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High pressure liquid chromatography with electrochemical detection was used to quantify the efflux, in the same sample, of endogenous 5-hydroxytryptamine (5-HT), noradrenaline (NA), and 5-hydroxyindoleacetic acid (5-HIAA) into superfusates of the rat spinal cord in vivo. The efflux of these three agents was measured prior to, and during, electrical stimulation of the nucleus raphe magnus (n.r.m.) and nucleus reticularis paragigantocellularis (n.r.p.g.), two medullary nuclei implicated in antinociception. In untreated rats, basal efflux of 5-HT and NA was 0.21 and 0.12 ng/ml of superfusate respectively; the basal efflux of 5-HIAA was 18.17 ng/ml. Stimulation of the n.r.m. and n.r.p.g. in these animals increased the efflux of 5-HT and 5-HIAA, but did not alter the efflux of NA. 60 min after administration of fluoxetine (10 mg/kg, I.P.), a 5-HT uptake inhibitor, basal efflux of 5-HT and NA was unaltered, but the basal efflux of 5-HIAA was decreased. In these rats, stimulation of the n.r.m. and n.r.p.g. increased the efflux of 5-HT and of NA. The efflux of 5-HIAA was not altered. In rats pre-treated with both fluoxetine and desipramine (10 mg/kg, I.P.), the basal efflux of NA was increased while that of 5-HIAA was decreased; the basal efflux of 5-HT was not affected. The efflux of NA, but not of 5-HT, was increased in these animals during stimulation of the n.r.m. and n.r.p.g. The efflux of 5-HIAA was not changed by stimulation. Addition of fluoxetine alone or with desipramine to the superfusate in high concentrations greatly increased basal efflux of 5-HT. Failure of stimulation of the ventromedial medulla to increase the efflux of 5-HT in these animals may be related to feed-back inhibition of release by the high concentration of 5-HT initially present in the superfusate. These results indicate that electrical stimulation of the n.r.m. and n.r.p.g. increases the efflux of endogenous 5-HT and NA from the spinal cord. These stimulation sites are coincident with brain-stem sites at which stimulation produces antinociception by activation of spinal serotonergic and noradrenergic receptors. Thus, the ability of stimulation at these sites to evoke the spinal release of the probable neurotransmitters further supports the hypothesis that the antinociceptive effect is mediated by activation of serotonergic and noradrenergic neurones projecting to the spinal cord.

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