Estimation of high K- and noradrenaline-induced 45Ca uptakes in isolated rat aorta: effects of washing in icecold solutions.

Isolated rat aorta rings, labelled for 10 min with 45Ca in control physiological saline solution (PSS), PSS with 10 microM noradrenaline (NA), or 80 mM potassium-PSS (80 K), were washed out into non-labelled, icecold PSS. NA and 80 K increased tissue 45Ca. Retention of stimulated 45Ca uptake during washout was studied by subtracting the control washout curve from the washout curves of the stimulated tissues. The measured stimulated uptake fell rapidly during the initial 15 min of washout, thereafter decreasing more slowly. These results suggested that a lengthy wash in icecold PSS would lead to an underestimate of stimulated cellular 45Ca uptake. The cellular location of the stimulated uptake was confirmed by using diltiazem (10 microM) and forskolin (10 microM) to block specifically the 45Ca influxes caused by 80 K and NA, respectively. A similar biphasic loss of stimulated 45Ca uptake was seen in icecold lanthanum (50 mM) solution. These data suggest that established methods for measuring cellular Ca in isolated blood vessels may have markedly underestimated the amount of Ca entering cells during stimulation.