Abstract

Nitric oxide has been shown to react under physiologic conditions with norepinephrine (NE) to produce 6-nitro-norepinephrine (6-NO₂-NE), a compound that enhances NE release in the brain. Previous studies suggest that 6-NO₂-NE is formed in the spinal cord and stimulates spinal NE release to produce analgesia. The purpose of the current studies was to examine the mechanisms by which 6-NO₂-NE stimulates NE release in the spinal cord. Crude synaptosomes were prepared from spinal cords of male Sprague-Dawley rats and loaded with [³H]NE. Incubation of synaptosomes with 6-NO₂-NE resulted in a release of NE, with a threshold of 1 μM 6-NO₂-NE and a maximum effect of 30% fractional release. NE transporter inhibitors desipramine and nomifensine blocked NE release from 6-NO₂-NE, and desipramine exhibited an IC₅₀ of 9.6 μM. NE release from 6-NO₂-NE was dependent on external Na⁺, but not Ca²⁺ or the activity of guanylate cyclase. 6-NO₂-NE also blocked uptake of [³H]NE into synaptosomes, with an IC₅₀ of 8.3 μM. These data are consistent with a direct action of 6-NO₂-NE on noradrenergic terminals in the spinal cord to release NE. This action is independent of guanylate cyclase activation, and most likely shares a common mechanism with classic monoamine releasers such as amphetamine that cause direct release of NE from vesicles into the nerve terminal cytoplasm, leading to extracellular release by reverse transport.