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This Article Full Text Full Text (PDF) All Versions of this Article: 297/4/G687    most recent 90683.2008v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Mishra, R. Articles by Hill, C. E. PubMed PubMed Citation Articles by Mishra, R. Articles by Hill, C. E.

LIVER AND BILIARY TRACT

Mg²⁺- and MgATP-inhibited and Ca²⁺/calmodulin-sensitive TRPM7-like current in hepatoma and hepatocytes

Departments of Biology and Medicine, Hotel Dieu Hospital and Queen's University, Kingston, Ontario, Canada

Submitted December 1, 2008 ; accepted in final form August 4, 2009

Although understood to be ubiquitously expressed, the functional identification and significance of Mg²⁺-inhibited, nonspecific cation currents has been established in only a few cell types. Here we identified an outwardly rectifying nonspecific cation current in quiescent rat hepatocytes and the proliferating and polarized rat hepatoma, WIF-B. Under whole cell recording conditions in which cells were bathed and dialyzed with Na-gluconate solutions, the latter Ca²⁺ and Mg²⁺ free, current reversed close to 0 mV, was time independent, and was greater than 10 times higher at +120 mV compared with –120 mV. Outward current at –120 mV developed slowly, from 17.7 ± 10.3 pA/pF at patch rupture to 106.6 ± 15.6 pA/pF at 12 min in WIF-B cells, and 4.9 ± 2.7 to 20.6 ± 5.6 pA/pF in rat hepatocytes. The nonspecific TRP channel inhibitor, 2-aminoethoxyphenylborate (2-APB), inhibited current (IC₅₀ = 72 ± 13 µM) and caused apoptotic cell death in WIF-B cells. Rat hepatocyte survival was more resistant to 2-APB. Dialysis of WIF-B cells with physiological concentrations of Mg²⁺ and Mg-ATP, but not ATP alone, inhibited current development, suggesting that Trpm7 rather than Trpm6 underlies this current. RT-PCR demonstrated that both Trpm6 and Trpm7 are expressed at similar levels in both cell types, suggesting that the functional differences noted are not transcript dependent. Intracellular Ca²⁺ (IC₅₀ = 125 ± 35 nM) also inhibited current development, and this could be partially relieved by the calmodulin and Ca²⁺/calmodulin-dependent kinase inhibitors W-7, staurosporine, KN-93, or calmodulin kinase II (CaMKII) inhibitory peptide. To summarize, our results show that in addition to their established Mg²⁺ sensitivity, Trpm7-like channels are inhibited by cytosolic Ca²⁺ in a CaMKII-dependent manner and may support hepatocellular survival during proliferation.

channel; liver; survival; kinase