Rat arterial smooth muscle devoid of ryanodine receptor function: effects on cellular Ca2+ handling

1 The roles of intracellular Ca2+ stores and ryanodine (Ry) receptors for v ascular Ca2+ homeostasis and viability were investigated in rat tail arteri al segments kept in organ culture with Ry (10-100 muM) for up to 4 days. 2 Acute exposure to Ry or the non-deactivating ryanodine analogue C-10-O-eq glycyl ryanodine (10 muM) eliminated Ca2+ release responses to caffeine (2 0 mM) and noradrenaline (NA, 10 muM), whereas responses to NA, but not caff eine, gradually returned to normal within 4 days of exposure to Ry. 3 Ry receptor protein was detected on Western blots in arteries cultured ei ther with or without Ry. 4 Brief Ca2+ release events (sparks) were absent after culture with Ry, whe reas Ca2+ waves still occurred. The propagation velocity of waves was equal (similar to 19 mum s(-1)) in tissue cultured either with or without Ry. 5 Inhibition of Ca2+ accumulation into the sarcoplasmic reticulum (SR) by c ulture with caffeine (5 mM), cyclopiazonic acid or thapsigargin (both 10 mu M) decreased contractility due to Ca2+-induced cell damage. In contrast, cu lture with Ry did not affect contractility. 6 Removal of Ca2+ from the cytosol following a Ca2+ load was retarded after Ry culture. Thapsigargin reduced the rate of Ca2+ removal in control cultu red rings, but had no effect after Ry culture. 7 It is concluded that intracellular Ca2+ stores recover during chronic Ry treatment, while Ry receptors remain non-functional. Ry receptor activity i s required for Ca2+ sparks and for SR-dependent recovery from a Ca2+ load, but not for Ca2+ waves or basal Ca2+ homeostasis.