The incorporation of [14C]adenosine into various metabolites was studied in a hippocampal slice preparation in order to assess the extent of adenosine metabolism via synthesis of S-adenosylhomocysteine, a potent inhibitor of transmethylation reactions. Highest incorporation of 14C occurred into nucleotides, with only a few percent being recovered in inosine + hypoxanthine, S-adenosylhomocysteine, and the free adenosine pool. Labeling of S-adenosylhomocysteine did not significantly increase with higher concentrations of added adenosine despite greater accumulation of free [14C]adenosine in the tissue. Addition of L-homocysteine significantly increased the labelling of S-adenosylhomocysteine. The results indicate that S-adenosylhomocysteine synthesis is a minor pathway of adenosine metabolism in brain tissue under steady-state conditions. Further, changes in adenosine concentration, without a concomitant change in L-homocysteine availability, are unlikely to lead to a significant accumulation of S-adenosylhomocysteine. S-Adenosylhomocysteine is therefore not likely to play a significant role in mediating the biological effects of adenosine in the CNS via inhibition of transmethylations.