Effects of vanadate, phosphate and 2,3-butanedione monoxime (BDM) on skinned molluscan catch muscle

Abstract

The effects of orthovanadate (V(i)), inorganic phosphate (P(i)) and 2,3-butanedione monoxime (BDM) on tension, force transients and the catch state (passive tension maintenance) were investigated in saponin-skinned fibre bundles of the anterior byssus retractor muscle (ABRM) of the bivalve mollusc Mytilus edulis at pH 6.7. During maximal Ca(2+) activation isometric force was depressed by V(i) (0.03-10 mM), P(i) (10 mM) and BDM (50 mM). Force transients following quick stretches (0.1-0.3% of fibre length) were accelerated substantially by 1 mM V(i), 10 mM P(i) or 50 mM BDM. These compounds also accelerated force responses in experiments in which ATP was released rapidly from caged ATP by flash photolysis at both pCa 4.7 (force rise) and at pCa>8 (force decline). The effects on the catch state were investigated in two types of experiments: (1) Ca(2+) removal after maximal Ca(2+) activation and (2) rapid ATP release during high-force rigor at pCa>8. In both cases rapid relaxation was followed by slow relaxation (slower than 2% of initial force per min). This later slow relaxation (catch) was insensitive to V(i) (1-10 mM), P(i) (10 mM) and BDM (50 mM) but was accelerated by 0.12 mM cAMP. Complete relaxation to almost zero force was attained by changing pH from 6.7 to 7.7 (pCa>8). We conclude that catch depends on cAMP- and pH-sensitive structures linking the myofilaments and not on the force-generating actomyosin cross-bridges that are sensitive to V(i), P(i) and BDM.

Effect of pH on the rate of myosin head detachment in molluscan catch muscle: are myosin heads involved in the catch state?

“…The pH changes applied in the present study were larger, ranging stepwise from pH 6.7 (control) to 7.2, 7.4 and 7.7. The reason for the different pH sensitivity of intact and skinned ABRM preparations is not known; it was observed also by other investigators (Rüegg, 1971;Siegman et al, 1998;Galler et al, 2005).…”

“…This was confirmed in saponin-treated fibre bundles in the present study and in one of our previous studies (cf. fig.·1A in Galler et al, 2005). When catch is abolished by serotonin in intact ABRM, the pH changes from approximately pH 7.4 and pH 7.6 (Zange et al, 1990).…”

“…Briefly, ABRMs of fresh blue mussels (Mytilus edulis L.), obtained from a local sea-food supplier, were isolated (shell length, 4-7·cm 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (Hepes), pH 7.2]. These bundles were skinned in two different solutions containing 0.05% (w/v) saponin (Cornelius, 1980;Chick and Stephenson, 1995;Galler et al, 2005 KCl, 150·mmol·l -1 sucrose, 1·mmol·l -1 DTE, 0.05% (w/v) saponin, pH adjusted to 6.7 with KOH. The skinning procedure took about 30·min in total and was carried out at 20-24°C.…”

“…Therefore, most studies investigating catch have been carried out at pH 6.5-6.8 (e.g. Rüegg, 1971;Siegman et al, 1998;Galler et al, 2005). A change in pH from 6.2 to 7.5 (Rüegg, 1964), from 6.5 to 7.7 (Rüegg, 1971) or from 6.7 to 7.7 induces relaxation of catch force.…”

The N-terminal Region of Twitchin Binds Thick and Thin Contractile Filaments

“…Their results supported our previous suggestion (22) that twitchin might provide a mechanical link between thick and thin filaments. The idea that catch force is maintained by a mechanical link not involving the high force myosin cross-bridge is also supported by studies showing that agents that block force output by myosin cross-bridges have no effect when added to permeabilized muscles in catch (23,24) and actually increase catch force when added to activated muscles (24). Also, there is no measurable turnover of the ADP bound to myosin when catch force is relaxed by phosphorylation of twitchin (25).…”