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Comparison of the properties of CLCA1 generated currents and I_Cl(Ca) in murine portal vein smooth muscle cells

Fiona C. Britton, Susumu Ohya, Burton Horowitz and Iain A. Greenwood

Calcium-activated chloride currents (I_Cl(Ca)) have been recorded in various smooth muscle cells but, to date, there has been no information as to the molecular nature of the channel underlying this conductance. We have characterised native I_Cl(Ca) in freshly dispersed smooth muscle cells isolated from murine portal vein using whole-cell voltage clamp. I_Cl(Ca) exhibited time-dependent activation at depolarised potentials and rapid deactivation upon repolarisation. The reversal potential of I_Cl(Ca) was close to the theoretical equilibrium potential (E_Cl) and was shifted by replacement of external Cl^- by SCN^- or isethionate. Dithiothreitol (DTT, 1 mM), a blocker of CLCA1, had no effect on the I_Cl(Ca) current in myocytes. RT-PCR demonstrated the expression of mCLCA1 transcripts, but not mCLCA3 transcripts, in various murine smooth muscle cells including portal vein, as well as cardiomyocytes, and the levels of mCLCA1 transcriptional expression were quantified by real time quantitative RT-PCR. Stable transfection of HEK293 cells with the cDNA encoding mCLCA1 cloned from murine portal vein smooth muscle yielded a current with notable differences in Ca²⁺ sensitivity, channel kinetics and modulation by DTT from the native I_Cl(Ca). However, there was some similarity in the pore properties and these data suggest that mCLCA1 alone does not comprise the Cl^- channel in portal vein smooth muscle cells.

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