Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, PC313

Poster Communications

Modulation of TRPM3 cation channel by intacellular Ca2+

V. A. Seymour1, K. Greenhalgh1, J. Naylor1, D. J. Beech1

1. IMSB, University of Leeds, Leeds, West Yorkshire, United Kingdom.


TRPM3 is a non-selective cation channel that is widely expressed throughout the body and it has been implicated in the regulation of vascular smooth muscle cell function (Naylor et al, 2010) and insulin secretion (Wagner et al, 2008). However, its regulation remains incompletely understood. Other members of the melastatin subgroup of TRP channels such as TRPM2, TRPM4b and TRPM5 are strongly activated by intracellular Ca2+. Therefore we tested the hypothesis that TRPM3 is also a Ca2+ activated ion channel. The effect of Ca2+ on TRPM3 function was investigated using a calcium indicator dye (fluo4) and the whole-cell patch-clamp technique. HEK 293 cells stably expressing human TRPM31325 and the TRPM3 agonist pregnenolone sulphate (PregS) were used in these experiments. Data are shown as mean ± s.e.m. TRPM3 expressing cells were treated with thapsigargin to elevate intracellular Ca2+. These cells displayed an increase in basal intracellular calcium levels and their response to PregS was decreased by approximately 20 % (n=3). This result indicates that TRPM3 is inhibited by increases in intracellular Ca2+. Patch-clamp experiments however, revealed that Ca2+ has a more complex effect on TRPM3. Under conditions of low Ca2+ buffering (0.1 mM EGTA in the pipette solution) two effects of Ca2+ were observed. In 4 out of 9 whole-cell patches where the extracellular solution was switched from a Ba2+- to a Ca2+-containing solution the PregS induced inward and outward currents were significantly potentiated (2.9 ± 0.3 and 1.9 ± 0.1 times respectively). In 4 of the remaining patches application of Ca2+ caused a significant inhibition of the inward and outward PregS induced currents (44 ± 4 and 77 ± 7 % respectively). All of these effects were lost under high Ca2+ buffering conditions (10 mM EGTA in the pipette solution) suggesting they arose due to changes in intracellular Ca2+. The data suggest that TRPM3 is both stimulated and inhibited by intracellular Ca2+. Experiments are ongoing to reveal the underlying mechanisms and physiological relevance.

Where applicable, experiments conform with Society ethical requirements