Correlation between response to norepinephrine and removal of 45Ca from high-affinity binding sites by extracellular EDTA in rabbit aortic smooth muscle

Abstract

Divalent ion chelators (EDTA, EGTA) were found to remove 45Ca from high-affinity binding sites in isolated rabbit aortic smooth muscle in a concentration-dependent manner. Measurement of 14C-labeled EDTA uptake (10 to 60-min incubation periods) yielded tissue/medium ratios equivalent to the extracellular space [14C]-sucrose tissue/medium ratio). Addition of high EDTA concentrations (0.5--1.5 mM) elicits large but reversible increases in 45Ca efflux, rapidly removes virtually all 45Ca from muscles previously incubated with 45Ca for as short a time as 7 min (filling of extracellular and superficial sites) or as long as 180 min (more complete equilibration of 45Ca), and blocks the norepinephrine-induced contractile response (this response is only partially inhibited by 0.05 mM EDTA, a concentration too low to remove most of the 45Ca present). Furthermore, phosphatidyl serine, a compound known to decrease exchangeability of 45Ca in this tissue, inhibits the EDTA-induced increase in 45Ca loss. Thus, those slowly depleted Ca++ stores that are important for the contractile action of norepinephrine are removed by EDTA even though this chelator is confined to the extracellular space. Possibly, release or removal of high-affinity Ca++ is regulated by EDTA-accessible Ca++ bound at relatively superficial membrane sites.