Localization of [ 3H]nitrobenzylthioinosine binding sites in rat spinal cord and primary afferent neurons

Abstract

The distribution of [ 3H]nitrobenzylthioinosine ([ 3H]NBI) binding to nucleoside transport sites in rat spinal cord and spinal roots was examined using membrane binding and autoradiographic techniques. A single class of high affinity binding sites having dissociation constants ( K D ) between0.42 ± 0.05 and0.088 ± 0.012 nM was observed in dorsal and ventral spinal cord and their associated roots. The maximal number of binding sites (B max) in dorsal and ventral spinal cord was110.1 ± 7.1 and73.6 ± 7.5fmol/mg protein, respectively. The highest levels of [ 3H]NBI binding were found in the dorsal grey matter of the cervical and lumbar enlargements. Autoradiographic studies showed that [ 3H]NBI sites were especially concentrated in the substantia gelatinosa of the dorsal spinal cord and the nucleus caudalis of the spinal trigeminal nucleus. The level of these binding sites in dorsal roots was nearly 4 times that observed in ventral roots; 98.5 and 23.0 fmol/mg protein, respectively. Adult animals depleted of unmyelinated sensory fibers by neonatal capsaicin treatment showed significantly reduced numbers of [ 3H]NBI sites (35%) in dorsal roots but not ventral roots, while K D values were unaffected. These results indicate that [ 3H]NBI sites are enriched in areas of the spinal cord and brainstem which subserve sensory functions and that these sites are located, in part, on unmyelinated primary afferent fibers.