Previous investigations have shown disparate effects of adenine nucleotides on epidermal cell proliferation. Our present study demonstrates that adenosine and its related nucleotides (ATP, ADP, AMP) are antiproliferative for normal human epidermal keratinocytes cultured in the absence or presence of exogenous epidermal growth factor. Furthermore, the inhibitory effects of these compounds occur at concentrations less than 100 microM, are reversible, and do not affect the viability of the keratinocyte cultures. Our current investigation also demonstrates that both selective and nonselective adenosine receptor agonists are themselves approximately as potent as keratinocyte proliferation inhibitors, but are all less potent inhibitors than adenosine. These observations are consistent with the theory that adenosine mediates its antiproliferative response via a novel or more poorly characterized adenosine purinoreceptor subclass. Moreover, our present study demonstrates that ATP and ATP-gamma-S are significantly more potent antiproliferative agents than either alpha,beta-methylene ATP or beta,gamma-methylene ATP. Based on previous studies that have demonstrated that P2y purinoreceptors possess this type of ligand specificity and that the P2y purinoreceptor may be expressed by keratinocyte cultures, we propose that ATP may mediate its antiproliferative effects via this purinoreceptor. Collectively, our results indicate that adenosine and adenine nucleotides abrogate exogenous epidermal growth factor-dependent and -independent keratinocyte proliferation at submillimolar concentrations and may be important physiologic regulators of keratinocyte growth in vivo. Further, these results suggest that these or related compounds may have application as treatments for epidermal growth factor receptor-signaling pathway has been activated.