Interactions of chloroethylclonidine with rauwolscine‐ and prazosin‐sensitive adrenoceptors in dog saphenous vein
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abstract
α1‐Adrenoceptors have been classified pharmacologically into four subtypes (α1A, α1B, â1c and α1D) on the basis of their differential affinity for novel antagonists such as chloroethylclonidine (CEC). While CEC is considered an α1B‐adrenoceptor antagonist, our earlier studies revealed that it also acted like an agonist in the dog saphenous vein (DSV). The present study characterized the contraction induced by CEC in endothelium‐denuded rings from DSV.Concentration‐response curves for CEC were constructed in the absence (EC50 value of 11.13 ± 3.6 μm, n = 8) and presence of propranolol (β‐adrenoceptor antagonist, 30 nm), rauwolscine (α2‐adrenoceptor antagonist, 30 nm), prazosin (α1‐adrenoceptor antagonist, 30 nm) or methysergide (5HT2 antagonist, 30 nm) or both prazosin and rauwolscine. Pretreatment with methysergide (9.83 ± 5.14μm, n = 4) or propranolol (23.78 ± 12.32μm, n = 4) had no consistent effect. In the presence of rauwolscine, the concentration‐response curve for CEC was significantly shifted to the right with an EC50 value of 48.82 ± 13.2 JAM (n = 8). In the presence of prazosin, the CEC concentration‐response curve had an EC50 value of 29.12 ± 6.42 μm (n = 8). Pretreatment with both prazosin and rauwolscine shifted the concentration‐response curve for CEC to the right with an EC50 value of 72.67 ± 10.69 JAM (n = 8, P < 0.05). Maximum responses were significantly reduced only in tissues that were treated with both prazosin and rauwolscine.CEC (100 μm) pretreatment abolished prazosin binding sites and reduced the Bmax for rauwolscine by 50% without affecting the IQ value or the Hill slope.In Ca2+‐free Krebs solution containing 50 /IM EGTA, CEC produced a small transient contraction, suggesting that it can mobilize internally‐stored Ca2+. Pretreatment with rauwolscine abolished the CEC‐induced contraction in Ca2+‐free medium; prazosin pretreatment reduced but did not abolish CEC response in Ca2+“free medium.Restoring Ca2+ (0.5‐2.5 μm) to the extracellular solution increased CEC contraction in a concentration‐dependent manner, reaching a plateau at around 1.5 mm Ca2+. The contraction was insensitive to nicardipine (1 μm), a voltage‐operated Ca2+channel blocker, but was blocked in a concentration‐dependent manner by the putative receptor‐operated Ca2+channel blockers, SK&F 96365 (1–10 μm) and genistein, also a tyrosine kinase inhibitor (10–100μm).We conclude that CEC acts on rauwolscine‐ and, to a less extent, prazosin‐sensitive adrenoceptors in DSV to release internally stored Ca2+ and to open receptor‐operated Ca2+ channels. The inhibitory effect on CEC‐induced contraction that depended on external Ca2+ by genistein suggests a role for tyrosine kinase in the regulation of dihydropyridine‐insensitive Ca2+ entry.
