IntroductionIn diaphragm sarcoplasmic reticulum (SR) the channel conductance and gating properties were not affected by physiological concentrations of Ca2+, Mg2+, and the channel appeared insensitive to glyburide (a selective blocker of ATP-sensitive K+ channels), and charybdotoxin (ChTx) (blocker of Ca2+-activated K+ channels) [5]
The nature of the large conductance K+ channel we previously identified in the rough endoplasmic reticulum (RER) membrane of hepatocyte [8] was investigated in a series of experiments aimed first to characterize the channel pharmacological profile, and second to establish if the KATP Kir6.x and sulfonylurea receptors (SURs) subunits were present in our RER membrane preparation
Both Maxi-KCa and KATP channels are sensitive to ATP, we considered the possible effect of Ca2+ ions and Maxi-KCa blockers on the K+ channel from RER membranes

Summary

Introduction

In diaphragm SR the channel conductance and gating properties were not affected by physiological concentrations of Ca2+, Mg2+, and the channel appeared insensitive to glyburide (a selective blocker of ATP-sensitive K+ channels), and charybdotoxin (ChTx) (blocker of Ca2+-activated K+ channels) [5]. It was proposed on the basis of these observations that SR K+ channels could serve in maintaining the Ca2+ homeostasis [5, 7]

Methods

Results

Conclusion