Protection of presynaptic α-adrenoceptors against irreversible blockade by phenoxybenzamine: preservation of the modulatory effects of exogenous noradrenaline and yohimbine

Summary

1. Receptor protection experiments were carried out in order to study the site of action of α-adrenoceptor agonists and antagonists on the release of noradrenaline. Cerebrocortical slices from rabbits were preincubated with ³H-noradrenaline. They were then superfused with medium containing cocaine 30 μmol/l and stimulated electrically (3 Hz) three times, after 60, 250 and 295 min of superfusion (S₁, S₂, S₃). Phenoxybenzamine 10 μmol/1 when used, was added between S₁ and S₂ for 30 min; putative protecting drugs (clonidine 100 μmol/1 or yohimbine 10 μmol/1) were present 5 min before and during the exposure to phenoxybenzamine and then washed out together with the latter. Either the voltage drop between the electrodes at S₂ and S₃ or the Ca²⁺-concentration of the superfusion medium at S₂ and S₃ was diminished, if necessary, in order to bring the overflow evoked by S₂ close to the overflow at S₁. Blockade by phenoxybenzamine, or protection against the blockade, was examined by addition of the test compounds noradrenaline 0.1 μmol/1 or yohimbine 1 μmol/1 before S₃. 2. In slices not exposed previously to α-adrenoceptor ligands, noradrenaline 0.1μmol/1 greatly reduced, whereas yohimbine 1 μmol/1 greatly increased the evoked overflow of tritium. Both effects were abolished in slices treated with phenoxybenzamine 10 pmol/1 alone between S₁ and S₂. 3. In contrast to phenoxybenzamine alone, exposure to phenoxybenzamine 10 μmol/1 in the presence of either clonidine 100 pmol/1 or yohimbine 10 μmol/1 failed to abolish the effects of the test compounds noradrenaline 0.1 μmol/1 and yohimbine 1 μmol/1, although the effects were reduced. 4. It is concluded that the irreversible antagonist phenoxybenzamine, the protecting agents clonidine and yohimbine, the test compounds noradrenaline and yohimbine, and by inference endogenous noradrenaline as well, all act at the same site, namely the presynaptic α-autoreceptor.