Diuretic and smooth muscle-relaxing peptides, designated atrial natriuretic peptides (ANPs), have been identified in human and rat atrial tissues and implicated in the control of fluid volume and vascular function. Recently, cDNAs encoding the human and rat ANP precursors have been sequenced. We previously isolated from human tissue a natriuretic peptide of molecular weight (MW) 13,000 (gamma-hANP) comprising 126 amino acid residues, the largest natriuretic peptide so far identified, and showed that it is directly derived from the 151-residue human ANP precursor by the removal of a signal peptide. We now report the isolation and sequence analysis of a novel rat atrial natriuretic peptide (gamma-rANP) of MW 13,000, which derives from the rat ANP precursor. We also report the molecular cloning and nucleotide sequence analysis of the cDNA of the 152-residue rat ANP precursor, which is remarkably similar to the human 151-residue precursor (pre-hANP) except at the C-terminus. Differences in the rat and human precursor nucleotide sequences around the termination codons lead to a difference in processing pattern.