Triton Skinned Preparations

In triton skinned and in a-toxin-permeabilized smooth muscle preparations, the Ca + sensitivity of force production is also decreased by cGMP (Pfitzer et al., 1984,1986 Nishimura efflZ., 1992). This may be due to an up-regulation of MLCP (Pfitzer etal., 1986). Clear evidence that modulation of the activity of MLCP would affect Ca + sensitivity of tension development was in fact obtained in triton skinned smooth muscle when it was shown that inhibition of MLCP by the black sponge toxin, okadaic acid, increased Ca + sensitivity (Takai et al., 1987 Bialojan et al., 1988). On the other hand, incubation of triton skinned chicken gizzard fibers with a purified phosphatase decreased Ca + sensitivity (Bialojan et al., 1987). 

Ca + sensitivity of force production may also be associated with a shift in the relation between force and MLC phosphorylation (Jiang and Morgan, 1989), indicating that additional regulatory mechanisms may contribute to Ca2+-sensitivity modulation. 

The agonist-induced Ca + sensitization can be mimicked by GTPyS and inhibited by GTPpS (Fu-jiwara et al., 1989 Kitazawa et al., 1991a,b Satoh et al., 

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As pointed out by Butler et al. (1994), the a-toxin-permeabilized preparation used by Kitazawa et al. might contain endogenous phosphatase inhibitors that have been lost in the Triton-skinned preparation. This could at least partially explain the observed difference in phosphatase rate, and is an intriguing possibility relating to the demonstration that receptor agonists are able to increase Ca2+ sensitivity in neuroeffector-coupled permeabilized preparations by a mechanism involving inhibited myosin light chain phosphatase (MLCP) activity (Kitazawa et al., 1991 Kubota et al.,... 

Triton skinned and glycerinated fibers have been very valuable in demonstrating that phosphorylation and dephosphorylation of MLC is sufficient to induce contraction and relaxation (see Section III.B). These preparations have also been used to study the influence of ionic strength (Arheden et al., 1988 Gag-elmann and Guth, 1985), free Mg + (Arner, 1983 Bar-sotti et al., 1987), pH (Mrwa et al., 1974), inorganic phosphate (Schneider et al., 1981), nucleotides such as ATP and ADP (Arner and Hellstrand, 1985) on isometric force development, shortening velocity, and ATP turnover. Some of these experiments have also been carried out in smooth muscle fiber bundles and single smooth muscle cells permeabilized with saponin, (3-escin, or a-toxin (Saida and Nonomura, 1978 lino, 1981 Warshaw et al., 1987 Crichton et al., 1993). 

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