Characterization in rat aorta of the binding sites responsible for blockade of noradrenaline‐evoked calcium entry by nisoldipine

1 The effectiveness of the calcium antagonist, 1,4‐dihydropyridine nisoldipine, as an inhibitor of contraction and 45 Ca entry evoked by noradrenaline in rat aorta has been investigated and correlated with binding characteristics in intact artery. 2 Contractions evoked by noradrenaline were concentration‐dependently depressed by nisoldipine (0.3–300 n m ). About 60% of the response was resistant to inhibition, while KCl‐induced contractions could be completely blocked. Noradrenaline‐induced contractions were also less sensitive to nisoldipine inhibition than were KCl‐induced contractions. 3 Preincubation of the aorta with nisoldipine in high KCl depolarizing solution increased the inhibition of the contraction evoked by a short application of noradrenaline or KCl to a similar extent. 4 The inhibition by nisoldipine of 45 Ca influx evoked either by KCl depolarizing solution or by noradrenaline correlated well with the inhibition of the contractile responses. However, while KCl‐stimulated 45 Ca influx was totally abolished by nisoldipine (300 n m ), 38% of the noradrenaline‐stimulated 45 Ca influx was resistant to inhibition by nisoldipine (300 n m ). 5 The study of [ 3 H]‐(+)‐PN 200‐100 ([ 3 H]‐(+)‐isradipine) binding in intact aorta showed the presence of a homogeneous population of specific binding sites. K D values were dependent on the KCl concentration in the bath while B max was unaffected. Binding of [ 3 H]‐(+)‐isradipine was also increased in tissue exposed to noradrenaline; in the presence of 10 −5 m noradrenaline, binding parameters of [ 3 H]‐(+)‐isradipine were close to the values obtained in aorta bathed in 20 m m KCl solution. 6 Displacement of [ 3 H]‐(+)‐isradipine specific binding by nisoldipine was determined in segments of mesenteric artery and of aorta. The potency of nisoldipine was dependent on the incubation conditions applied to the vessel, as follows: KCl (100 m m ) depolarizing solution > noradrenaline (10 −5 m ) = KCl (25 m m ) solution > physiological solution. The K i value measured in aorta exposed to noradrenaline (10 −5 m ) was close to the IC 50 value of nisoldipine on the noradrenaline‐evoked contraction. 7 The membrane potential value of rat aorta was estimated by the distribution of [ 3 H]‐tetraphenylphosphonium bromide ([ 3 H]‐TPP + ). [ 3 H]‐TPP + uptake concentration‐dependently decreased when the KCl concentration in the bath was increased from 5.9 to 130 m m . Noradrenaline also concentration‐dependently decreased [ 3 H]‐TPP + uptake; the maximum effect (1–10 μ m noradrenaline) was comparable in amplitude to the effect of 25 m m KCl solution. 8 It is concluded that in rat aorta, noradrenaline activates voltage‐operated calcium channels that contain the specific, voltage‐sensitive binding sites for calcium antagonistic dihydropyridines. The existence of a fraction of noradrenaline‐stimulated 45 Ca entry that is resistant to nisoldipine blockade suggests that another Ca 2+ entry pathway is also opened by the agonist.