Adenosine A1 receptors were characterized in membranes from rat dorsal and ventral spinal cord using [3H] cyclohexyladenosine [( 3H]CHA) and compared with those in brain. For determination of anatomical loci of adenosine A1 receptors in the dorsal and ventral spinal cord, various lesions were employed, including kainic acid injections directly into the lumbar dorsal spinal cord, spinal cord hemitransections, dorsal rhizotomies, and neonatal capsaicin treatment. In control animals a single high affinity binding component was observed in dorsal and ventral spinal cord with KD values of 2.3 and 2.6 nM and Bmax values of 170 and 123 fmol/mg of protein, respectively. In comparison, [3H]CHA binding to whole brain membranes exhibited KD and Bmax values of 2.3 nM and 301 fmol/mg of protein, respectively. The IC50 values for CHA, (-)-phenylisopropyl adenosine, adenosine-5'-ethylcarboxamide, 2-chloroadenosine, (+)-phenylisopropyl adenosine, and theophylline to displace [3H]CHA were 3.6, 2.3, 15, 17, 21, and 30,500 nM for dorsal horn and 5.1, 2.7, 9.8, 24, 25, and 21,000 nM for ventral horn. The potencies of the various ligands are similar to those found for brain tissue. Injection of kainic acid directly into the dorsal spinal cord significantly reduced specific [3H]CHA binding by 33% in this tissue when compared to values from saline-injected control animals. This decrease was accompanied histologically by the depletion of intrinsic neuronal cell bodies and extensive gliosis at the injection site. Terminals of descending or primary afferent systems appear not to contain [3H]CHA-binding sites since lesions which interrupt these systems failed to alter the levels of [3H]CHA receptors in denervated spinal cord tissue.(ABSTRACT TRUNCATED AT 250 WORDS)