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Comparison of functional properties of the Ca2+-activated cation channels TRPM4 and TRPM5 from mice
Nina D Ullrich1, Thomas Voets, Jean Prenen
1Laboratorium voor Fysiologie, Department of Physiology, Campus Gasthuisberg, KU Leuven, Herestraat 49, B-3000 Leuven, Belgium.
Abstract:
Non-selective cation (NSC) channels activated by intracellular Ca2+ ([Ca2+]i) play an important role in Ca2+ signaling and membrane excitability in many cell types. TRPM4 and TRPM5, two Ca2+-activated cation channels of the TRP superfamily, are potential molecular correlates of NSC channels. We compared the functional properties of mouse TRPM4 and TRPM5 heterologously expressed in HEK 293 cells. Dialyzing cells with different Ca2+ concentrations revealed a difference in Ca2+ sensitivity between TRPM4 and TRPM5, with EC50 values of 20.2+/-4.0 microM and 0.70+/-0.1 microM, respectively. Similarly, TRPM5 activated at lower Ca2+ concentration than TRPM4 when [Ca2+]i was raised by UV uncaging of the Ca2+-cage DMNP-EDTA. Current amplitudes of TRPM4 and TRPM5 were not correlated to the rate of changes in [Ca2+]i. The Ca2+ sensitivity of both channels was strongly reduced in inside-out patches, resulting in approximately 10-30 times higher EC50 values than under whole-cell conditions. Currents through TRPM4 and TRPM5 deactivated at negative and activated at positive potentials with similar kinetics. Both channels were equally sensitive to block by intracellular spermine. TRPM4 displayed a 10-fold higher affinity for block by flufenamic acid. Importantly, ATP4- blocked TRPM4 with high affinity (IC50 of 0.8+/-0.1 microM), whereas TRPM5 is insensitive to ATP4- at concentrations up to 1 mM.
Insights
TRPM4 and TRPM5 are calcium-activated cation channels. TRPM5 activates at lower calcium levels than TRPM4, and they differ in ATP sensitivity.
Area of Science:
- Molecular Biology
- Cell Physiology
- Ion Channel Research
Background:
- Non-selective cation (NSC) channels are crucial for calcium (Ca2+) signaling and cellular membrane excitability.
- Transient Receptor Potential Melastatin (TRPM) channels, specifically TRPM4 and TRPM5, are implicated as key Ca2+-activated cation channels.
Purpose of the Study:
- To compare the functional properties of mouse TRPM4 and TRPM5 channels.
- To elucidate differences in Ca2+ sensitivity and pharmacological profiles between TRPM4 and TRPM5.
Main Methods:
- Heterologous expression of mouse TRPM4 and TRPM5 in HEK 293 cells.
- Whole-cell patch-clamp recordings with varying intracellular Ca2+ concentrations.
- UV uncaging of Ca2+ using DMNP-EDTA.
- Inside-out patch-clamp recordings.
- Assessment of channel block by spermine, flufenamic acid, and ATP4-.
Main Results:
- TRPM5 exhibited significantly higher Ca2+ sensitivity (EC50 ~0.70 µM) compared to TRPM4 (EC50 ~20.2 µM).
- Ca2+ sensitivity was reduced in inside-out patches for both channels.
- TRPM4 showed higher affinity for flufenamic acid block and was potently blocked by ATP4- (IC50 ~0.8 µM), while TRPM5 was insensitive to ATP4-.
Conclusions:
- TRPM4 and TRPM5 possess distinct Ca2+ activation thresholds and pharmacological properties.
- TRPM4's sensitivity to ATP4- differentiates it from TRPM5, suggesting distinct physiological roles.
- These findings contribute to understanding the specific functions of TRPM4 and TRPM5 in Ca2+ signaling pathways.
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