Structural determinants of phorbol ester binding in synaptosomes: pharmacokinetics and pharmacodynamics

The present study used structurally distinct phorbol esters to investigate the relationship between their pharmacokinetics of binding to protein kinas e C (PKC) in rat brain cortex synaptosomes, their affinity for PKC in synap tosomes and ability to enhance noradrenaline release from rat brain cortex. Affinity binding studies using [H-3]phorbol 12,13-dibutyrate (PDB) yielded a rank order of potency for the phorbol esters in binding to synaptosomal PKC of phorbol 12-myristate-13-acetate (PMA)> deoxyphorbol 13-tetradecanoat e (dPT)= PDB much greater than 12-deoxyphorbol 13-acetate (dPA)= phorbol 12 ,13-diacetate (PDA). In intact synaptosomes PDB, dPA and PDA rapidly displa ced bound [H-3]PDB whereas PMA and dPT were comparatively slow. However, th e displacement rates for all the phorbol esters were equally rapid in synap tosomal membranes or synaptosomes permeabilised with Staphylococcus alpha-t oxin. These results suggest that the lipophilic phorbol esters (dPT and PMA ) are slower to displace [H-3]PDB binding because they are hindered by the plasma membrane. In brain cortex slices it was found that the rate of displ acement of [H-3]PDB binding was closely correlated with the degree of eleva tion of transmitter noradrenaline release. Thus kinetic characteristics may determine biological responses and this may be particularly evident in eve nts which occur rapidly or where there is fast counter-regulation. (C) 1999 Elsevier Science B.V. All rights reserved.