PERSISTENT ACTIVATION BY AND RECEPTOR RESERVE FOR AN IRREVERSIBLE A(1)-ADENOSINE RECEPTOR AGONIST IN DDT1MF-2 CELLS AND IN GUINEA-PIG HEART

The p- and m-isothiocyanate adenosine derivatives yl]methyl]anilino]ca rbonyl]methyl]phenyl]adenosine (p- and m-DITC-ADAC) were examined for irreversible agonist effects at the A(1)-adenosine receptor (A(1)-AdoR ) in DDT1 MF-2 (DDT) cells and a functional A(1)-AdoR response in the guinea pig isolated heart. The p- and m-DITC-ADAC inhibited (-)-isopro terenol stimulated cAMP accumulation in DDT cells in the low nanomolar range, and the maximal responses elicited by both compounds were simi lar to that for N-6-cyclopentyladenosine. Once established, the p-DITC -ADAC-mediated inhibition of cAMP accumulation in DDT cells was not af fected by the addition of the AdoR antagonist 8-cyclopentyl-1,3-diprop ylxanthine (CPX). Pretreatment of DDT cells with p-DITC-ADAC (1 mu M), followed by washing, reduced [H-3]CPX binding to the A(1)-AdoR by 44% without altering the K-d value for the radioligand to the remaining r eceptors. The relationship between irreversible A(1)-AdoR occupancy by p-DITC-ADAC and inhibition of cAMP accumulation revealed a relatively large receptor reserve (64%) for the maximal response. In guinea pig isolated hearts, m-DITC-ADAC (5 mu M) prolonged the stimulus to His bu ndle (SH) interval by 2.1-fold; this response could be prevented by th e antagonist 8-cyclopentyltheophylline (5 mu M). However, after the SH interval prolongation was established, extensive washout or the addit ion of 8-cyclopentyltheophylline had little reversal effect on the m-D ITC-ADAC response. Binding of [H-3]CPX to the guinea pig ventricular m embranes after m-DITC-ADAC treatment and washing was reduced by 35%. T he A(1)-AdoR occupancy response relationship for m-DITC-ADAC to prolon g the SH interval indicated a small (10-20%) receptor reserve. Both p- and m-DITC-ADAC seem to be irreversible full agonists at the A(1)-Ado R and may prove to be useful probes to further investigate A(1)-AdoR s tructure-function relationships.