Functional and molecular expression of volume‐regulated chloride channels in canine vascular smooth muscle cells

摘要: We examined the possibility of functional and molecular expression volume-regulated Cl− channels in vascular smooth muscle using whole-cell patch-clamp technique quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) on cells from canine pulmonary renal arteries. Decreasing external osmolarity induced cell swelling, which was accompanied by activation Cl−-dependent outward-rectifying membrane currents with an anion permeability sequence SCN− > I− Br− aspartate−. These were sensitive to block DIDS, extracellular ATP antioestrogen compound tamoxifen. Experiments performed determine whether form chloride channel (ClC-3) is expressed arteries. Quantitative RT-PCR confirmed ClC-3 both types muscle. 76.4% β-actin artery 48.0% artery. We conclude that are exhibit properties similar those found other cells, presumably contributing regulation volume, electrical activity and, possibly, myogenic tone. Membrane stretch or increases transmural pressure cause contraction a response referred as tone (Meininger & Davis, 1992). This mechanical force changes conductance, cytoskeleton inactivation variety second messenger systems (Osol, 1995). Early studies (Sparks, 1964; Uchida Bohr, 1969; Harder, 1984) revealed associated depolarization exhibits some dependence Ca2+. Although modulation large-conductance Ca2+-activated K+ (Brayden Nelson, 1992) stretch-activated non-selective cation (Davis, Donovitz Hood, has been implicated response, exact mechanisms responsible for initial unknown. It recently shown pressure-induced cerebral inhibited various blockers, suggesting may be these arteries (Nelson, Conway, Knot Brayden, 1997). known ubiquitous (Large Wang, 1996), involved insensitive niflumic acid, effective blocker channels, leading speculation activated stretch, like volume many tissues (Strange, Emma Jackson, 1996). However, there presently no direct evidence showing functionally molecularly muscle. In tissues, possess common biophysical pharmacological properties, including hypotonic solutions, outward rectification symmetrical selectivity F−, sensitivity hypertonic stilbene compounds, nucleotides tamoxifen (Strange et al. 1996; Okada, three different proteins, P-glycoprotein (P-Gp; Valverde, Diaz, Sepulveda, Gill, Hyde Higgins, 1992), pIcln (Paulmichl, Li, Wickman, Ackerman, Peralta Clapham, ClC-2 (Grunder, Thiemann, Pusch Jentsch, have proposed candidates volume-sensitive recent data suggest P-Gp instead regulators endogenous fail characteristic (ICl,vol) native (Okada, identified another member ClC family, ClC-3, gene encoding ICl,vol heart mammalian (Duan, Winter, Cowley, Hume Horowitz, therefore tested isolated conventional techniques, examine ClC-3.