Ca concentration and flux in Ca-deprived arteries

1. Extracellular Ca and EDTA concentrations in arterial smooth muscle in Ca-free solutions with and without EDTA were estimated from (45)Ca and [(14)C]EDTA washout studies. Ca concentrations were calculated from the measured rate of (45)Ca washout at a given moment, combined with separate determinations of the coefficient of diffusion of Ca in the tissue, while EDTA concentrations were calculated by analysing an exponential phase of [(14)C]EDTA washout and checked by separate determinations of coefficient of diffusion of EDTA in the tissue.2. Extracellular Ca concentrations after 30 min in simple Ca-free saline at 36 degrees C was 0.049 mM at the centre of the tissue and averaged 0.033 mM throughout the tissue; these were well above the value of 0.005 mM-Ca that was found to be the threshold external level for contraction after tissue stores of Ca were removed.3. Extracellular EDTA concentration at the centre of the tissue reached 6.25 mM after 1.9 min in EDTA 12.5 mM at 36 degrees C or after 4.9 min at 5 degrees C. It reached 12.45 mM after 30 min at 5 degrees C, enough to keep free extracellular Ca at this point well below threshold even if all of the Ca in the tissue, determined as 0.42 mumole/g, were suddenly released on warming.4. Ca efflux increased greatly on warming after long periods in cold EDTA, and all measurable Ca left the tissue within 30 min in EDTA 12.5 mM at 36 degrees C although tissue Mg did not fall significantly during this time.5. Contractions elicited by noradrenaline in Ca-free saline, with or without EDTA present, were not associated with any increases in the rate of external loss of Ca greater than 0.001 mumole.g(-1).min(-1).6. Electronmicrographs showed numerous microvesicles that communicated with the extracellular space; also smooth endoplasmic reticulum among other structures that might have contained non-communicating Ca stores.7. The results provide evidence that large responses given by the arteries to noradrenaline in simple Ca-free saline were due to persisting extracellular Ca or to labile Ca stores dependent on this, while small responses obtained after long periods in cold EDTA depended on non-communicating Ca stores whose loss was too temperature-dependent to be limited by diffusion.