Maitotoxin activates an endogenous non-selective cation channel and is an effective initiator of the activation of the heterologously expressed hTRPC-1 (transient receptor potential) non-selective cation channel in H4-IIE liver cells

The structures and mechanisms of activation of non-selective cation channel s (NSCCs) are not well understood although NSCCs play important roles in th e regulation of metabolism, ion transport, cell volume and cell shape. It h as been proposed that TRP (transient receptor potential) proteins are the m olecular correlates of some NSCCs. Using fura-2 and patch-clamp recording, it was shown that the maitotoxin-activated cation channels in the H4-IIE ra t liver cell line admit Ca2+, Mn2+ and Na+, have a high selectivity for Na compared with Ca2+, and are inhibited by Gd3+ (half-maximal inhibition at 1 muM). Activation of the channels by maitotoxin was inhibited by increasin g the extracellular Ca2(+) concentration or by inclusion of 10 mM EGTA in t he patch pipette. mRNA encoding TRP proteins 1, 2 and 3 at levels comparabl e with those in brain was detected using reverse transcriptase-polymerase c hain reaction in poly(A)(+) RNA prepared from H4-IIE cells and freshly-isol ated rat hepatocytes. In H4-IIE cells transiently transfected with cDNA enc oding hTRPC-1, the expressed hTRPC-1 protein was chiefly located at intrace llular sites and at the plasma membrane. Cells expressing hTRPC-1 exhibited a substantial enhancement of maitotoxin-initiated Ca2+ inflow and a modest enhancement of thapsigargin-initiated Ca2+ inflow (measured using fura-2) and no enhancement of the highly Ca2+-selective store-operated Ca2+ current (measured using patch-clamp recording). In cells expressing hTRPC-1, maito toxin activated channels which were not found in untransfected cells, have an approximately equal selectivity for Na+ and Ca2+, and are inhibited by G d3+ (half-maximal inhibition at 3 LM). It is concluded that in liver cells (i) maitotoxin initiates the activation of endogenous NSCCs with a high sel ectivity for Na+ compared with Ca2+; (ii) TRP proteins 1, 2 and 3 are expre ssed; (iii) maitotoxin is an effective initiator of activation of heterolog ously expressed hTRPC-1 channels; and (iv) the endogenous TRP-1 protein is unlikely to be the molecular counterpart of the maitotoxin-activated NSCCs nor the highly Ca2+-selective store-operated Ca2+ channels. (C) 2001 Elsevi er Science B.V. All rights reserved.