Biochemical, single-channel, whole-cell patch clamp, and pharmacological analyses of endogenous TRPM4 channels in HEK293 cells

Amarouch, Mohamed Yassine; Syam, Ninda Ratna Maharani; Abriel, Hugues (2013). Biochemical, single-channel, whole-cell patch clamp, and pharmacological analyses of endogenous TRPM4 channels in HEK293 cells. Neuroscience letters, 541, pp. 105-110. Elsevier 10.1016/j.neulet.2013.02.011

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Human embryonic kidney cells 293 (HEK293) are widely used as cellular heterologous expression systems to study transfected ion channels. This work characterizes the endogenous expression of TRPM4 channels in HEK293 cells. TRPM4 is an intracellular Ca(2+)-activated non-selective cationic channel expressed in many cell types. Western blot analyses have revealed the endogenous expression of TRPM4. Single channel 22pS conductance with a linear current-voltage relationship was observed using the inside-out patch clamp configuration in the presence of intracellular Ca(2+). The channels were permeable to the monovalent cations Na(+) and K(+), but not to Ca(2+). The open probability was voltage-dependent, being higher at positive potentials. Using the whole-cell patch clamp "ruptured patch" configuration, the amplitude of the intracellular Ca(2+)-activated macroscopic current was dependent on time after patch rupture. Initial transient activation followed by a steady-increase reaching a plateau phase was observed. Biophysical analyses of the macroscopic current showed common properties with those from HEK293 cells stably transfected with human TRPM4b, with the exception of current time course and Ca(2+) sensitivity. The endogenous macroscopic current reached the plateau faster and required 61.9±3.5μM Ca(2+) to be half-maximally activated versus 84.2±1.5μM for the transfected current. The pharmacological properties, however, were similar in both conditions. One hundred μM of flufenamic acid and 9-phenanthrol strongly inhibited the endogenous current. Altogether, the data demonstrate the expression of endogenous TRMP4 channels in HEK293 cells. This observation should be taken into account when using this cell line to study TRPM4 or other types of Ca(2+)-activated channels.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > DBMR Forschung Mu35 > Forschungsgruppe Ionenkanalkrankheiten
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > DBMR Forschung Mu35 > Forschungsgruppe Ionenkanalkrankheiten

04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR)

UniBE Contributor:

Amarouch, Mohamed Yassine; Syam, Ninda Ratna Maharani and Abriel, Hugues

Subjects:

600 Technology > 610 Medicine & health

ISSN:

0304-3940

Publisher:

Elsevier

Language:

English

Submitter:

Verena de Serra Frazao-Bill

Date Deposited:

24 Jun 2014 14:43

Last Modified:

09 Jan 2015 05:10

Publisher DOI:

10.1016/j.neulet.2013.02.011

PubMed ID:

23428507

BORIS DOI:

10.7892/boris.43782

URI:

https://boris.unibe.ch/id/eprint/43782

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